The Changing Market Structure For The American Beef Industry by Harlan Hughes1 I. Introduction

Change is now a common everyday word in the beef marketing. Change spells out “Cattlemen Have A New Great Encore.” You and I are witnessing this encore as the beef industry divides into two distinctly different marketing systems based on two distinctly different production systems. First, we have the traditional beef system producing and marketing commodity beef. Second, we have a value-based beef system producing and marketing high quality beef designed to meet tight consumer specifications. A critical decision that today’s beef cow producers have to make is to decide which system – commodity beef or value-based beef – they are going to target their production towards. The beef industry’s transition into value-based marketing is generating rapid change in today’s beef industry. Today’s beef industry has roughly 800,000 beef cow producers marketing to 2100 feedlots; who in turn, are marketing to 4 major beef packers who are marketing to 250 million plus domestic consumers.2 In spite of the wide spread belief that agriculture in general, and the beef industry in particular, are different from other U.S. industries, today’s beef industry is also caught up in the U.S. economy’s overall drive for efficiency. Consumers are demanding value in anything and everything they purchase. He who can produce the beef that consumers want, in the form that they want it, when they want it, at the lowest price wins.

1Harlan Hughes is Livestock Economist and Professor Emeritus, North Dakota State University. Paper presented at Western Animal Science Symposium entitled “Using Science And Marketing In Producing And Delivering Value Added Products For The Consumer” Bozeman, Montana June 20, 2001.

2Topper Thorp, CattleFax, NCBA Annual Meeting, San Antonio, Texas, Feb 2001.

1 A. Commodity Beef By definition, commodity beef is a homogenous product with little or no product differentiation and there are no brand names associated with commodity beef. Commodity beef is priced based on averages. Average beef prices are public information reported daily by the USDA Agricultural Marketing Service. Since one beef producer’s commodity beef is about the same as another beef producer’s commodity beef, the primary way that commodity beef producers compete is through being a lower-cost producer. Traditionally, research and extension educational programs have targeted their research and educational programs towards increased production efficiency of commodity beef. The operational assumption has been that the increased production efficiency implies lower costs of production, which in turn, implies higher profits. Research, however, is changing. Using this conference as a specific example, current research and extension programs are changing to a focus on the production and marketing of consumer determined value-added production. Change is everywhere. Commodity beef producers are trying to lower costs of production by adopting new genetics, new production practices, and new management tools that will increase their individual herd’s production efficiency. Producers generally perceive that production efficiency is a sufficient condition for profitability (economic efficiency). This certainly has been true in past decades; but, once again, things are changing. IRM (integrated Resource Management) data suggests that production efficiency is a necessary condition but production efficiency is not now a sufficient condition to guarantee economic efficiency. Since the mid 1980s, financial management has increased in importance in beef production. In today’s profitable beef production, economic efficiency now also requires financial efficiency.3 I find that some beef producers have made this transition to financial

3I have had senior ranchers talk to me about why the current generation “can not survive on the ranch that momma and I lived on for 40 years.” What these senior ranchers have not recognized is the change that has occurred in the ranching business. For the last 30 years, profit margins have decreased with each and every added cattle cycle. This drop in profit margins has been heavily related to the 30 year decrease in beef demand. I really think that most ranchers have no concept of what 30 years of decreasing beef demand had done to them and to their neighbors. The good news, today, is that demand is increasing (see section in this paper on changing demand).

2 efficiency very well while others are still struggling. The traditional business model followed by most commodity beef producers is to produce commodity beef during those time periods that best fit their production resources. There is little or no focus in the typical commodity business model on year-around supply, seasonal production patterns, or seasonal market price patterns. Consumers are expected to consume whatever is the easiest for beef producers to produce; in fact, consumer demand is ignored in the commodity beef business model. The rest of the marketing chain is considered the enemy. The enemy’s fight with ever decreasing beef demand is frequently interpreted as the enemy attempting to take advantage of beef cow producers. If prices are low, it must be the fault of the next participant in the marketing chain. Commodity beef producers typically are going it alone and often view their neighbor as their competitors. The general practice in the commodity beef business model is to not share any production or financial performance data with other participants in the marketing chain and there even is a general reluctance towards sharing with fellow beef cow producers. The management power of sharing and benchmarking to increase beef cow profits is not possible due to non- standardized accounting systems and the reluctance to share.4 A. Change Came Out Of The Financial Crisis Of The 1980s Change came out of the agricultural financial crisis in the 1980s when many beef producers found out that high production, in itself, did not guarantee financial survival. In addition, most beef producers were let down by their on-farm, production and financial management information systems; in fact, bankers were called upon to determine when a business failed. By that time all equity had been used up.5

4Out of North Dakota’s 12,000 operations with beef cattle, 250 participate in CHAPS Herd Performance System, share data, and benchmark against each other. Another 180 participated in the North Dakota Farm Business Management System, share data, and benchmark against each other. The other 11,570 do not share data and can not practice benchmarking.

5A favorable observation in the 1990s is that some farmers and ranchers decided to get out of farming while there was still some equity left in the business. In the 1980s, if you had equity, you farmed or ranched one more year. There is nothing wrong with selling the business while you still have equity but you need the financial records to know how much equity you have and how fast it is changing.

3 In the late 1980s, a group of NCBA cattlemen came to the Cooperative Extension Service and requested help in designing a new management information system for commodity beef producers that integrated financial management and production management into a single management information system. This new management information system was called IRM -- Integrated Resource Management. IRM was conceived and driven by grassroots NCBA cattlemen who witnessed neighbors forced out of business during the financial crisis of the 1980s. These grassroots producers wanted to make sure that their ranch did not succumb to the next financial crisis in the 1990s. Over the last 10-12 years a completely new IRM educational program has been launched by State Cooperative Extension Services, NCBA, and State Cattlemen’s Associations to change how ranchers and beef farmers manage their commodity beef production. NCBA launched its Cattlemen’s Colleges, State Cattlemen Associations launched their Cattlemen’s Colleges, and State Extension Services launched intensive IRM educational programs all based around one common set of Standardized Performance Analysis (SPA) Guidelines. Ranchers were taught the IRM way of integrating financial management into the production management of their beef cow businesses. Since IRM’s conception, almost every state in the Nation has launched an IRM educational program for their beef producers. Profits can and are being made from producing commodity beef. In fact, low-cost commodity beef producers are currently making good profits. My IRM work identified the key Critical Control Points (CCP) for running a profitable beef cow herd is intensified management. My Northern Plains IRM Database clearly indicates that the economic rewards from commodity beef production through intensified management are going to those that have the lowest unit costs of producing a hundredweight of calf (see Figure 1). For example, the average net income per cow for all IRM Cooperators in 1999 was $129 per cow. My highest profit herd in 1999 generated $281 net returns Figure 1: Unit Costs Of Production to unpaid labor, management, and equity capital per For Northern Plains IRM Database

4 cow with his 1999 calves. My low-cost IRM Cooperators have made a profit each and every year of the cattle cycle. Yes, IRM is changing the economics of producing commodity beef. B. Value-Based Beef Value-based beef, the newer production and marketing system, is being marketed based on selected quality factors targeted towards consumers’ expressed wants and desires. Value-based prices are based on a specified set of quality specifications implemented through some form of a grid pricing system. The largest example of value-based beef is Certified Angus Beef (CAB).6 CAB is one of the oldest (founded in 1978) and perhaps the best-known of all beef carcass alliances.7 Today there are many alliances being offered to beef producers. As early as 1996, Kansas State University (Sartwelle, Marston and Bolze) listed and described 13 alliances in a paper.8 Each different value-based marketing program focuses on a different set of quality specifications. New value-based marketing programs are starting up every year. The challenge for today’s value-based beef cow producers is to match their herd’s genetic characteristics to a specific value- based marketing program. CattleFax reported that 40-50 percent of the beef produced is now or soon will be marketed on some contract, grid, or formula price structure.9 While costs of production are still important in value-based beef production, value-based beef producers spend considerable management energy focusing on adding value to their beef production. Value-based beef producers are recognizing that consumers have specific quality specifications for what they consume and that consumers will reward quality through paying a

6I once mentioned to an Angus breeder that CAB was really taking off. His response was: “Well, it is about time! We have been working on this for over 20 years.” The message is that developing an effective alliance takes years.

7 James Sartwell, Twig Marson, and Ron Bolze, Jr. “Marketing And Informational Alliances In The Fed Cattle Sector.”, paper presented at AAEA-LMIC Commodity Outlook Conference, San Antonio, Texas, July 26, 1996.

8See previous footnote.

9Topper Thorp. Presentation at NCBA annual Conference, San Antonio, Texas Feb 2001.

5 premium for that quality product. On-the-other-hand, consumers will pay a discount through reduced consumption of those products that do not meet consumers’ quality specifications . In addition, consumers demand a year-around supply of that quality product leading producers to the recognition that producers have to band together in business relationships to meet year-around consumer demand. Value-based beef is characterized by the fact that value is linked to the production practices at each and every level of the supply chain. As a result, a value-based marketing system has to focus on all producers in the supply chain. The marketing challenge is to make sure that the marketing program recognizes these linkages equitably. The marketing system needs to recognizes that what one producer does in the supply chain directly impacts other producers in that supply chain. Fellow participants in the total value-based beef supply chain are business partners -- not their competitors. Retail value is the collective sum of all “value added” actions taken in the total supply chain. How the complete value-based beef supply chain is managed determines the overall marketing success of any value-based beef marketing program. Value-based beef producers need to carefully select their value-based marketing program with care as not all value-based beef marketing programs will survive. The logical next step in value-based beef marketing will be the move to branded beef; but, that is a subject of another complete paper. Key Factors Underpinning Change In The Market Structure

Key production, economic, and financial factors are underpinning the change in the beef marketing system. My goal in this section of this paper is to briefly discuss 6 of the key underpinnings. The six key underpinnings causing change are: 1. Cattle cycle and its resulting beef price cycles 2. Economic hurt in the mid-1990s 3. Increasing production per cow 4. Long-term decreasing demand for beef 5. The good news on the recent up-turn in beef demand 6. Ranchers are currently marketing on an up-market

1. Cattle Cycle And Its Resulting Beef Price Cycle

6 The single most important economic underpinning of the beef marketing system is the cattle cycle and its resulting beef price cycle. Beef cattle prices tend to go in 10-year cycles that correspond to each decade. History demonstrates that we can expect a “U” or “V” beef price cycle during each decade. Cattle prices typically start the decade with high prices, move to low prices in the mid-decade, and increase again towards the end of the decade. Figure 2 illustrates the “V” shaped price Figure 2: US Calf, Feeder, and Slaughter Prices In 1990s. cycle generated in the decade of the 1990s. Price spreads between feeder calves, feeder cattle, and slaughter cattle started out the decade wide but narrowed in the mid-1990s. The magnitude of the price depression in 1996 is best illustrated by the fact that feeder calf prices per hundred weight went below the per hundredweight price of slaughter steers.10 Since 1996, the price spreads have been widening. This a the typical price pattern during a typical cattle cycle. Price spreads start out the decade wide, narrow in mid- decade, and widen again toward the end of the decade. Where, then, are we in the current cattle cycle? a. Current Cattle Cycle implications Cattle cycles can be divided into three phases – contraction, expansion and turn-around. We have completed the 1996 to 2000 contraction phase, now have completed the turn-around phase and are just now entering into the expansion phase. The expansion phase should last through 2003.

10For 22 years and two complete cattle cycles I had talked about 1974 being a unique time in history when calf prices per hundred weight averaged below slaughter cattle prices per hundred weight. Twenty-two years and two cattle cycles later it happened again. In 1996, the average price of feeder calves went below the average price of slaughter cattle. Both record lows – the 1974 and 1996 lows – were accentuated by record feed grain prices.

7 Beef production peaked in year 2000 and is projected to decrease in 2001-2003 time period. As the increased heifer retention materializes in the expansion phase, beef production should start increasing once again in year 2004. This cycle’s peak beef prices are currently projected for year 2003. The next downturn in market prices is projected for 2004

through 2007. Prices are projected to increase Figure 3: North Dakota Long-Run towards the end of the decade (see Figure 3). Planning Prices For 500-600 Lb Steer Calves

As beef cow producers decide to divert heifers from feeding to breeding, the supply of feeder cattle will be reduced. The expansion phase is characterized by a smaller supply of feeder cattle, excess feedlot capacity, excess packer capacity and by overall industry consolidations.11 CattleFax reports a 15 percent increase in feedlot capacity in the last 15 years.12 Feedlots, trying to utilize their excess feedlot capacity, will increase their competitive bid for feeder cattle. Existing feedlots will turn to added imported Mexican feeder cattle to try and utilize existing feedlot capacity. Due to higher feeder cattle prices, some feedlots will feed cattle to minimize feedlot losses rather than to maximize profits. All of this suggests that the beef industry will see the cow calf producer move into the drivers seat with respect to feeder cattle sales. They will decide when they want to market their calf production -- at weaning, after backgrounding, or through retained ownership. As more beef cow producers sell at weaning, the popularity of retained ownership will decline. Reduced retained ownership will lead to more cattle owned directly by the feedlots. Feedlots, trying to ensure a feeder cattle supply for their feedlot, will strive to develop

11Topper Thorp, CattleFax at NCBA Annual meeting, San Antonio, Texas Feb 2001.

12Dave Weber, CattleFax presentation to NCBA Annual Conference, San Antonio, Texas, Feb 2001.

8 and expand business relationships directly with beef cow producers. Reduced feeder cattle supplies will change how beef cow producers interact with cattle feeders. 2. The Economic Hurt During The Last Cattle Cycle The economic impact of the mid-1990's price depression was devastating for many beef cow producers. Figure 4 presents the average profit per cow earned by North Dakota’s Farm Business Management Association Members during the1990s.13 Profit per cow was high in the early part of the decade averaging $152 to $192 per cow. Nineteen-hundred-and-ninety-four signaled the beginning of the market downturn with a 74 percent drop in beef cow profits in that one year. Average profits went negative in 1995 and went even more negative in 1996. Average profits again Figure 4 Average North Dakota’s Beef turned positive in 1997, 1998 and 1999. Profits Cow Profits During The 1990s continued increasing in 2000 to $ 124 per cow. Clearly, better times are returning for cow calf producers. An important point for this conference is that we recognized the fact that over the five- year period – 1994 thru 1998 – producers earned an average annual profit of only $3 per cow for this five year period. It appears that profit margins are getting smaller for each progressive cattle cycle. A signal that beef cow profit margins are getting smaller and smaller is the fact that this 5-year $3 average in the decade of the 1990s compares to the 5-year average low of a positive $33 for the decade of the 1980s. In fact, none of the 1980 years averaged below zero while two years in the 1990s averaged below zero. Clearly, something was rasing havoc with the beef

13Profit here is defined as the earned returns to unpaid family and operator labor, management, and equity capital. These are the three resources that a ranch family contributes to the ranch business.

9 industry! Research now suggests that something was decreasing consumer demand for beef. More on this in a later section of this paper. 3. Increased Beef Produced Per Cow While the cattle cycle’s influence on cattle numbers is a primary factor determining beef supply, it certainly does not explain the total beef supply picture. Figure 5 illustrates the USDA All-Cattle Inventory numbers from 1960 through year 2000 via the dashed line measured on the right axis. You can see that cattle numbers peaked in the mid-1970s, again in the mid-1980s, Figure 5: Beef Production and All-Cattle Inventory 1960 - 2000. and once again in the mid-1990s. You also see that the long run trend in cattle numbers from 1975 through year 2000 has been downward. The dark line in Figure 5 presents beef production measured on the left axis. While beef production varies during each cattle cycle, the long-term trend in beef production since 1965 has been upward. I particularly call your attention to the 1980 through year 2000 time period. While the all-inventory numbers trended downward over this 20-year period, beef production trended upward over this same time Figure 6:Beef Produced Per Cow 1960-2000. period. In fact, beef production in 1998 exceeded the record beef production in 1975. How could this be? The beef industry has 34

10 million less cattle in year 2000 as compared to 1975? One key reason for the upward trending beef production from less and less cattle is “increasing beef production per cow.” Over the last 30 years, beef production per cow has trended upward (see Figure 6). In 1980, each cow produced approximately 450 pounds of carcass beef per year. By 2000, production had risen to 620 pounds of carcass beef per cow per year. Figure 7: North Dakota’s Increase In Weaning This 170 pound increase in a 20 year time Weights During 1980s. span represents an average annual productivity gain of 1.8 percent.14 Several factors are responsible for this production increase including weaning weights. Figure 7 confirms that in the decade of the 1980s, North Dakota beef cow producers added an average of 10 pounds per calf weaned per year. This totals to 100 pounds added to weaning weights in the decade of the 1980s. I fully believe that this North Dakota increase in weaning weights is indicative of the U.S. beef industry in general. Canada, on the other hand, shifted to a higher percentage exotic breeding programs and, as a result, increased weaning weights even more than the U.S. Figure 8: Carcass Weight Increased Through Time. Certainly, genetic improvements in both countries have resulted in bigger and bigger

14Ibid.

11 carcass weights making carcass weight a major component of the increased beef production per cow. Adding 100 pounds to carcass weight results in an additional 3.6 billion pounds of beef.15 Another primary source of productivity gains is heavier carcass weights (see Figure 8). Over the past 20 years, while annual productivity gains have averaged 1.8 percent, weight gain has accounted for 1.1 percent of the total 1.8 percent growth per year. All other productivity increases have accounted for about 0.7 percent per year.16 These other productivity increases involve the ability to produce more calves from a given number of brood cows and include fewer non-productive cow days and higher calving rates. Better husbandry practices have certainly contributed to this higher beef cow production as have improved health programs. The popularity of slaughtering veal calves has decreased so today the beef industry is taking a higher percentage of all calves to slaughter weights. Finally, imports have also contributed to this increased production (see Figure 9). Cattle imports make up of all types of cattle – breeding, feeders, and slaughter. Most Mexican exports into the U.S. are feeder cattle while Canadian exports to the U.S. are processed meats, slaughter cattle, cull cows, and feeder cattle. In any case, U.S. imports add to the total beef supply produced in the U.S. While the amount of beef produced per cow is rising at a rate of about 1.8 percent per year, domestic and foreign demand is only increasing at about 1.3 Figure 9: Live Cattle Imports From Canada and percent per year. This suggests that Mexico.

15Assumed 36 million head slaughtered annually. 36 million times 100 pound increase comes to 3.6 billions added pounds of beef. Carcass weights have continued to increase for the last 30 years.

16Ibid.

12 productivity growth will likely outpace demand growth, and as a consequence, brood cow numbers over the long-run will likely decrease over time.17 Current beef cow numbers have also been dropping since 1995 and yet total beef production has continued to rise. This short-run increase in beef production primarily due again to short-run increases in carcass weights, increased feeder cattle imports, and lower veal calf slaughter.

4. Decreasing Demand For Beef Figure 10: Nominal Steer Prices 1949- 1995. During the 1980s beef consumption in North America declined, in part, due to concerns about human health and grazing pressures on public lands. At the same time, poultry and fish enjoyed increases in consumption due to lower prices and alleged health advantages. With real incomes also declining in the 1980s, consumers became more cost conscious and began to pay more attention to the quality of meat obtained per dollar spent. Decreasing demand for beef led to continual decrease in the real price of beef . While nominal steer prices trended upward since the early 1960s (see Figure 10), inflation drove the real price of slaughter steers ever downward (Figure 11). By 1995, the real steer price was one-half of the real steer price in 1979. Figure 11: Real (Inflation Adjusted) Slaughter Seer Prices 1949-1995. Most cattlemen do not appreciate the impact that 25 years of decreasing demand for beef and the continued drop in the real price of beef was

17Kris Hurt, “Cycle Nears End Of Contraction” Livestock Outlook Web Page, http://web.aces.uiuc.edu/farm.doc/marketing/livestockoutlook/0700cattle_text.html.

13 having on the ranching industry. Over the last 25 years, ranch after ranch has been forced out of business by decreasing demand. Let’s look at some ranching changes driven by decreasing demand. A. Producers Turn To Exotics – Consumers Heighten Demand For Quality Information Generating the family living draw from beef cows was one of the more challenging implications of reduced real prices of beef. To remain in business, beef cow producers had to increase production efficiency in an attempt to maintain family living draw during these times of reducing real market prices. Increased production efficiency became about the only way for a ranch business to survive. Consolidation of smaller beef ranches into larger, more efficient ranches was one way to reduce family living draw per cow. Ranch after ranch was sold and consolidated into a neighboring ranch operation. Each ranch consolidation meant one less family’s living draw had to be generated from beef cows. Producers looked to the introduction of Continental European Breeds (exotics) for significant increases in size and cutability as a means of increasing production efficiency. The genetic drive of the 1970s and 1980s was to increase weaning weights, carcass weights and to produce beef more efficiently. Figures 7 illustrates the dramatic increases in weaning weights and Figure 8 illustrates the dramatic increase in carcass weights. Carcass weights are still trending upward. B. More Select Beef and Less Choice Beef This focus on increasing weaning weights and the larger carcass weights led to the rise in popularity of producing USDA Select quality grade beef. In 1986 USDA , under pressure from the cattlemen, changed the name of existing beef quality grade from Good to Select. The amount of beef graded Select has increased markedly since that time. For example, in 1986 only 1.3 million pounds per month of Select beef were graded as compared to 30.2 million pounds per month in 1999.18 During this time period, the production of Choice beef has remained relatively stable at around 46 million pounds per month.

18Jayson L. Lusk and Thomas L. Marsh, “Wholesale Demand For USDA Quality Graded Boxed Beef And Effects On Seasonality.” Selected paper, Western Agricultural Economics Association Annual Meeting, Vancouver, British Columbia, Jun 29-July 1, 2000.

14 In a market of increasing differentiation, USDA beef quality grades played an important role in distributing quality throughout the marketing chain and should have provided signals to cattle producers of consumer desires at the retail level. Increased use of the USDA beef quality grading system reflected heightened consumer demand for quality information and segregation at the retail level. According to USDA data, over 90 percent of beef from steer and heifer slaughter was quality graded in 1999 as compared to just 67 percent in 1986. Consumers wanted quality information and quality segregation at the market. C. Beef Demand Research Tells More Of The Story Although the amount of beef graded has increased markedly, not much is known about the price sensitivity of the USDA beef quality grades or the substitutability between grades and other meat products. Recently published research has shed some added light on beef demand.19 For example, chicken is only a substitute for Select beef, not Choice beef. Thus, chicken may be substituted for low quality, but not high quality beef. Research shows that the two beef quality grades are good substitutes for one another during the fall and winter; however, during the summer Select beef is not a substitute for Choice beef. Apparently, retailer demand for Choice beef cannot be met by changes in relative prices of lower quality beef during the cookout season. In addition, demand for both Choice and Select beef becomes much more inelastic ( % change in quantity demanded verses % change in price is less than 1.0) during the spring and summer than during the fall and winter. Increases in retail beef prices increases the amount of wholesale pork and poultry demanded by meat retailers. Increases in retail beef prices have as much, if not more, of an impact on wholesale pork and chicken demand than equivalent changes in wholesale pork and poultry prices. A previously unreported result, presented in the above study, is the cross price elasticity between Choice and Select beef. Results indicate that a 1 percent increase in the price of Choice beef is associated with a 0.28 percent increase in the quantity of Select beef demanded. On-the-other-hand a 1 percent increase in the price of Select is associated with a 0.19 percent increase in the quantity of Choice beef demanded by meat retailers. Price changes in Choice

19Ibid.

15 have a larger impact on the quantity of Select demanded than the reverse. The same pattern of beef demand may apply to producers marketing fed cattle. If these same seasonal fluctuations in beef demand are transmitted to the farm level, cattle producers, as a whole, may be able to benefit from timing the feeding of cattle. Feeding cattle to heavier weights during the summer and fall to achieve the Choice quality grade may prove to be a profitable strategy. However, if the seasonal pattern of beef demand is not transmitted to the farm level, packers may be capturing this economic surplus. Packers may be able to take advantage of changes in retailers’ demand for quality graded beef by increasing its margins through strategic seasonal cattle purchasing. D. Market Is Demanding More Choice Cattle And The Beef Industry Is Producing More Select

It appears that today’s beef industry is currently living with the genetics brought on board in the 1970s and 1980s. During the 1990s, the percent of the carcasses grading Choice went down (see Figure 12) while the percent of the carcasses grading Select went up. The net result of all of this is that the market is demanding more Figure 12: Percent Cattle Grading Choice & Select 1993 Thru Mid 2000. choice cattle and producers are producing more select cattle. This is further demonstrated by looking at Kansas State University’s Choice/Select Price Spread Index (Figure 13). A large index suggest a larger Choice/Select price spread. The Choice/Select price spread has followed a distinct pattern. The Choice/Select prices index is low during the first four months of the year. By May, the index gets large and stays large the

16 rest of the year peaking in November.20 The price spread tends to be widest in the fall months. This Kansas research suggests that during 8 out of the 12 months the premium for Choice cattle is large. Once again, the market is signaling that it wants Choice cattle. The Choice-Select price spread averaged $15.09 per hundredweight in October 1999. This Choice/Select price spread is the Figure 13: KSU Choice/Select Index. fundamental source of premiums paid for value-based beef. D. National Beef Quality Audit The 1995 National Beef Quality Audit (NBQA) suggested that current USDA quality grading system segregates carcasses into: 1 percent Prime, 47 percent Choice (11 percent upper 2/3 Choice and 36 percent Low Choice), 47 percent Select, and 5 percent Standard. 21 Slaughter quality in the 1995 Audit quality actually decreased from the earlier 1991 NQBA. Based on USDA 1974 data, the percent cattle grading prime and choice decreased 36 percent from 1975 to 1995. Decreasing demand since 1975 has paralleled this beef quality decrease since 1975. The 2000 National Beef Quality Audit showed some improvement over the 1995 audit. The 2000 Survey showed that U.S. cattle producers have responded to the market place, delivering higher quality product to consumers then they did in the mid-1990s.22 The percentage of Choice and Prime carcasses went from 48 percent in 1995 to 51 percent in 2000. Prime percentages went from 1.3 percent to 2 percent in 2000.23 Clearly, the beef industry is moving in the right direction but progress is slow.

20Get this footnote from ??

21Kenneth Eng, “Conference Focuses On Optimizing Performance, Improving Feeding Efficiency,” Page 16, Feedstuffs, April 15, 1996.

22 Page 1. “NBQA 2000 Gaining Ground,” NCBA, 2001.

23Ibid.

17 Retailers indicate that 50 percent of the beef marketed through their stores must be Select to meet consumer demands. Exporters, on-the-other-hand, say they need 30 percent prime, 42 percent upper 2/3 Choice and 28 percent low Choice with no Select to satisfy their customers. Food service representatives need another product mix different yet from retailers and exporters. To adequately meet the needs of all customers, the U.S. beef industry should produce cattle grading 7 percent prime, 21 percent upper 2/3 Choice, 34 percent low Choice, and 38 percent Select carcasses.24 Ranchers will increase market segmentation and targeting will be necessary to efficiently match production resources to consumer demand specifications. E. Increased Demand For Value-Based Beef Increased demand for quality graded beef at the retail level has resulted in beef packers offering cattle producers premiums and discounts based upon the quality and yield grades of their cattle. With the current flurry of marketing alliances, it behooves alliance participants to understand how the market’s increasing demand for Choice beef is in direct conflict with the past genetic trend in the beef industry. This all suggests that, as the quantity of Choice slaughter cattle goes down, price premiums for Choice cattle will go upward. In the mean time, meeting consumer demand for Choice beef necessitates the importation of Choice beef and cattle from Canada. 5. The Good News Is That Demand May Have Changed Upward The most exciting news impacting the beef industry in years is the recent change in beef demand. Kansas State University researchers have documented 1) the magnitude in the drop in demand during the 1990s and 2) the apparent turn around in demand towards the end of the decade Figure 14: Demand Turned Around in 1999.

24Warren Kester, “Consistency’s The Quest,” Pg 46-47, Beef, November 1996.

18 (see Figure 14).25 Figure 14 illustrates the decrease in demand during the 1990s where 1990 was set as the base of 100. By 1997, retail beef price was down to 83 percent of what it would have been if demand had stayed at the 1990 level. This represents a 13 percent drop in demand over the 7-year 1990 to 1997 period. Beef cutout demand went down 30 percent over this same 7-year period and live slaughter steer demand also went down 30 percent over this same 7- year period. All three demand values went down even more in 1998. The apparent turnaround in demand is illustrated in the 1999 data Figure 15: Demand Increase Signal Has Been and preliminary year 2000 data as both Received. years generated increases in all three values (again see Figure 15). By June 2000, the retail beef index went up to 91, beef cutout was up to 83 and the live steer price index went up to 76. While two years do not make a trend, this apparent increase in demand is the best news that the beef industry has received in many, many years. Increasing demand changes the dynamics of marketing beef. 6. Ranchers Are Now Marketing Beef Cattle On An Up Market Figure 16 presents the opening slide that I Figure 16: Introduction Slide In Recent have used over the past year in my beef producer Presentations

25Source: Jim Mintert’s Kansas State University Monthly Beef Newletter.

19 conferences and meetings. Note the champagne bottle on the slide. Also, note the fist-of- money on the other slide of Figure 16. Why would I begin my beef presentations with a bottle of champagne and a fist-full of money? After showing the bottle of champagne and the fist-of- money to last year’s audiences, I have asked the question: “ARE YOU READY TO MAKE SOME MONEY??” As this point in my introduction, I then draw the conference participants attention to the relative profitability of beef by asking: “What other commodity group could I stand in front of in year 2001 and ask if they are ready to make some money?” It is interesting to see the response that this discussion gets from beef producers. Ten to twenty percent respond by vigorously shaking their head yes and immediately echoing “You bet we are ready to make some money!” The rest are either in shock that I would ask such a question or in complete disbelief of the whole situation. I certainly would not do this in front of wheat producers. I assure you that I would not do it with corn producers. I would not do it with swine producers nor dairy producers. My whole point with all of this is that the beef cow industry is currently positioned in the cattle cycle to have several very favorable years. Let me summarize all of this by stating that the beef business goes in 10-year cycles and we are in the upward phase of the current beef price cycle. Figure 17 presents my current planning prices for North Dakota beef cow producers. These are October’s average monthly prices for 500-600 pound steer calves marketed in Western North Dakota. The left-hand side presents historical prices from 1990 through year 2000. The right-hand side presents my projected planning prices for years 2001 through 2009. These projections suggest that we will see steer calf prices trend upward for the next 2-plus years with the projected price peak in year 2003.

Let’s take a more in-depth look at short-run Figure 17: North Dakota Calf Planning feeder calf planning prices. The Extension Prices 1990 thru 2008.

20 Economics Section, Department of Agribusiness and Applied Economics, North Dakota State University, posts weekly my Futures based planning prices generated from the weight/price spreads of last week’s feeder cattle auction sales and the Futures market.26 Figure 18, prepared from May 23, 2001 Western North Dakota salebarn prices and May 30, 2001 feeder cattle Futures prices, compares October 2001 projections with historical prices for three previous Octobers. Prices are presented for steer calves weights ranging from 400 to 925 pound feeders. Two price trends are evident in Figure 18. First, feeder steer prices for all feeder weights have trended upward from 1998 through 2000 and are projected to continue this trend into the Fall 2001. Second, cheap feed grains, and the resulting low-cost gains, are driving the price of light-weight feeder calves up relatively more than the price for heavier-weight feeders. This is evident by the clock-wise rotation of the price lines in Figure 18. Figure 18 Western North Dakota Feeder In summary, this section has highlighted Calf Prices for Oct 1998-2001 one more change for ranchers. Ranchers are now marketing beef cattle on an up market. Summary Change is now a common everyday word in beef marketing and change is the central theme of this paper. Today’s beef industry is also caught up in the U.S. economy’s overall drive for efficiency. Consumers are demanding value in everything they purchase and beef purchases are no exception. Changes in beef marketing are in the works. The beef industry is changing into two distinctly different marketing systems based on two distinct production systems – one production system for commodity beef and one production system for value-based beef. By definition, commodity beef is a homogenous product with little or no product differentiation. Without product differentiation, commodity beef producers

26: The web page is www.ag.ndsu.nodak.edu/cow under the “weekly prices” hot button.

21 compete by being the lowest cost producer. In order to increase economic efficiency, reduce production costs and increase overall beef cow profits, the beef industry has developed and implemented the Integrated Resource Management(IRM) and the Standardized Performance Analysis (SPA) Systems. The key focus of IRM is on integrating financial management and production management into a single management information system that sends integrated management signals for enhancing beef cow profits. Today’s leading IRM cooperators are generating profits of over $200 per cow producing commodity beef. Value-based beef is the newer beef marketing system and is being marketed and priced based on selected quality factors targeted toward consumers’ expressed wants and desires. Value-based beef producers are recognizing that consumers have specific quality specifications for what they consume and consumers are willing to pay a premium price for that quality product. Consumers demand for year around supply necessitates value-based producers banding together to meet this year around demand. Certified Angus Beef is the best known example of value-based beef alliance. Today, there are many difference beef alliances and new ones are forming each year. The logical next step in value-based marketing will be branded beef. There are six fundamental factors underpinning the change in the current beef marketing system. These six factors are: 1. Cattle cycle and its resulting beef price cycles 2. Economic hurt in the mid-1980s 3. Increasing production per cow 4. Decreasing demand for beef 5. The good news is that consumer demand has turned up 6. Ranchers are marketing on an up-market

The single most important factor impacting beef marketing is the cattle cycle and it resulting beef price cycle. Beef prices go in 10-year cycles that correspond to the decades. Market prices are typically high in the beginning and ending years of the decade. Produces can expect the cycle low in beef prices to occur in the mid-part of each decade. Farm Business Management data also suggests that beef cow profit margins are getting smaller and smaller with each progressing cattle cycle.

22 Change is again impacting feeder cattle marketings. As beef cow producers now decide to divert heifers from feeding to breeding, the supply of feeder cattle will decrease. This reduction in feeder cattle supply over the next 2 years will put beef cow producers in the drivers seat with respect to feeder cattle sales. Over the past 30 years, beef cow numbers have trended downward and total beef production has trended upward. Beef production per cow has increased over the years due to increased carcass weights, increased feeder cattle imports, and reduced veal calf slaughter. Beef production per cow has been increasing at such a rapid pace that lagging beef demand has led to reduced beef cow numbers. While the amount of beef per cow is rising at a rate of 1.8 percent per year, domestic and foreign demand is only increasing at about 1.3 percent per year. This means that the production increases per cow will continue to decrease brood cow numbers over time. Long-term decreasing demand for beef led to a long-term decrease in the real price of beef. While nominal steer prices trended upward since the early 1960s, inflation has driven the real (inflation adjusted price) ever downward. By 1995, the real steer price was only one-half of the real steer price in 1979. To remain competitive during times of decreasing demand, beef producers have had to increase production efficiency. This was the only way that they could even begin to maintain the family living draw from the ranch in times of reducing real market prices. Over the last 25 years, ranch after ranch has been forced out business by decreasing demand. This pressure for generating family living led to the general consolidation of smaller ranches into larger, more efficient ranches as a way of reducing family living draw per cow. Producers feeling continued pressures to increase production efficiency, looked to the exotic breeds for significant increases in size and cutability as another means of increasing production efficiency. This industry wide focus on increasing weaning weights lead to the rise in the production of USDA Select grade beef. At the same time, meat retailers turned to increased the use of USDA beef quality grading systems in response to the heightened consumer demand for quality information and segregation at the retail level. It appears that today’s beef industry is currently living with its focus on weaning weights

23 in the 1970s and 1980s. During the 1990s, the percent of the carcasses grading Choice went down while the percent of the carcasses grading Select went up. The net result of all of this is that the market is demanding more Choice cattle and producers are producing more Select cattle. Quality price premiums are now being established in the market. Increased demand for quality beef at the retail level has resulted in beef packers offering value-based cattle producers premiums based upon the quality and yield grades of their cattle. Today, meeting the demand for Choice cattle includes importing Choice cattle from Canada. Change spells out “Cattlemen Have A New Great Encore” and the most exciting encore is that beef demand may have turned upward for the first time in 25 years. This is the best news that the beef industry has received in many, many years. Increasing demand changes the dynamics of beef marketing. Ranchers are once again marketing in an up-market. Part of this up-market is due to the cattle cycle and part of this up-market is due to increased demand. Yes, change is here -- “Cattlemen should Have A New Great Encore.”

24 Preweaning survival in swine1

D. C. Lay, Jr.*2, R. L. Matteri†, J. A. Carroll†, T. J. Fangman‡, and T. J. Safranski§

*Livestock Behavior Research Unit, ARS, USDA, Purdue University, West Lafayette, IN 47907 and †Animal Physiology Research Unit, ARS, USDA, ‡College of Veterinary Medicine, and §Department of Animal Sciences, University of Missouri, Columbia 65211

ABSTRACT: A limited ability to cope with environ- intake, and disease. Causes of mortality may be more mental stressors (cold, disease, limited nutrition), par- closely linked with one another than previously be- ticularly over the first 2 to 3 d of life, predisposes the lieved. Interactions exist between disease, thermoregu- piglet to relatively high rates of neonatal morbidity and lation, and nutrition. Piglets with disease and nutri- mortality. Due to the serious economic impact, numer- tional problems experience hypothermia and express altered behaviors that increase the likelihood of their ous surveys of preweaning losses have been conducted being laid on by the sow. High probabilities of neonatal over the last century. Although losses are still signifi- losses are associated with low birth weights, cold ambi- cant, the existing literature indicates a significant im- ent temperatures, and scouring. An understanding of provement in piglet survival over time, as determined the interactions between environmental stressors and by reports of 35% preweaning mortality in 1924 and 13 the biology of the piglet forms the basis for strategies to 15% in 2000. Major sources of mortality have been and recommendations for improving preweaning categorized as overlying by the sow, insufficient energy survival.

Key Words: Mortality, Piglets, Preweaning Period, Survival

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E74–E86

Introduction Within the United States, of the preweaning losses 48.1% are due to the sow’s lying on the piglets, 15.3% A survey of 712 swine farms, representative of 95% are due to starvation, and 13.3% are due to scours of U.S. producers, was conducted for the first time by the (NAHMS, 1995). Other reviews of preweaning mortal- National Animal Health Monitoring System, USDA- ity are in general agreement with regard to major cate- APHIS-VS in the year 1990 (NAHMS, 1991) and again gories of crushing, starvation, and disease (Svendsen in 1995 (representing 90% of producers, NAHMS, et al., 1986; Varley, 1995). Reflecting the economic im- 1995). These surveys estimated a national average of portance of neonatal survival, numerous surveys of pig- 9.9 piglets born alive per litter and 8.4 piglets weaned let losses have been conducted over the last century per litter, representing a 15% preweaning mortality for (Zavoral, 1924; Hutchinson et al., 1954; Koketsu, 2000). the year 1990, and they estimated a national average In 1928, average preweaning mortality was reported of 9.5 piglets born alive per litter and 8.6 piglets weaned at 35% of all liveborn piglets (Zavoral, 1924). Losses in per litter, representing a 9% preweaning mortality rate excess of 30% are still reported in some less-developed for the year 1995. According to survey results, piglets countries (Chabo et al., 2000). These numbers should be born alive per litter has decreased during the 5 yr be- viewed carefully, though, because many factors affect tween surveys but piglets weaned has increased, effec- what is reported. tively decreasing preweaning mortality. The percent- age of farms using total confinement during farrowing Overview did not change, remaining constant at 81%. Economic Impact

1Mention of trade names or commercial products in this publication The value of saving additional piglets can be dra- is solely for the purpose of providing specific information and does matic. The National Swine Improvement Federation not imply recommendation or endorsement by the U.S. Department estimates that each additional piglet born has a value of Agriculture. of $13.50 (NSIF, 1997). The value of saving piglets could 2Correspondence: 216 Poultry Sci. Bldg. (E-mail: layd@ purdue.edu). be calculated using this value assuming there is no Received April 5, 2001. added value prior to weaning. Until recently there was Accepted October 29, 2001. not a market for weaned piglets, and little effort was

E74 Preweaning survival in swine E75 placed on determining the value of a weaned piglet. during the early postnatal period, because continuous With the move to segregated-early-weaning, however, cross-fostering later in the preweaning period can de- it has become necessary to determine the value of pig- crease overall growth (Straw et al., 1998). lets at less than 21 d of age. Dhuyvetter (1996) has Dystocia, and the resulting oxygen deprivation, has estimated that a weaned piglet has a value of $33.00. long been associated with stillbirths (Jackson, 1975). Some of the substandard piglets that typically die are Hypoxia during farrowing is an important contributing not this valuable, even though they cost that much factor to the 70 to 90% of stillbirths when death occurs to produce. Therefore, estimating the value of saving during the farrowing process (English and Morrison, additional piglets using this figure may be overinflated. 1984). Some decrease in blood flow to the fetus is com- According to USDA statistics (USDA, 2000) there mon during normal uterine contractions at farrowing. were 11,462,000 litters farrowed in the United States However, more serious reductions can occur through in 2000. The average litter size weaned was 8.84 piglets. damage or occlusion of the umbilicus, or placental de- An assumed 9% preweaning mortality suggests an av- tachment. The resulting hypoxia can result in still- erage of 9.71 liveborn piglets, and a loss of 0.87 piglets births and reduced postnatal viability. Piglets born in per litter. The total value of preweaning loss in 2000 the latter part of the birth order experience a greater can then be calculated either conservatively or liberally degree of hypoxia. Hypoxia increases the amount of as follows: time between birth and first suckling and is associated with hypothermia, reduced postnatal growth, and 11,462,000 litters × 0.87 piglets per litter higher neonatal mortality (Herpin et al., 1996). Minor × $13.50 per piglet = $134,621,190 11,462,000 litters direct effects of hypoxia on thermoregulatory ability × 0.87 piglets per litter × $33.00 per piglet may exist (Herpin et al., 1999); however, the major = $329,074,020 detrimental effects are likely related to depressed colos- trum intake and carbohydrate metabolism (Herpin et Accepting these values as lower and upper limits sug- al., 1996). gests that the annual cost of preweaning mortality to Large litters result in a longer farrowing duration the U.S. swine industry is between $130 and $330 and thus may be critical to survival for piglets born million. toward the end of farrowing. Litter size can also influ- Many of the underlying causes of neonatal mortality ence piglet survival after birth; research has shown are interlinked. For example, neonatal losses due to that piglet losses are greater in large litters (Fahmy and cold stress are rarely recorded, although the resulting Bernard, 1971; Dyck and Swierstra, 1987; Marchant et hypothermia may lead to starvation, crushing, and(or) al., 2000). These losses are largely attributed to the disease (Curtis, 1974; Kelley, 1985; English, 1993). within-litter variation in piglet body weight (Marchant Weaker piglets that are not able to effectively compete et al., 2000). Pigs born to and reared in large litters for colostrum and milk become hypothermic (Svendsen have lower preweaning survival rates and depressed et al., 1986). As hypothermia and lack of nutrition fur- preweaning gain and are older at 105 kg than counter- ther weaken the piglet, problems with orientation and parts born to and reared in smaller litters (Stewart and locomotion occur (DeRoth and Downie, 1976; Svendsen Diekman, 1989). et al., 1986), which increase the probability of crushing. Preweaning mortality is influenced by birth weight, Prolonged Farrowing litter size, duration of farrowing, dystocia, birth order, thermal environment, nutritional status, disease, sow Dystocia, or difficult farrowing, in sows is generally and piglet behavior, sex, and genetics. the result of uterine and maternal exhaustion. Primary uterine inertia associated with a deficiency in the con- Piglet Size and the Birth Process tractile potential of the myometrium is uncommon in the sow (Britt et al., 1999). Secondary uterine inertia The low-birth-weight piglet is particularly at risk for is not a cause of the dystocia, but rather is a result of preweaning morbidity and mortality. Piglets weighing the dystocia associated with fetal malpresentation or less than 0.8 kg (1.8 lb) at birth have a 32% survival maternal obstruction (Britt et al., 1999). rate, compared with 97% for piglets weighing 2 kg (4.4 Pigs born after an extended labor are more likely to lb) or more (Gardner et al., 1989). The low-birth-weight experience a higher death loss associated with hypoxia piglet is physiologically compromised in terms of energy (Randle, 1971). In the early 1970s it was reported that stores and susceptibility to cold and is at a disadvantage an 11.8% perinatal mortality rate was observed in lit- in competing with larger littermates at the udder. ters farrowed in less than 6 h, compared to a 21.3% Within-litter variability in piglet weights is known to perinatal mortality rate in those litters farrowed over be associated with higher preweaning losses (English more than 6 h (Bille et al., 1974a,b). Housing conditions et al., 1982; Marchant et al., 2000). In some instances, during gestation, litter size, season, and disturbances cross-fostering to create more uniformity in piglet from stockpersons influence overall duration of the ex- weights has been shown to reduce mortality by 40% pulsion phase of parturition in sows (Naaktgeboren, (English et al., 1982). Cross-fostering should be utilized 1979; Svendsen and Bengtsson, 1983). E76 Lay et al. Thermal Environment the early neonatal piglet does not increase its intake in response to cold temperature. Colostrum intake actu- Cold stress is a critical factor affecting piglet survival ally decreases during cold exposure, exacerbating the over the first several days of life (Curtis, 1970, 1974; likelihood of starvation (Le Dividich and Noblet, 1981). Kelley, 1985). Cold exposure begins at birth with a rapid Behavioral adaptations also play an important role 15 to 20°C transition from the uterine environment to in maintaining body temperature. Neonatal piglets are ambient temperature conditions. The newborn piglet able to choose microenvironments that reduce exposure has a relatively high thermoneutral zone (TNZ), which to cold temperatures. Such behavior has been demon- is defined by upper critical and lower critical tempera- strated in laboratory settings (Balsbaugh et al., 1986); tures (UCT and LCT, respectively). Ambient tempera- however, piglets still prefer to huddle close to the sow tures higher than the UCT and lower than the LCT and littermates during the first 3 d of life regardless of activate the animal’s thermoregulatory mechanisms in thermal environment or the presence of supplemental order to maintain core body temperature within normal heat (Hrupka et al., 1998). Proximity to the sow is a limits. Due to the high UCT, heat stress is rarely a concern with regard to increasing the risk of piglet problem for the newborn piglet; however, a LCT of crushing (Wechsler and Hegglin, 1997); therefore, the 34.6°C highlights the concern raised by cooler tempera- design of microenvironments that provide warmth and tures. However, the piglet’s ability to tolerate cooler keep the piglets at a safer distance is of considerable temperatures increases rapidly with age. Between 24 interest. It is important to keep in mind that this strat- and 48 h of age, the LCT decreases from 33.3 to 30.1°C egy must be balanced with keeping the piglets by the (Berthon et al., 1993), and by 1 wk the LCT of the piglet dam’s side during the first 24 h to ensure that they has rapidly declined to approximately 25°C (Svendsen ingest enough colostrum to maintain their health. In- et al., 1986). terestingly, neonatal piglets show a significant reduc- In addition to thermal environment, physiological tion in huddling behavior during the acute-phase re- factors also play an important role in the thermoregula- sponse to an immunological challenge (Matteri et al., tory ability of the neonate. Brown fat, for example, is 1999). Consistently, piglets with scours prefer cooler an important source of metabolic heat production in ambient temperatures when allowed to choose locations many neonates (Benito, 1985). An important peculiar- within an experimental thermocline (Balsbaugh et ity in the thermoregulatory ability of the piglet relative al., 1986). to other species stems from the absence of brown fat Piglet size is also an important factor associated with (Trayhurn et al., 1989). Uncoupled oxidative metabo- thermoregulation in the newborn. The low-birth-weight lism of brown fat mitochondria can generate a sustained piglet is particularly at risk for hypothermia, with a thermogenic response that can constitute one-third of greater body surface-to-volume ratio and reduced en- ergy stores (English and Morrison, 1984). The rate of the overall metabolic rate in neonatal animals (Foster, body temperature loss during cold exposure is inversely 1984). The newborn piglet, however, is able to mount related to piglet weight; however, Meishan piglets are other thermoregulatory responses (Stombaugh et al., better able to maintain body temperature than Large 1973). Exposure to cold elicits piloerection, although Whites on a body-weight basis (Le Dividich et al., 1991). the piglet’s hair coat limits the effectiveness of this The increased cold resistance of the Meishan piglet is heat-conserving mechanism. Other physiological fac- not related to fat stores; the well-known difference in tors such as reduced blood flow to the periphery, shiv- body fat composition between Meishan and European ering, and increasing metabolic rate also aid in heat breeds develops after the neonatal period. The colos- production. Shivering is an important heat-generating trum of the Meishan sow, however, has a high fat con- mechanism in the neonatal piglet (Noblet et al., 1997). tent that can provide her piglets with the energy needed Although not highly efficient thermogenically, shiv- to maintain body temperature (Le Dividich et al., 1991). ering in piglets is associated with increased body tem- Sow feed intake is likely a greater concern at high perature and metabolic rate (Herpin and Le Dividich, ambient temperatures for piglet survival. Increasing 1995; Berthon et al., 1996; Lossec et al., 1998). ambient temperature has been shown to decrease in- The newborn piglet increases its metabolic rate lin- take in lactating sows, with a suppression of piglet early as environmental temperatures decrease from ap- growth, but not survival, at temperatures above 25°C proximately 34 to 18°C. At 18°C, metabolic rate reaches (Quiniou and Noblet, 1999). In fact, a slight increase an upper limit (summit metabolic rate). Summit meta- in piglet survival was observed at 27°C. The ambient bolic rates in piglets of 36.6, 43.3, and 46.8 kJ/h/kg have temperatures used to produce heat stress in the sows been reported at 2, 24, and 48 h of age, respectively were closer to the TNZ of the piglets, which would be (Berthon et al., 1993). At these higher metabolic rates, expected to decrease hypothermia and metabolic energy the importance of adequate intake becomes critical. requirements, allowing the piglets to perform well at Body heat production is linearly related to colostrum the expense of their dam. intake (Le Dividich and Noblet, 1983). At the summit metabolic rate, the newborn piglet utilizes its energy Nutritional Status stores in 11 to 12 h without nutritional intake (Herpin The piglet is born with relative physiological defi- and Le Dividich, 1995). In contrast to older animals, ciencies that predispose it to mortality and morbidity. Preweaning survival in swine E77 The newborn piglet has a low energy reserve, approxi- and starvation likely reflect a variety of etiologies. Pig- mately half that of lambs and calves (Mellor and Cock- let losses tend to be the result of noninfectious causes burn, 1986; Herpin and Le Dividich, 1995). Adequate and are strongly associated with management prac- ingestion of colostrum is crucial for neonatal survival tices. However, epidemics of certain neonatal diseases (Noblet et al., 1997). Accordingly, factors related to lim- can occur and may result in extremely high levels of ited intake and(or) increased energy expenditure have mortality for limited periods of time (Vaillancourt et a serious impact on survival (NAHMS, 1991). Body fat al., 1994). Accordingly, the immunological development is only 1 to 2% of body weight at birth, with most lipids of the neonatal piglet has been extensively studied. In existing in cell membrane structure (Mersmann, 1974; all cases it is important to maintain high levels of sani- Svendsen et al., 1986). The ability of the newborn piglet tation and biosecurity (see Amass and Clark for re- to oxidize fatty acids is extremely low (Mersmann and view, 1999). Phinney, 1973). Accordingly, the early neonatal piglet The piglet is immunologically immature at birth and relies heavily on carbohydrate stores, with glycogen depends on early postnatal transfer of maternal anti- levels in liver and muscle of approximately 200 and bodies from colostrum for immune protection. Colos- 120 mg/g tissue weight, respectively (Mersmann, 1974). trum is a rich source of immunoglobulin-G (IgG), which Liver and muscle glycogen stores reach minimum levels provides the piglet with circulating immunity against at 12 to 18 and 36 to 48 h of age, respectively. The disease organisms. Immunoglobulins are maximally piglets’ respiratory quotient usually decreases by 12 h, absorbed from the jejunum into lymphatic vessels reflecting an increased utilization of dietary fat (Mers- within the first 12 h of life. The ability of the immuno- mann, 1974). globulins to pass through the gut declines thereafter. Increasing levels of colostral fat have a positive linear By 48 h of age, no further transfer occurs (gut closure). effect on the piglet’s energy intake and fat deposition, However, IgG have a relatively long half-life of 14 d, although total colostral intake is somewhat reduced (Le which provides some immunological overlap with the Dividich et al., 1991). Increasing dietary fat for the sow in late gestation and lactation can significantly increase onset of endogenous antibody production at approxi- the fat content of colostrum and improve survival of low- mately 10 d of age (Wilson, 1974; Gaskins and Kelley, birth-weight piglets (Pettigrew, 1981; England, 1986). 1995). Antigen-stimulated lymphocyte proliferation in- This becomes especially important in induced far- creases and T-cell differentiation markers appear and rowings when parturition occurs 1 to 2 d prior to natural increase during the first weeks of life (Becker and Mis- farrowing (Jackson et al., 1995). The use of long- and feldt, 1993). As colostrum production ends and milk medium-chain triglycerides (LCT and MCT, respec- production begins, the primary antibody transferred to tively) in sow feeds is also of interest, because the con- the piglet is IgA (60% of the immunoglobulin content version of these compounds into ketones has a glucose- of milk). Rather than being absorbed, as is the IgG, sow- sparing effect that may be passed on to nursing piglets. derived IgA coats the gut mucosal surfaces, forming a In particular, the MCT are readily metabolized. Consis- barrier against enteric diseases. Fangman et al. (1996a) tent with earlier findings (Pettigrew, 1981), Azain have speculated that scouring in weaned piglets re- (1993) found that the survival of low-birth-weight pig- sulting from transmissible gastro-enteritis virus (TGE) lets (< 900 g) was significantly improved by feeding exposure could be related to the withdrawal, associated triglycerides to sows. Survival rates of these small pig- with early weaning, of maternal IgA as the piglet lets in control, LCT, and MCT treatment groups were switches from a diet of sow’s milk to a solid feed. 32, 53, and 68%, respectively. Based on such findings, Neonatal mortality has been associated with low con- earlier authors have concluded that feeding sows high- centrations of serum IgG (Klobasa et al., 1981). Obvi- triglyceride diets could be beneficial in herds with a ously, factors that decrease colostrum/milk ingestion history of high preweaning losses (Pluske et al., 1995). reduce energy intake and protection against disease. Considering the importance of piglet nutrition for For instance, the decrease in suckling induced by cold neonatal survival, maintaining the lactational perfor- stress reduces the acquisition of maternal antibodies mance of the sow is a high priority. Sow feed intake (Kelley, 1985). Although birth order may have less in- during lactation has a recognized influence on milk fluence on total colostrum intake than once believed yield and the associated nutritional status of piglets as (Fraser et al., 1995), it may still influence the total well as rebreeding performance (Verstegen et al., 1985; amount of colostral IgG ingested by the piglet. Colostral Britt, 1986; England, 1986; Koketsu et al., 1996). Pro- IgG levels drop by 50% within 6 h of the first nursing; viding a healthy and a minimally stressful environment thus, late-born piglets may receive significantly lower for the lactating sow is an important management goal, levels of passive immunity than littermates born earlier because environmental and disease stressors all can in the farrowing order (Gaskins and Kelley, 1995). Ele- contribute to depressed intake (Matteri et al., 2000). vated environmental temperatures may also be a con- cern, because stress during late gestation is associated Disease with elevated circulating cortisol concentrations in As indicated above, causes of neonatal mortality are sows and their newborn piglets, increased cortisol con- tightly interlinked. Major death losses due to crushing centrations in colostrum, and lower serum IgG concen- E78 Lay et al. trations in piglets at 24 h of age (Machado-Neto et does decrease piglet mortality (e.g., Friend et al., 1988; al., 1987). McGlone et al., 1994). This management practice The thermal environment can also significantly in- gained momentum in the 1960s (Blackshaw et al., 1994) fluence disease susceptibility and subsequent survival as more economic pressures were applied to the swine (Kelley, 1985). The ability to maintain body tempera- industry. Unfortunately, piglet crushing remains a ture is lower in the diarrheic piglet (Balsbaugh et al., problem for swine producers. Data from NAHMS indi- 1986). Pigs that die of cold and chilling have lower cate that the rate of piglet crushing has remained at a gamma globulin concentrations on d 1 than those that high, stable rate from 1991 to 1995 (NAHMS, 1991, survive (Blecha and Kelley, 1981). The interaction be- 1995). The incidence of preweaning mortality continues tween thermal environment and disease status in the to average approximately 15% (NAHMS, 1991; Ko- neonatal piglet is multifaceted. Whereas a loss of body ketsu, 2000), representing a significant source of eco- temperature in newborn piglets can be caused by dis- nomic loss to the swine industry. Interestingly, approxi- ease (Matteri et al., 1998), exposure to a cold environ- mately 50% of these preweaning death losses occur dur- ment increases the neonatal piglet’s susceptibility to ing the first 3 d of life (Bauman et al., 1966; English immunological challenges (Carroll et al., 2000), thus and Smith, 1975; Cieslak, 1983; NAHMS, 1991). More increasing the risk of hypothermia. This work indicates specifically, it is during the first 48 h after farrowing that the pathogenic environment may be an important that the majority of crushing deaths occur (Rudd and factor with regard to chilling of newborn piglets. Inter- Marchant, 1995). Unfortunately, the incidence of crush- estingly, hypothermia resulting from an endotoxin chal- ing can be attributed to many factors such as low birth lenge can be blocked by ensuring a warm ambient tem- weights, environmental temperature, facilities, and perature (Carroll et al., 2000) or by preventative treat- disease, which contributes to the complexity of solving ment with aspirin-related compounds (Matteri et al., this problem. The implications of crushing on piglet 1999). Hypothermia, whether induced by thermal envi- well-being are obvious, but crating sows has raised con- ronment or disease challenge, increases the ther- cerns for the well-being of the sow, which must remain motaxic response of neonatal piglets, which often seek in an environment in which she may not perform typical warmth from the sow. Additionally, piglets experienc- maternal behaviors and in which her mobility is re- ing hypothermia due to disease challenge are typically stricted to such a point that she may not turn around. more lethargic and less active than healthy piglets, However, production parameters such as feed consump- and thus the risk of being crushed by the sow becomes tion and return to estrus remain high for the sow, which compounded by piglet behavior under these circum- suggests that the sow’s well-being is adequate in stances. these systems. The endemic pathogen status of the sow herd can Many experiments have been conducted to investi- also influence the disease status of the neonatal piglet. gate the effect of the design of the sow’s housing during Increased deaths due to infectious causes including farrowing in reducing preweaning mortality (Curtis et Streptoccocus sp., Escherichia coli, and Clostridium al., 1989; Blackshaw et al., 1994; Morris and Hurnik, perfringens and viruses such as rotavirus (Rota), coro- 1995). The typical farrowing crate of approximately 0.6 navirus (Corona), and TGE are more likely in litters of × 2.2 m is the prevalent form of housing today, but sows affected with these postparturient diseases (Fang- animal well-being continues to stimulate interest in the man et al., 1996b). farrowing pen. Most studies indicate that fewer pigs There has been a considerable shift in the prevalence die in crates than in pens. For instance, Blackshaw et of agents causing neonatal diarrhea in swine. Entero- al. (1994) reported mortality in farrowing crates of 14% toxigenic E. coli and TGE were once the most common and in farrowing pens of 32%. Ba¨ckstro¨m (1973) re- pathogens identified. Now these organisms are detected ported crushing of 3.4% in crates and 5.9% in pens. in only a small percentage (15%) of neonates with Also, crate design has been found to influence piglet scours. However, Rota continues to be a significant mortality; Curtis et al. (1989) found more crushing in problem and has been joined by Clostridium difficle and wide crates (64 cm wide) than in narrow crates (55 cm Porcine Reproductive and Respiratory Syndrome virus wide). Unfortunately, relatively little work has been as the most common pathogens identified in neonates conducted to determine the effect of gestation housing with diarrhea (Yaeger, 2000). on subsequent piglet survival. One study (Cronin et al., 1996) in which sows were housed in a gestation crate Sow and Piglet Behavior or pen and then farrowed in a pen or crate found that gestation environment did not affect the number of pig- The initial move to decrease piglet crushing consisted lets born or weaned, although the authors believed of confining the sow to a smaller pen than was tradi- there was a trend for the crate-housed sows to give tional. The incidence of crushing and related piglet mor- birth to fewer piglets. In contrast, Marchant et al. tality has significantly decreased since the popular (1994) found that litter size was greater for sows that adoption of the farrowing crate in the 1950s (Arey, gestated in crates; however, mortality rate was also 1993). Indeed, most studies have found that housing greater compared to that of piglets from sows that ges- sows in a small pen, or farrowing crate (0.6 × 1.7 m), tated in groups. He suggests that the differences in Preweaning survival in swine E79 mortality rate are likely due to the sow’s having a larger been selected to produce piglets that grow quickly and litter. This research also found that sows that were produce a large amount of lean mass. This selection group-housed during gestation produced more pigs if pressure has caused not only large piglets, but large they farrowed in a crate rather than in a pen. More sows as well. Unfortunately, as the sows have become research in this area will help to build complementary larger, the gestation crate and farrowing crate have not gestation and farrowing accommodations. changed in size. This mismatch between sow size and Several researchers have evaluated alternate meth- crate size is of important concern for both sow well- ods of confinement to determine whether sows could be being and productivity. Data collected by the NAHMS confined in more space but still maintain a low inci- (1995) indicate that 40.7% of sows are culled due to dence of crushing (Friend et al., 1988; McGlone and “age or size.” Sows kept in gestation crates also receive Morrow-Tesch, 1990; Cronin et al., 1996). McGlone and less exercise than sows kept in gestation pens or out- Morrow-Tesch (1990) noted a reduced piglet mortality side, resulting in reduced cardiovascular and muscular when sows farrowed in pens with a slope (approxi- fitness (Marchant et al., 2000). All these factors may mately 8%), whereas Grissom et al. (1990) found no increase the crated sow’s difficulty in lying down care- effect on piglet mortality when sows were housed in fully. Rudd and Marchant (1995) proposed that crates similar accommodations. Environmental factors such compensate for careless lying behavior, but they inhibit as types of flooring also have been found to influence careful lying behavior. the rate of piglet crushing (Walker and Knox, 1996). Activity level of the sow has also been found to affect These researchers found that although some floor types piglet mortality. Svendsen et al. (1986) found that most and crate configurations may save more piglets ini- traumas to piglets occur when the sow changes position; tially, all systems tested produced the same number of standing up, lying down, walking, and so on. This sug- weaned piglets. This was thought to be due to a system’s gests that a sow that spends more time lying quietly is saving small piglets early on, which altered death loss less prone to crushing her piglets than a sow that is prior to weaning. Cronin and van Amerongen (1991) more active. Sows housed in farrowing pens spend ap- found that sows housed in crates but given access to proximately 90% of the time lying during the 3 d follow- straw and covered with a hessian cover during far- ing farrowing (Hohenshell et al., 1996; Minnick et al., rowing were more responsive to the distress vocaliza- 1997). Compounding the chances of getting crushed is tions of their piglets, and piglet mortality was de- the finding that sows housed in crates performed twice creased. Sows farrowing in pens have been found to as many position changes between lie:sit and sit:lie have shorter birth intervals, wean the same number, compared to sows housed in pens, although pen-housed but heavier, piglets (Biensen et al., 1996), and have an sows performed more rolls (Weary et al., 1996a). In increased expression of maternal behavior (Cronin et addition, Haussmann et al. (1999) noted that some sows al., 1996). Unfortunately this increase in the expression possessed pressure point sores on their shoulders and of maternal behavior was not associated with an in- limb joints and that this may increase the number of crease in piglet survival. A great deal of variation exists position changes a sow makes; therefore, they adminis- in the current literature as to whether farrowing crates decrease piglet mortality. Several studies (McGlone and tered a potent analgesic to sows (butorphanol tartrate) Blecha, 1987; Arey and Sancha, 1996; McGlone and and found that butorphanol administration to sows Hicks, 2000) have found no difference in piglet mortal- within 4 h after farrowing decreased the total number ity when comparing a crate and pen (or outdoor) system. of body position changes that a sow makes during the One major reason changes in pen sizes and shapes 3 d after farrowing. These data indicate that research may not have been successful in decreasing crushing on improving farrowing crate flooring may improve sow is because piglets are attracted to their dam’s udder well-being and piglet survival. immediately upon birth and prefer to lie there the ma- Maternal Behavior. In conjunction with research on jority of time during the first 3 d after birth. After this different housing methods for sows to address the prob- initial 3 d, piglets are often seen using the heat lamp lem of piglet crushing, several researchers have exam- instead of the sow’s udder. This change of preference ined the sows’ behavior to determine exactly how piglets for lying area may help the piglets avoid death due to become crushed (e.g., Svendsen et al., 1986; Rohde Par- crushing. Although the sow’s housing environment has fet et al., 1989; Weary et al., 1996a). These researchers been shown to have a profound effect on piglet crushing, found that piglets are crushed when the sow changes the physical constraints of the sow and the behavior of position, essentially moving between lying and stand- both the sow and piglet should not be overlooked. ing and vice versa. However, Weary et al. (1996a) found Physical Constraints. The physical attributes of the that pen-housed sows also crushed a significant number sow may be one important factor in piglet crushing. For of piglets while changing lying positions and that most instance, the larger the sow, the less physical control crushings occurred during the 1st d of life. Interest- she may have over her body in the given space. The ingly, evidence exists to suggest that early experience percentage of piglets crushed has been found to be posi- affects maternal ability, and sows reared in group-hous- tively correlated with sow body length (Rudd and ing systems have been found to exhibit a lower piglet Marchant, 1995). During the previous 10 yr, sows have mortality rate (Wechsler, 1996). E80 Lay et al. Sows are capable of exhibiting beneficial maternal savaged, the decrease in savaging across parities could behavior, and confinement has been shown to prevent not be attributed to culling for the later parities. In a their natural “anti-piglet-crushing” behavioral reper- more comprehensive survey of commercial sows, Harris toire (Blackshaw and Hagelsø, 1990; Algers, 1992), thus et al. (2001b) evaluated the incidence of savaging and suggesting a reason for the variable success of crates. some factors that are correlated with this deleterious Feral sows perform pre-lying behavior that is designed behavior. They collected data from seven farms, repre- to remove piglets from the lying area (Rudd and senting 8,800 gilts and 5,232 sows. Their data revealed Marchant, 1995). This pre-lying behavior, consisting of that 5.3% of gilts expressed piglet directed aggression rooting through the bedding and vocalizing to the pig- and 2.9% of these gilts fatally savaged at least one of lets, may decrease piglet mortality. Blackshaw and Ha- their piglets. Aggressive behavior of gilts toward their gelsø (1990) reported that 79% of sows rooted before offspring resulted in a 0.6% death loss and 0.14% of lying down on the day after farrowing when housed in piglets were injured. Interestingly, these authors found pens. Marchant et al. (2001) found that sows housed in that if the lights were left on in the farrowing house, a group farrowing system were much more likely to a reduction in the incidence of savaging was realized. perform dangerous lying behavior and increase the risk Additionally, animals that savaged piglets as gilts were of crushing piglets when they did not perform piglet- more likely to savage during their second parity. The directed behaviors prior to lying. Interestingly, these behavioral differences at parturition in wild boar sows authors found that sows performed the greatest amount that savage may be indicative of problems in commer- of piglet-directed behaviors during the 1st d after far- cial sows. Appleyard et al. (2000) found that savaging rowing, the precise time that piglets are the most vul- sows were more active prior to farrowing, showing more nerable to becoming crushed. Sows confined to crates, ventral lying and shifting between positions. Similarly, in which they cannot turn, are hindered from per- Marchant et al. (2001) found that savaging sows are forming pre-lying behavior repertoires. more fearful of humans and that sows that readily inter- An outstanding anomaly in the piglet mortality prob- acted with humans were not savaging and were more lem is that the majority of sows do not respond to the protective of their litters. Fear of the piglets (Harris et distress vocalizations of their piglets when they are al., 2001a), lack of experience during adolescence (Knap being crushed. However, sows that are responsive to and Merks, 1987), and the pain associated with parturi- piglet distress calls are better able to release trapped tion (Hansen and Curtis, 1981) have all been implicated piglets prior to crushing (Wechsler and Hegglin, 1997). in savaging behavior; however, the definitive cause(s) One theory to explain the nonresponsiveness is that of savaging remains elusive. sows in farrowing crates are subjected to the distress Piglet Behavior. Piglet behavior during the early post- vocalizations of neighboring piglets, and regardless of natal period (d 1 to 3) is another area that has received their responses they cannot make the neighboring pig- considerable attention with regard to trying to reduce let stop vocalizing and thus they learn to be nonrespon- neonatal mortality. Similar to newborn mice, gerbils, sive when piglets vocalize. guinea-pigs, and rabbits (Dawes and Mestyan, 1963; Housing methods to reduce crushing may have met Eedy and Ogilvie, 1970), the newborn piglet huddles with variable success largely because research efforts with littermates against the dam, presumably to con- have concentrated on controlling and(or) altering the serve heat and metabolic fuel (Mount, 1959). This in- behavior of the sow and have largely ignored the piglets’ nate behavior ultimately increases the neonatal piglet’s role in crushing mortalities. Weary et al. (1998) found likelihood of being crushed by the sow when it is lying that it was a combination of increased sow parity, small down or rolling over (Wechsler and Hegglin, 1997). piglet size, and large litters that increased piglet crush- Weary et al. (1996) found that low-weight piglets spent ing, although one factor alone could not be implicated more time by the sow’s udder, actively massaging her as being most important because they are confounded. udder, and therefore were in greater danger of becom- Another anomaly in preweaning piglet deaths is due ing crushed. Intuitively, if piglets are huddling with to savaging of piglets by the sow. Savaging behavior is littermates and the sow to conserve heat, then provid- characterized by a sow that is overtly aggressive to her ing an alternative heat source to attract piglets away piglets and may result in injury and death to a portion from the sow should decrease the incidence of piglet of the litter. Savaging behavior has been noted in cap- crushing. In support of this idea are prior studies that tive wild boars. Harris et al. (2001a) found that 8.3% demonstrated a thermotaxic response of newborn pig- of wild boar sows used in their study killed piglets and lets when exposed to a thermocline (Balsbaugh et al., that one genetic line in particular was characterized 1986). Thus, initial research efforts focused on ex- by more aggression and a longer duration of parturition. ploiting the neonatal piglet’s innate thermotaxic re- English and Morrison (1984) examined the incidence sponse as a method for drawing the piglet away from the of savaging attempts (any attempt by the sow to bite sow by evaluating the location of various heat sources her piglet) of 31 farrowings and found that 89% of the (conventional heat lamps, heaters, and hovers) in far- gilts attempted to savage their piglets, and this percent- rowing crates. age decreased to approximately 20% for subsequent Early studies by McGinnis et al. (1981) and Saldierna farrowings. Because these sows were not culled if they et al. (1987) suggested that the use of supplemental Preweaning survival in swine E81 heat sources can be used to reduce the incidence of (two paths that would lead them away from the udder). diarrhea, and thus supplemental heat may be more The piglets’ attraction to the udder is most likely depen- advantageous to the young piglet’s overall health dur- dent on odor. When a sow farrows, the majority of the ing periods of pathogenic exposure. Although providing piglets move directly toward the udder, and very few supplemental heat to newborn piglets during the first venture the long way around the back. This indicates 2 d of life has been shown to increase survival rates that the piglets have a drive to move toward the udder, (Adams et al., 1980; McGinnis et al., 1981; Saldierna despite their known lack of significant vision at this et al., 1987), the addition of more supplemental heat time. Several factors are likely responsible for orien- sources, the type of heat sources (i.e., conventional heat tation. lamps, heaters, and hovers), and the location of supple- It is this attraction of the piglet, due to warmth, odor, mental heat sources in the farrowing crate (rear, front, and tactile stimuli, to the sow’s udder that puts the or lateral) do not result in further improvement in sur- piglet into a position to become crushed. In fact, Weary vival rates (Ogunbameru et al., 1991; Hrupka et al., et al. (1996) have shown that it is more probable to 1998). Hrupka et al. (1998) reported that during the find the smaller, weaker piglets near the udder, thus first 3 d of life the neonatal piglet tends to lie in close increasing their chance of becoming crushed. These au- proximity to the sow regardless of the heat source loca- thors hypothesized that these smaller piglets were hun- tion or the environmental temperature. Additionally, gry and thus maintained contact with the sow’s udder 1-d-old piglets spend the majority of their time (60 to in order to gain access to her warmth and milk. 75%) suckling the sow or huddling near the sow (Tit- Following up on these earlier studies, Lay et al. (1999) terington and Fraser, 1975; Lewis and Hurnik, 1985). explored the possibility that newborn piglets are at- Of the time spent in close proximity to the sow during tracted to the odor, as well as the warmth and tactile the 1st d of life, only about 40% of this time is associated properties, of the sow’s udder. They demonstrated that with suckling (Lewis and Hurnik, 1985). Therefore, the the presence of a simulated udder, possessing olfactory, newborn piglet spends 20 to 35% of its time in close tactile, and thermal characteristics of the sow, in the proximity to the sow without suckling, increasing the farrowing crate could attract piglets away from the sow risk of death by crushing for no apparent reason. Thus, more effectively than heat lamps alone. Further work in order to make significant progress in reducing pre- by Hrupka et al. (2000a,b) examining the thermal, vi- weaning mortality losses, an understanding of environ- sual, and physical cues that may act as attractants mental factors other than temperature that modulate to neonatal piglets demonstrated that piglets choose the behavior of the newborn piglet must be addressed. physical contact with an anesthetized littermate in a It is intuitive that piglets should have a high at- cold environment (24°C) as opposed to a warm environ- traction to their dam’s udder and its associated warmth, ment (45°C) without the presence of a littermate. Addi- nutrition, and comfort. Several studies have examined tionally, they demonstrated that visual recognition the auditory, olfactory, visual and tactile cues in order does not influence huddling; piglets demonstrated no to identify those factors that drive the neonatal piglet preference in huddling activity between littermates and to huddle with littermates and the sow (Welch and nonlittermates. Collectively, these data suggest that Baxter, 1986; Rohde Parfet and Gonyou, 1991; Hrupka tactile and olfactory cues are the primary innate stimuli et al., 2000a,b). Welch and Baxter (1986) reported that that increase the risk of piglet crushing from birth to 3 piglets are attracted to the physical and thermal proper- d of age. It should be mentioned, however, that although ties of the sow’s udder, and Rohde Parfet and Gonyou supplemental heat may not be considered a primary (1991) reported that the attraction to the sow’s udder attractant, its positive role in the prevention of hypo- is associated with the odor of the sow’s milk. Rhode thermia and illness in the newborn piglet is without Parfet and Gonyou (1991) noted that piglets have a question. Until we can develop sows with good maternal tendency to move in the direction of the dam’s orienta- behavior, future studies that perfect simulated udder tion, thus directing them toward the udder. Upon birth, designs such as that used by Lay et al. (1999) may piglets have a highly developed sense of smell and are prove to be the most effective means to reduce neonatal attracted toward the dam’s udder. Morrow-Tesch and mortality caused by crushing in the newborn piglet. In McGlone (1990) found that piglets were highly at- the meantime, the presence of a trained attendant may tracted to the odor of their dam’s feces and teat wash- be highly beneficial in reducing piglet loss due to crush- ings and that they learned this attraction within the ing (Holyoake et al., 1995). first 12 h of life. Similarly, Horrell and Hodgson (1992) Facility design was shown to be an important influ- noted that piglets were able to distinguish odors associ- ence on sow behavior, and the same may be true for ated with their dam (urine, feces, udder) compared to the piglet. Rohde Parfet et al. (1989) have shown that an unfamiliar dam and that they were particularly at- crate design affected important behaviors of the piglet. tracted to the wood shavings that had been previously In fact, Blackshaw et al. (1994) found that piglets would associated with their dam’s udder. Rhode-Parfet and spend twice the amount of time at their dam’s udder if Gonyou (1991) found that piglets were attracted to the they were housed in a crate compared to a pen. If a odor of the sow’s milk. However, they were also at- sow’s piglets would move to the heat lamps or pads tracted to the odor of birth fluids and sow vocalizations provided to them, they would certainly decrease their E82 Lay et al. chances of becoming crushed. Svendsen et al. (1986) maternal characteristics in sows. Interestingly, it was noted that it took piglets up to 48 h to start using these probably the introduction of farrowing crates into the heat sources, and Blackshaw and Hagelsø (1990) noted swine industry that allowed producers to put less em- that 94% of piglets would stay under their heat source phasis on maternal behaviors and more emphasis on by d 8 after parturition. Obviously, no matter how care- other reproductive traits, thus inadvertently imple- ful and attentive a sow may be, if her piglets will not menting a selection program for nonmaternal behaviors move when she lies, she has little choice but to crush that contribute to piglet crushing. In support of this them. theory are the maternal characteristics of the Meishan sow from China. These sows are known to produce large Sex litters, and Meishan piglets have been found to have a 5% advantage in piglet survival rate over Large White Although slightly more males than females are born piglets (Bidanel et al., 1990). Meunier-Salaun et al. in a litter, females have a greater survival advantage (1991) reported that Meishan sows had 13.6 piglets than males (Bereskin et al., 1973; Svendsen et al., 1986; Becker, 1995). In addition, a larger litter was found to born alive and weaned 12.4 piglets, whereas Large be more detrimental to survival for male than for female White sows farrowed 8.6 live piglets and weaned 7.4. piglets. Males are more susceptible to stillbirth, weak- The superior production of the Meishan has been attrib- ness/starvation, and crushing (Svendsen et al., 1986). uted to behavioral characteristics and a greater number McGlone et al. (1993) reported that castration of male of teats. Hohenshell et al. (1996) studied the behavior piglets at any age leads to less time spent nursing and of Meishan sows to determine how their behavior imme- standing and more time spent lying, which may lead diately after parturition differed from that of our mod- to more clinical and subclinical disease, resulting in ern production sow. Their preliminary research on greater mortality compared to females. More recently, these sows indicates that the Meishan sow may be more Becker (1995) reported that the greater mortality in vigilant and aware of her piglets’ location and then males was due to more males being crushed and chilled. she quickly lies down without crushing them. Common Although the underlying mechanism(s) responsible breeds used in U.S. pork production may also vary in for this sexual dimorphism in preweaning mortality their rate of crushing piglets. Curtis et al. (1989) found rates has not been elucidated, there are some unique that Duroc-sired sows crushed fewer piglets than Lan- sex differences that may account for these observations. drace- or Hampshire-sired sows. Unfortunately, mod- For instance, the higher basal concentration of cortisol ern genetic selection of breeding stock has emphasized observed in the male piglet as compared to the female piglet growth characteristics, number of piglets born piglet (Ruis et al., 1997) may cause male piglets to be alive, and number of piglets weaned, not sow maternal more susceptible to the deleterious effects of stress and behavior. Therefore, maternal behavior varies widely therefore succumb to subsequent disease challenges. among sows. One possible factor that may account for a greater The increase in neonatal mortality typically seen rate of crushing in male piglets is their potential for with increasing litter size differs significantly among an increased sensitivity to pheromones. In a study by swine breeds. With 10 liveborn piglets, preweaning Dorries et al. (1995), male piglets were reported to have mortalities in Large White and Pietran swine have been a fivefold increase in their sensitivity to androstenone reported at approximately 12 and 20%, respectively compared to female piglets. Although the focus of this (Blasco et al., 1995). A positive effect of heterosis on study was on sensitivity of adult piglets to a sex phero- piglet survival is known to exist, and an improvement mone, it is plausible that a more highly developed vom- of 5 to 6% in survival can be expected in litters of cross- eronasal sensitivity in the neonatal male piglet could bred, compared with purebred, piglets (Blasco et al., result in an increased amount of time spent near the 1995). An additional concern is raised by rapid changes sow due to pheromones associated with the sow’s udder, in genetics related to lean growth. Selective breeding thus increasing its risk of being crushed. could result in metabolic and compositional changes that affect the newborn piglet’s ability to adapt to the Genetics postnatal environment. Relative to Meishan and Euro- The sow is apparently the only mammal that pro- pean white breeds, crossbred piglets selected for en- duces large litters and lies on a significant portion of hanced lean tissue growth have been shown to possess them within days of parturition. Because of the risk lower body fat stores and a reduced ability to metabolize associated with parturition and the repartitioning of triglycerides (Herpin et al., 1993). The genetic correla- nutrients associated with producing young, piglet tion between backfat thickness and daily feed intake crushing by the dam is not consistent with current evo- is 0.55 (Stewart and Schinckel, 1991), thus selection lutionary theory. Likely, the modern sow has been al- for decreased fatness results in decreased feed intake. tered through genetic selection, resulting in its being This could result in sows that are unable to consume a “poor” mother. Algers (1992) suggests that farrowing adequate feed to meet the milk production needs of a crates have reduced the selection pressure for beneficial large litter. Preweaning survival in swine E83 Sow and Piglet Well-Being aging gifts. In: Proc. 34th Int. Congr. Int. Soc. of Appl. Ethol., Florianopolis, Brazil. p 62. The use of gestation and farrowing crates is certainly Arey, D. S. 1993. The welfare of pigs in confined and non-confined one of the major targets of growing international con- farrowing systems. Pig News Info. 14:81–84. Arey, D. S., and E. S. Sancha. 1996. Behavior and productivity of cerns for swine well-being. Sows housed under re- sows and piglets in a family system and in farrowing crates. stricted housing conditions, such as the gestation and Appl. Anim. Behav. Sci. 50:135–145. farrowing crate, and restricted feed conditions during Azain, M. J. 1993. Effects of adding medium-chain triglycerides to gestation are known to perform stereotypic behaviors sow diets during late gestation and early lactation on litter per- such as bar biting and excessive rooting. In addition, formance. J. Anim. Sci. 71:3011–3019. Ba¨ckstro¨m, L. 1973. Environment and animal health in piglet produc- a large percentage of sows have limb abrasions, lame- tion. Acta Vet Scand. 41:1–240. ness, stomach ulcers, and a high group mortality rate Balsbaugh, R. K., S. E. Curtis, R. C. Meyer, and H. W. Norton. 1986. (Deen and Xue, 1999; Geiger et al., 1999) indicating Cold resistance and environmental temperature preference in that their well-being is not adequate. However, sows diarrheic piglets. J. Anim. Sci. 62:315–326. housed in less-restrictive environments, as groups, Bauman, R. H., J. E. Kadlec, and P. A. Powlen. 1966. Some factors affecting death loss in baby pigs. Purdue Univ. Agric. Exp. Sta. must contend with competition from penmates and are Bull. 810. West Lafayette, IN. often subjected to aggression from which they cannot Becker, B. A. 1995. Effects of season and sex on piglet mortality. J. escape. Obviously, this raises a different set of well- Anim. Sci. 73(Suppl. 1):131 (Abstr.). being concerns for group-housed sows. A gestation sys- Becker, B. A., and M. L. Misfeldt. 1993. Evaluation of the mitogen- tem such as that developed by Hurnik and Morris (Mor- induced proliferation and cell surface differentiation antigens of lymphocytes from pigs 1 to 30 days of age. J. Anim. Sci. ris and Hurnik, 1995) may well improve the well-being 71:2073–2078. of group-housed sows; these researchers found that Benito, M. 1985. Contribution of brown fat to neonatal thermogenesis. sows in the Hurnik-Morris system performed fewer ste- Biol. Neonate 48:245–249. reotyped behaviors and produced more piglets. Al- Bereskin, B., C. E. Shelby, and D. F. Cox. 1973. Some factors affecting though more scientific data are needed, the popular pig survival. J. Anim. Sci. 36:821–827. Berthon, D., P. Herpin, R. Bertin, F. De Marco, and J. Le Dividich. concept is that sow well-being is improved in systems 1996. Metabolic changes associated with sustained 48-hr shiv- that provide more natural physical and social environ- ering thermogenesis in the newborn pig. Comp. Biochem. Phys- ments (Wechsler, 1996). Mortality and morbidity in the iol. B Biochem. Mol. Biol. 114:327–335. piglets, however, appear to be compromised in a large Berthon, D., P. Herpin, C. Duchamp, M. J. Dauncey, and J. Le Divi- portion of such alternative production environments dich. 1993. Modification of thermogenic capacity in neonatal pigs by changes in thyroid status during late gestation. J. Dev. (Arey, 1993; Wechsler, 1996; Marchant et al., 2000). Physiol. 19:252–261. Bidanel, J. P., J. C. Caritez, and C. Legault. 1990. 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H. Ca´ rdenas2 and W. F. Pope

The Ohio State University, Columbus 43210

ABSTRACT: Follicular recruitment and atresia are ovulation rate in swine. Administration of testosterone important processes associated with ovulation rate in or the nonaromatizable androgen dihydrotestosterone swine. Follicle-stimulating hormone regulates granu- increased the number of ovulations in gilts in a dose- losa cell division, differentiation, and steroidogenic dependent manner. Furthermore, administration of di- function, and, as such, significantly influences follicular hydrotestosterone increased the amounts of FSH recep- growth and development. Follicle-stimulating hormone tor mRNA in pig preovulatory follicles. Steady-state is an inducer of follicular recruitment in swine and an amounts of FSH receptor mRNA are relatively high inhibitor of granulosa cell apoptosis, and it seems to be during the early follicular phase but decrease signifi- a major regulator of ovulation rate in swine. Although cantly as follicles grow and approach ovulation, sug- local factors, such as growth factors and steroid hor- gesting that major changes in amounts of FSH receptor mones, might regulate follicular development by con- mRNA occur during late follicular development in pigs. trolling the expression of gonadotropin receptors or by Local ovarian factors that regulate follicular respon- modulating other related processes, the dominant role siveness to gonadotropins seem to be important compo- of FSH cannot be ignored. Recent results indicate that nents of the mechanisms that control ovulation rate androgens might be among the local factors regulating in pigs.

Key Words: Ovarian Development, Ovulation Rate, Pigs

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E36–E46

Introduction grow or how many follicles die (atresia). Follicular growth or atresia will be the main focus of the present A fundamental function of the ovary is to support review. Initially this review will describe follicular de- development of oocytes. Oogenesis begins during fetal velopment and then will partition factors that affect life and is highly influenced, later during the reproduc- recruitment from atresia. This overview will continue tive period, by follicular development. Beginning at pu- by reviewing some recent observations from our labora- berty, about 10 to 25 follicles develop to the ovulatory tory that suggest a novel role of androgens in regulating stage and release oocytes every estrous period. The total follicular development. Finally, a brief description of number of ovulations is called the ovulation rate and how some other factors might regulate recruitment or it is an important parameter of swine reproductive effi- atresia will be discussed. ciency. It should be noted that although ovulation rate is a limiting factor in determining the number of off- Folliculogenesis spring born, numerous additional factors act upon the uterus and conceptuses throughout gestation and con- Primordial follicles form the stock from which all folli- tribute to the number of healthy fetuses that develop cles emerge (Peters, 1978). Approximately 500,000 pri- to term. mordial follicles are present in both ovaries by 10 d after Factors that alter ovulation rate might function by birth in swine (Black and Erickson, 1968). Primordial modulating, directly or indirectly, how many follicles ovarian follicles in this population are not stimulated to grow at the same time, and only a small number begin their development while the rest remain quiescent. In 1Salaries and research support provided by state and federal funds mammals, this characteristic increases the possibilities appropriated to The Ohio Agric. Res. and Dev. Center and The Ohio of having progeny throughout a rather long reproduc- State Univ. Manuscript no. 16-01AS. tive life. 2 Correspondence: Dept. of Anim. Sci., The Ohio State Univ., 2027 Initiation of growth of primary follicles involves endo- Coffey Rd. (phone: 614-292-0714; fax: 614-292-7116; E-mail: [email protected]). crine actions and regulatory effects of local factors from Received April 5, 2001. the somatic cells of the follicle (Hirshfield, 1991) and Accepted August 29, 2001. probably from the growing oocyte (reviewed by Picton

E36 Ovulation rate in swine E37 et al., 1998). Early follicular growth is characterized by et al., 1987). Follicular dominance, which occurs in cat- an increase in the number and layers of granulosa cells, tle (Ko et al., 1991) and primates (Zeleznik and Kubik, which subsequently separate from each other resulting 1986), has not been demonstrated in pigs. in formation of the antrum, a fluid-filled cavity (Zam- Within each mammalian species, ovulation rate is boni, 1974). Initial studies using hypophysectomy tech- regulated within a relatively narrow range. Changes niques, and more recent experiments using knockout in rates of follicular recruitment or atresia could alter mice lacking a functional FSH receptor (FSHR) gene the number of follicles that ovulate. This notion has (Dierich et al., 1998), indicated that FSH seems to be been supported in recent studies. For instance, Meishan absolutely necessary for antrum formation and postan- sows have a higher ovulation rate than Large White tral follicular development. Therefore, development of hybrids (27.7 vs 17.6 CL, respectively) and, probably responsiveness to FSH might be a key factor controlling related, Meishan sows have more follicles ≥ 2mmond follicular differentiation and function. 16 than Large White sows. Also, in Large White sows Growth rates of antral follicles of pigs and other spe- the number of follicles decreased from d 16 to 19, but cies follow an exponential curve because of a dramatic this decrease does not occur in Meishan sows (Miller increase in proliferation of granulosa cells and an in- et al., 1998). Collectively, these observations suggest crease in size of the antrum (Clark et al., 1975; Grant that the greater ovulation rates in the Meishan breed et al., 1989). Activated primary follicles of pigs require than in Large White hybrids is related to differences approximately 84 d for growing to the antral stage and in both recruitment and atresia. Alternatively, the pos- an additional 19 d to grow to the preovulatory size of sibility that ovulation rate can be altered by manipulat- 10 mm (Morbeck et al., 1992). The rate of growth of ing the incidence of atresia alone has been shown in porcine follicles from 3 to 10 mm in diameter has been mice that exhibit increased ovulation rate and litter estimated to be 1.14 mm/d (Dailey et al., 1976). During size following overexpression of the antiapoptotic factor final maturation, cells of the theca and granulosa layers bcl-2 (Hsu et al., 1996). synthesize and secrete significant amounts of steroids, peptide hormones, prostaglandins, and other sub- Follicle-Stimulating Hormone is an Effective stances. These hormones are important because they Inducer of Follicular Recruitment participate locally in follicular development, and some of them convey signals that coordinate the functions of The number of follicles that are recruited might de- the hypothalamic-hypophyseal-ovarian axis. pend on concentrations of FSH or other regulatory fac- Follicular recruitment refers to the formation of a tors, or on the number of follicles responsive to FSH pool of antral follicles from which the ovulatory folli- present at the time of recruitment. Whether the latter is cle(s) is subsequently selected (Fortune et al., 1991). related to some endocrinological event occurring before Follicular recruitment in pigs occurs between d 14 and recruitment, or what other factors might influence it, 16 of the estrous cycle (Clark et al., 1982; Foxcroft and is not known. Several relationships support a regula- Hunter, 1985) or shortly after weaning. On d 16, ap- tory role of FSH on follicular recruitment in swine, and proximately 40 to 50 follicles, 2 to 6 mm in diameter, are some have been known for many years. Treatment with present in both ovaries (Grant et al., 1989). A number of FSH or PMSG on d 15 and 16 of the estrous cycle in- these follicles are growing and represent the population creases the number of ovulations in gilts, but the effect of recruited follicles. Most follicles, and the oocytes they significantly diminishes when given after d 16 (Hunter, contain, degenerate and disappear from the ovaries 1979). Administration of FSH or PMSG increases ovula- through the process of atresia. Although atresia may tion rate in estrus-synchronized sows (Guthrie et al., occur any time during development of antral follicles, 1997) and induces follicular recruitment in prepuberal most follicles, for a given species, are lost during the gilts (Paterson, 1982) or during the luteal phase in post- transition from the small to large size (Fortune, 1994). pubertal gilts in a manner that is dose-dependent Therefore, only a small proportion of recruited follicles (Hunter, 1979). Partial suppression of FSH by adminis- (about 30 to 40%) are selected to complete final matura- tration of porcine follicular fluid to gilts during the early tion and ovulate while the remaining become atretic, follicular phase reduced the subsequent number of most of them before reaching 6 mm in diameter (Grant small, medium, and large follicles, possibly due to in- et al., 1989; Dailey et al., 1976). Ovulatory follicles are hibin present in follicular fluid (Guthrie et al., 1987). not readily identifiable until d 20 (Grant et al., 1989), In another experiment, the numbers of medium-sized which suggests that the process of selection is not com- follicles were partially restored when FSH was given pleted until near ovulation. Therefore, the final number following treatment with porcine follicular fluid of ovulatory follicles is determined by how many folli- (Guthrie et al., 1988; Knox and Zimmerman, 1993). cles are recruited and by the ability of recruited follicles Active immunization against inhibin increased concen- to continue to grow and avoid atresia. Because follicles trations of FSH (area under the curve) by 27% and might become atretic, even close to the time of ovula- number of ovulations by 39% in gilts (King et al., 1993). tion, the process of selection probably takes place con- A similar increase of 35% in ovulation rate was reported tinuously, over the entire follicular phase (from luteo- by other investigators in gilts immunized against in- lysis at approximately d 13 to 14 to ovulation; Foxcroft hibin (Brown et al., 1990). However, diminishing the E38 Ca´rdenas and Pope decrease in FSH that occurs after weaning in sows by creased FSH stimulation might allow only those folli- passive immunization against inhibin did not alter cles that have reached certain developmental condi- numbers of piglets born (ovulation rate was not evalu- tions to continue to grow, to avoid atresia, and to ovu- ated; Wheaton et al., 1998). late. Although this concept seems attractive as a Despite the effects of exogenous FSH on ovulation mechanism of follicular selection in pigs, it will need rate, which indicate that it is a major inducer of follicu- to be examined further. As FSH stimulation becomes lar recruitment in pigs, a clear increase in FSH concen- attenuated, additional support for follicular develop- trations coincident with the period of follicular recruit- ment could come from growth factors, LH, and ste- ment has not been detected in gilts (Flowers et al., 1989; roid hormones. Guthrie and Bolt, 1990) or sows (Shaw and Foxcroft, Unlike FSH, LH remains almost unchanged during 1985; Foxcroft et al., 1987). A small increase in plasma the follicular phase, except near the onset of estrus FSH concentrations occurs around estrus in postpuber- when the preovulatory LH surge occurs (Brinkley, tal gilts, coincident with the preovulatory LH surge, 1981; Van de Wiel et al., 1981; Guthrie and Bolt, 1990). which is followed by a second increase approximately Luteinizing hormone binding in theca and granulosa 72 h later. No significant changes in concentrations of cells increase as follicles become larger during the follic- FSH occur during the luteal phase until the time of ular phase (Nakano et al., 1977; Foxcroft and Hunter, luteolysis, when FSH concentrations decrease gradu- 1985), increasing the ability of late-developing follicles ally through the period of estrus (Brinkley, 1981; Van to respond to LH. Amounts of LH receptor mRNA in de Wiel et al., 1981; Guthrie and Bolt, 1990; Guthrie granulosa and theca interna cells increases as follicles et al., 1993). Likewise, during the follicular phase, FSH grow from 2 to 6 mm in diameter and then decrease in binding and FSH-stimulated adenylate cyclase activity 8-mm preovulatory follicles (Liu et al., 2000), indicating by granulosa cells decrease as follicular size increases that the LH receptor gene is down-regulated as follicles (Nakano et al., 1977; Lindsey and Channing, 1979). approach ovulation. Moreover, amounts of FSHR mRNA determined using Specifically, what FSH does to stimulate follicular in situ hybridization (ISH) decrease from the time of recruitment in swine is an important question. In vitro recruitment to undetectable levels at estrus (Liu et al., experiments conducted in swine or other species indi- 1998). In another study, FSHR mRNA became unde- cate that FSH might influence granulosa cell division by tectable by ISH procedures when follicles reached 4 mm stimulating mitogen-activated protein kinases (Babu in diameter in sows after weaning (Liu et al., 2000). et al., 2000) or cell cycle regulators such as cyclin D2 Experiments using reverse-transcription polymerase (Sicinski et al., 1996). Also, FSH significantly stimu- chain reaction (RT-PCR), a very sensitive technique, lates estradiol synthesis by several mechanisms includ- indicated that FSHR mRNA in surface walls of the ing regulation of expression of the CYP 19 gene that largest follicles present from d 13, 15, and 17 of the encodes for aromatase cytochrome P450 enzyme estrous cycle were not different but decreased by d 19 (Hickey et al., 1988), synthesis of progesterone (Ford (about fivefold relative to d 13, Ca´rdenas and Pope, and Howard, 1997), and expression of LH receptors, unpublished data). This significant down-regulation of an important factor in the availability of aromatizable FSHR around estrus is only present in late-developing androgens. Inhibin secreted by maturing follicles is a follicles, because small antral follicles permanently ex- potent enhancer of LH-stimulated androgen synthesis hibit high amounts of FSHR throughout the estrous by theca cells (Hillier et al., 1994). Estradiol, in turn, cycle (Yuan et al., 1996; Liu et al., 1998). stimulates granulosa cell proliferation, enhances the The decrease in plasma FSH concentrations and actions of FSH and LH on steroidogenesis (Daniel and amounts of FSHR mRNA in late-developing follicles Armstrong, 1980; Richards, 1980), and appears to be suggests a diminished influence of FSH on porcine folli- absolutely necessary for follicular growth and matura- cles as they advance to the preovulatory stage. This tion (reviewed by Drummond and Findlay, 1999). Dif- does not necessarily mean that FSH is not needed for ferences in litter size not necessarily related to ovula- recruitment. The relatively greater concentrations of tion rate have been associated to the chromosomal locus FSH before recruitment, and the concentrations pres- where the estrogen receptor alpha gene is located (re- ent during recruitment, although decreasing, are prob- viewed by Rothschild et al., 1997). ably enough to support follicular growth and function. Although FSH seems to be important in the control of For example, synthesis of estradiol, a process partially follicular recruitment, a relationship between ovulation regulated by FSH, is still increasing even though FSH rate and peripheral concentrations of FSH has not been and FSHR mRNA are decreasing. The significance of clearly demonstrated. For instance, lines of pigs dif- the FSH decrease is not apparent. Perhaps, lesser con- fering in ovulation rate did not exhibit differences in centrations of FSH would inhibit incorporation of more LH, FSH, progesterone or estradiol during the estrous follicles into the cohort of recruited follicles, thus de- cycle (Mariscal et al., 1998). Similarly, no associations creasing follicular heterogeneity. Reduced diversity were found between plasma concentrations of FSH and among follicles is important to ensure uniform response ovulation rate in other species such as sheep (Gibbons to ovulation or decrease diversity in oocyte or embryonic et al., 1999). However, differences in the ability of folli- development (Pope et al., 1990). Furthermore, de- cles to respond to FSH through the influence of other Ovulation rate in swine E39 factors such as the number of FSHR might exist and androgens in the follicular compared to nonatretic folli- could help to explain the differences in ovulation rate. cles (Maxson et al., 1985; Guthrie et al., 1993; Ca´rdenas Cell signaling by FSH is mediated by a plasma mem- and Pope, 1994). Atresia of pig follicles was also associ- brane receptor, which is present exclusively in granu- ated with a decrease in expression of mRNA for losa cells in females and is associated with the cAMP- aromatase and gonadotropin receptors (Tilly et al., protein kinase A and probably calcium second messen- 1992a) and with a loss of aromatase activity (Maxson et ger systems (Flores et al., 1992; Simoni et al., 1997; al., 1985). Although these findings did not demonstrate Touyz et al., 2000). Factors that regulate expression cause-effect relationships, they seemed to indicate a of the FSHR gene have been investigated in different role for estrogens and gonadotropins as inhibitors of species. In cultured rat granulosa cells, amounts of atresia. This notion was supported by other experi- FSHR mRNA were increased by treatment with FSH ments in which estrogen treatment decreased apoptosis (Tilly et al., 1992b). The effect of FSH on FSHR mRNA in granulosa cells of rats while testosterone antago- was enhanced by IGF-I, which apparently occurred due nized the effects of estrogens (Billing et al., 1993). More to an increase in mRNA stability (Minigeshi et al., recently, Meishan sows that exhibited lesser rates of 2000). In cultured granulosa cells of swine, small atresia than Large White hybrids during the follicular amounts of FSH increased FSHR mRNA (Sites et al., phase were observed to have greater concentrations of 1994); however, more recent results demonstrated that estradiol in follicular fluid (Miller et al., 1998). FSH, and other ligands that signal through cAMP, de- Follicle-stimulating hormone decreased apoptotic creased steady state amounts of FSHR mRNA (Murphy DNA fragmentation by 60% and was regarded as a and Dobias, 1999). Activin increases the number of FSH major suppressor of apoptosis in cultured rat granulosa receptors in preantral follicles and prevents premature cells or follicles (Chun et al., 1994, 1996). Likewise, luteinization of large antral follicles (Findlay, 1993; FSH, as well as IGF-I, decreased DNA fragmentation Minigishi et al., 2000). Findlay (1993) speculated that in porcine granulosa cells in culture (Guthrie et al., the effect of activin on FSHR could be of particular 1998). Kaipia and Hsueh (1997) proposed that FSH importance for initiation of follicular development. In could directly regulate apoptotic factors or influence primates, testosterone administration increased FSHR other inhibitors of granulosa cell death or growth pro- mRNA in granulosa cells of primary follicles but did moters. Recently, mcl-1, a new member of the bcl-2 not alter FSHR mRNA in more advanced follicles (Weil family, inhibited apoptosis in rat granulosa cells, and et al., 1999). We have determined the effects of andro- amounts of its message were enhanced by gonadotropin gens on amounts of FSHR mRNA in porcine follicles treatment (Leo et al., 1999). and the results are discussed below. The Role of Androgens in Follicular Follicle-stimulating Hormone is an Inhibitor of Development and Ovulation Rate Atresia in Granulosa Cells Follicular theca and granulosa cells undergo steroido- Follicular atresia in swine (Tilly et al., 1991; Guthrie genesis in a cooperative manner (Short, 1962). Aromati- et al., 1995), and probably most mammals (Kaipia and zable androgens (testosterone and androstenedione) Hsueh, 1997), is primarily induced by programmed cell are synthesized by thecal cells upon stimulation by LH, death or apoptosis of granulosa and theca cells. which then diffuse to granulosa cells for subsequent Apoptosis is characterized by internucleosomal DNA conversion into estradiol (pigs, Evans et al., 1981). In fragmentation, cell shrinkage, plasma membrane bleb- pigs, synthesis by theca cells and follicular fluid concen- bing, and formation of apoptic bodies (Hsu and Hsueh, trations of androstenedione during the mid- and late- 2000). Although considerable progress has been made follicular phase are severalfold greater than those of in understanding the mechanisms of cell death by testosterone (Evans et al., 1981; Tsang et al., 1985). apoptosis, the description of the apoptotic pathway in The conversion of androgens into estradiol requires granulosa cells is not complete. Some results indicate stimulation by FSH (Dorrington et al., 1975; Moon et that, as in other cell types, caspases are the final in- al., 1975). Androstenedione can be converted into tes- ducers of granulosa cell degradation (Hsu and Hsueh, tosterone in a reversible reaction catalyzed by 17β-hy- 1998). The members of the bcl-2 family of apoptotic and droxysteroid dehydrogenase or aromatized to estrone, antiapoptotic factors are upstream in this pathway and which then can be converted to estradiol-17β. Testoster- some of them have been shown to be functional in gra- one is directly aromatized into estradiol-17β (Peters nulosa cells (Hsu and Hsueh, 1998). and McNatty, 1980). The specific factors that induce atresia in porcine Evidence has accumulated to indicate that androgens follicles have been difficult to identify. Initially, at- play regulatory functions in follicular development. An- tempts to answer this question were made by determin- drogens enhanced FSH-stimulated progesterone pro- ing hormonal changes in follicles undergoing atresia duction and aromatase activity in rat and primate gra- relative to healthy follicles. For example, atretic folli- nulosa cells in vitro (Armstrong and Dorrington, 1976; cles had lower concentrations of estradiol and contained Daniel and Armstrong, 1980; Harlow et al., 1986). Simi- similar or greater concentrations of progesterone and larly, androgens stimulated [3H]thymidine incorpora- E40 Ca´rdenas and Pope Table 1. Effects of low amounts of androgens on numbers of corpora lutea and conceptuses or offspring in pigs

Corpora lutea Conceptuses or offspring

Treatment Control Androgen Control Androgen Reference

Testosterone (d 17 and 18, 1 mg/d) 14.4 16.7* 12.3 (d 11) 15.0* Ca´rdenas and Pope, 1994 — — 9.6 (Total born) 10.7* Unpublished Testosterone (d 13 to estrus, 1 mg/d) 14.8 16.2* 12.8 (d 11) 15.3* Ca´rdenas and Pope, 1997 13.3 14.9* 12.5 (d 4–7) 13.6* Ca´rdenas and Pope, 1997 Androstenedione (d 13 to estrus, 1 mg/d) 14.3 15.1 12.5 (d 11) 12.9 Ca´rdenas and Pope, 1997 DHT (d 13 to estrus, 6 ␮g/[kg BWؒd]) 17.4 20.3* 15.9 (d 11) 9.4* Ca´rdenas, unpublished DHT (d 13 to estrus, 60 ␮gؒ[kg BWؒd]) 19.5 26.6* 15.5 (d 3) 15.7 Ca´rdenas, unpublished *Different from control (P < 0.05). tion (Bley et al., 1997) and lipoprotein utilization would be translated into more piglets born. Results (Schreiber et al., 1984) by granulosa cells. Androgens (unpublished) of a field trial indicated that treatment also stimulated development of mouse follicles in vitro of multiestrous gilts with vehicle (85 gilts treated and (Murray et al., 1998), and administration of androgens mated) or 1 mg of testosterone (84 gilts treated and to primates enhanced early development of ovarian fol- mated) on d 17 and 18 of the estrous cycle increased licles (Vendola et al., 1998, 1999). Moreover, we demon- litter size by approximately one pig (9.6 in vehicle- vs strated for the first time that administration of andro- 10.7 in testosterone-treated gilts, P = 0.03). Farrowing gens during the follicular phase increased ovulation rates (gilts farrowing/gilts treated and mated) were 80 rate in postpubertal gilts. A summary of results using and 76.2% in vehicle- and testosterone-treated gilts, re- small doses of androgens is presented in Table 1. In spectively. one experiment, testosterone administration on d 17 It is possible that the effects of testosterone on ovula- and 18 of the estrous cycle increased ovulation rate in tion rate described above could have been mediated gilts, apparently by decreasing the incidence of atresia by estradiol. Alternatively, it might be possible that (Ca´rdenas and Pope, 1994). In another experiment, a testosterone, independent of estradiol, acting via the longer period of testosterone administration, from d 13 androgen receptor (AR), enhanced transcription of cer- to estrus, not only increased ovulation rate, but also tain genes involved in follicular development. In addi- increased the percentage of blastocysts surviving to d tion, androgens have been shown to induce rapid + 11 of gestation (Ca´rdenas and Pope, 1997). The total changes in intracellular Ca 2 in luteinizing granulosa number of blastocysts was therefore increased by the cells (Machelon et al., 1998). Effects of this nature are combined effects of testosterone on ovulation rate and so-called nongenomic and apparently do not involve the survivability of conceptuses. It is possible that factors AR; however, participation of membrane receptors has that enhance follicular development, such as testoster- been proposed for nongenomic actions produced by one, could optimize oocyte growth, resulting in concep- other steroids (reviewed by Wehling, 1997). Nongeno- tuses with greater ability to survive. mic actions of androgens have not been demonstrated The effect of testosterone on ovulation rate was also in vivo, and it is not clear whether they could be involved observed when follicular recruitment and ovulation in the effects of testosterone observed in our exper- were artificially induced during the luteal phase, indi- iments. cating that this effect of testosterone is not necessarily Kreider et al. (2001) recently reported that active im- related to or influenced by the physiological conditions munization against androstenedione increased ovulation present during the follicular phase. In this experiment, gilts were administered daily injections of vehicle or 1 Table 2. Numbers of induced corpora lutea (CL) in mg of testosterone on d 11 to 16 or 4 to 9 of the estrous gilts treated with vehicle or testosterone coincident cycle. Follicular recruitment was induced the 1st d of with induction of ovulation using PMSG (750 IU) testosterone treatment using PMSG, and follicles were and hCG (750 IU) during the luteal phase induced to ovulate by a single injection of hCG, 72 h of the estrous cycle later. Testosterone treatment increased (P < 0.05) the number of induced corpora lutea when treatment began Beginning of treatment Vehicle Testosterone = on d 4, but the effect (P 0.09) was not clear when Day 11a 15.7 ± 2.4 22.4 ± 2.6 testosterone treatment began on d 11 (Table 2). Super- Day 4b 5.9 ± 1.1c 11.9 ± 2.5d imposing testosterone treatment on ovulation induction aExperiment 1. Gilts (n = 10 per treatment) received PMSG on d using PG600 in prepubertal gilts did not alter ovulation 11, hCG on d 14, and daily doses of 1 mg of testosterone on d 11 to rate but had a significant effect on increasing embryonic 16. Induced CL were counted on d 23. survival (Table 3). An important practical question has bExperiment 2. Gilts (n = 18 per treatment) received PMSG on d 4, hCG on d 7, and daily doses of 1 mg of testosterone on d 4 to 9. been whether the increase in ovulation rate and embry- Induced CL were counted on d 12. onic survival produced by treatment with testosterone c,dMeans within a row with different superscripts differ (P < 0.05). Ovulation rate in swine E41 Table 3. Corpora lutea (CL) and d-11 blastocysts in prepubertal gilts induced to ovulate with PG600 and treated daily with 1 mg of testosterone from day of PG600 administration to mating

Treatment Number of CL Number of blastocysts Blastocyst survival, %

Vehicle (n = 19) 16.1 ± 1.3 11.4 ± 1.3 71.3 ± 6.1a Testosterone (n = 19) 16.7 ± 1.6 14.5 ± 1.6 85.6 ± 3.2b

a,bMeans within a column with different superscripts differ (P < 0.05). rate in gilts by approximately three corpora lutea (ovula- whether the effects of DHT on ovulation rate were associ- tions). It is not clear how immunization against andro- ated with specific periods of the follicular phase. Gilts stenedione, a weak androgen that can be converted into received daily i.m. injections of vehicle from d 13 to estrus testosterone and estrogens in ovarian follicles, would or they received 60 ␮g/kg of DHT from d 13 to estrus increase ovulation rate in pigs. In a previous experiment, (entire follicular phase), d 13 to 16 (follicular recruit- active immunization against androstenedione increased ment), or d 17 to estrus (postrecruitment). Results dem- ovulation rate in gilts, did not affect concentrations of onstrated that all three periods of DHT administration FSH and LH, but increased concentrations of andro- increased ovulation rate and that numbers of embryos stenedione in follicular fluid (McKinnie et al., 1988). Per- in gilts treated from d 13 to estrus were not different haps it can be speculated that an increase in androstene- compared with controls but increased in gilts treated dione would increase testosterone and estradiol in the from d 13 to 16 or d 17 to estrus (Ca´rdenas et al., 2002). follicular fluid, which in turn would stimulate follicu- In another trial, gilts received daily injections of vehi- lar development. cle or 6 ␮g/kg BW of testosterone or DHT from d 13 to The AR and receptors for other steroid hormones, thy- estrus. Tissue from the largest follicles was collected the roid hormones, retinoids, and vitamin D are structurally 1st d of estrus and processed for determination of AR related proteins that regulate gene transcription (Tsai and FSHR mRNA by RT-PCR. Amounts of AR mRNA and O’Malley, 1994). A long (A, 110 kDa) and a short were not altered by androgen treatment. However, a (B, 87 kDa) form of AR, having similar functional activi- significant increase (about onefold) in amounts of FSHR ties, are present in different organs and are derived from mRNA occurred in gilts treated with DHT compared differences in translation initiation of the AR mRNA with those that received vehicle (Ca´rdenas et al., 2002). (Gao and McPhaul, 1998). Similar to that in rats and These results using DHT demonstrate that stimula- primates (Tetsuka et al., 1995; Hillier et al., 1997), the tion of AR, probably at the ovary, is capable of altering AR protein in swine ovaries was localized mainly in gra- late follicular development, resulting in increased ovula- nulosa cells of small and medium-sized follicles during tion rate. The effects of DHT were more pronounced the 1st wk following ovulation (Garret and Guthrie, relative to those of testosterone, which might be ex- 1996). We determined the relative amounts of the AR plained by the greater androgenic activity of DHT com- protein and mRNA in ovarian follicles of swine from d pared with testosterone in other cell types. This effect 13 to 19 of the estrous cycle using immunohistochemistry of DHT is unique, and based on present knowledge only and RT-PCR. These preliminary results (Ca´rdenas and treatment with FSH, eCG or, in the recent study, DHT Pope, unpublished data) suggested that AR protein and has increased ovulation rate at this magnitude in pigs. mRNA are 1) present in maturing follicles during the Although the mechanisms involved in this effect of DHT follicular phase and 2) their relative amounts do not seem are probably numerous, the up-regulation of FSHR mRNA might be one of the mechanisms in place. An to change during follicular growth from recruitment to effect of this kind would increase the ability of follicles the preovulatory stage. to respond to the stimulatory effects of FSH during re- To examine the involvement of the AR in ovulation cruitment or to continue growing and be selected into rate in pigs, we administered daily injections of 0, 6, 60, the ovulatory pool, when otherwise they could become or 600 ␮g/kg BW of dihydrotestosterone (DHT) from d atretic. These effects could also increase the estrogenic 13 to estrus. Dihydrotestosterone binds and activates activity of growing follicles due to the effects of FSH on the AR, but unlike testosterone it cannot be aromatized aromatase. The relationships just described might be due to 5 -reduction of the A ring (Wilson, 1975). Ovula- α helpful in understanding the control of ovulation rate in tion rate significantly increased by approximately 3, 10, pigs and are summarized and integrated with previous and 17 with each additional dose of DHT. However, the knowledge in Figure 1. number of d-11 blastocysts drastically decreased as dose of DHT increased and no blastocysts were recovered in Other Factors That Alter Ovulation Rate May gilts that received the highest dose of DHT (Ca´rdenas Modulate the Actions of Major Regulators et al., 2002). Low recovery rates of blastocysts were asso- of Follicular Growth or Atresia ciated with opaque uterine flushings, indicating that uterine function could be altered by overexposure to It is well recognized that nutritional conditions may DHT. In a subsequent experiment, it was determined affect reproductive performance of domestic mammals E42 Ca´rdenas and Pope

Figure 1. Follicle-stimulating hormone (FSH) is a major factor that stimulates follicular growth and induces follicular recruitment during the early follicular phase (d 14 to 16 of the estrous cycle) in swine. Recruited follicles continue their phase of rapid growth and most of them (60 to 70%) become atretic before d 17. The number of recruited follicles and incidence of atresia are key determinants of the number of follicles that ovulate. Follicular growth following recruitment might continue to be supported by FSH; however, other factors such as luteinizing hormone (LH), estradiol, and growth factors possibly become increasingly important due to decreasing concentrations of FSH and amounts of FSH receptor (FSHR). Up-regulation of FSHR (dotted line) might be a way to increase responsiveness to FSH, resulting in enhanced follicular growth and decreased atresia. including swine, and that maintaining normal levels not alter ovulation rate or embryonic survival (Zak et of nutrition is important for optimal ovarian function. al., 1997, 1998). Likewise, administration of GnRH ev- Under certain conditions, particularly related to animal ery 6 h during feed restriction from d 22 to 28 of lacta- age, follicular development becomes responsive to tion failed to affect ovulation rate, embryonic survival, changes in feed intake. For instance, increasing feed or plasma concentrations of progesterone in primipa- intake (flushing) in pubertal gilts increased ovulation rous sows (Mao et al., 1999). rate, but the treatment was less effective in postpuber- The effects of nutrition on ovulation rate might be tal gilts (Rhodes et al., 1991). In the review by Prunier mediated by changes in secretion of growth factors, and Quesnel (2000), it was concluded that in postpuber- metabolic hormones, and gonadotropins (Flowers et al., tal gilts, nutrition restriction consistently decreased 1989; reviewed by Cox, 1997) and(or) their interactions. ovulation rate when applied during the luteal and follic- Components of the IGF system have been identified in ular phases; however, in most experiments with sows, ovarian follicles of swine and have been shown to be nutrition restriction did not alter ovulation rate when associated with follicular growth (Mondschein et al., imposed during lactation or after weaning. Recent ex- 1991; Yuan et al., 1996, Liu et al., 2000) and partial periments partially support these conclusions. For in- inhibition of follicular atresia (Guthrie et al., 1998). stance, in gilts that were fed a high plane of nutrition Transforming growth factor β has also been shown to during the entire estrous cycle or were restricted during influence granulosa cells (Gangrade and May, 1990; d 1 to 7 or 8 to 15, ovulation rate was not altered but Chang et al., 1993). Interestingly, although there have embryonic survival by d 28 was lower in gilts restricted been some indications that administration of insulin fromd8to15than in the other groups (Almeida et al., increases ovulation rate, the results have not been con- 2000). In another experiment, 50% restriction or 125% sistent (reviewed by Cox, 1997). Administration of insu- overfeeding during lactation in primiparous sows did lin to primiparous sows after weaning tended to in- Ovulation rate in swine E43 crease the number of follicles on d 5 but reduced concen- the estrous cycle in gilts on subsequent fertility. J. Anim. Sci. trations of estradiol, IGF-I, and IGF-I mRNA in large 78:1556–1563. Armstrong, D. T., and J. H. Dorrington. 1976. Androgens augment follicles (Whitley et al., 1998b). In a recent study (van FSH-induced progesterone secretion by cultured granulosa cells. den Brand et al., 2000a), changes in plasma insulin Endocrinology 99:1411–1414. concentrations were induced by feeding sows a high- or Babu, P. S., H. Krishnamurthy, P. J. Chedrese, and M. R. Sairam. low-energy diet (with fat or starch as energy source) 2000. Activation of extracellular regulated kinase pathways in during a 21-d lactation period. Postprandial plasma ovarian granulosa cells by the novel growth factor type 1 follicle- glucose and insulin were greater and ovulation rate stimulating hormone receptor. J. Biol. Chem. 275:27615–27626. Billing, H., I. Furuta, and A. J. W. Hsueh. 1993. Estrogens inhibit tended to be greater in sows fed the starch diet than and androgens enhance ovarian granulosa cell apoptosis. Endo- in sows fed the fat diet, and there was no association crinology 133:2204–2212. between glucose or insulin and LH concentrations. In Black, J. L., and B. H. Erickson. 1968. Oogenesis and ovarian develop- a follow-up experiment (van den Brand et al., 2000b) ment in the prenatal pig. Anat. Rec. 161:45–55. ovulation rate tended to be greater in sows fed the high- Bley, M. A., P. E. Saragueta, and J. Lino-Baran˜ ao. 1997. Concerted stimulation of granulosa cell deoxyribonucleic acid synthesis by energy than in those fed the low-energy diet. These sex steroids and follicle-stimulating hormone. J. Steroid Bio- results indicate that increasing dietary energy, and the chem. Mol. Biol. 62:11–19. consequent increase in insulin, did not clearly affect Brinkley, H. J. 1981. Endocrine signaling and female reproduction. ovulation rate. Similarly, ovulation rate was not altered Biol. Reprod. 24:22–43. when insulin was administered for4dtoprimiparous Brown, R. W., J. W. Hungerford, P. E. Greenwood, R. J. Bloor, D. F. Evans, C. G. Tsonis, and R. G. Forage. 1990. Immunization sows beginning the day after weaning (Whitley et al., against recombinant bovine inhibin alpha subunit causes in- 1998a) or when feed-restricted primiparous sows re- creased ovulation rate in gilts. J. Reprod. Fertil. 90:199–205. ceived insulin for 5 d during the weaning period (Ques- Ca´rdenas, H., J. R. Herrick, and W. F. Pope. 2002. Increased ovulation nel and Prunier, 1998). Although altering nutrition has rate in gilts treated with dihydrotestosterone. Reproduction not been able to consistently change ovulation rate in 123:(In press). Ca´rdenas, H., and W. F. Pope. 1994. Administration of testosterone sows, some follicular characteristics were altered, and during the follicular phase increased the number of corpora lutea it is not clear whether these changes could perhaps in gilts. J. Anim. Sci. 72:2930–2935. affect oocyte development. Ca´rdenas H., and W. F. Pope. 1997. Administration of testosterone Differences in ovulation rate between breeds, or lines from day 13 of the estrous cycle to estrus increased the number within breeds, have been demonstrated. Ovulation rate of corpora lutea and conceptus survival in gilts. J. Anim. can respond to selection and lines of pigs with greater Sci.75:202–207. Cassady J. P., R. K. Johnson, and J. J. Ford. 2000. Comparison of ovulation rate than others have been developed (John- plasma FSH concentrations in boars and gilts from lines selected son et al., 1999). Selection for FSH concentrations to for ovulation rate and embryonic survival, and litter size and increase ovulation rate was estimated to be effective estimation of (co)variance components for FSH and ovulation and more practical than direct selection on ovulation rate. J. Anim. Sci. 78:1430–1435. rate (Cassady et al., 2000). This supports the impor- Chang, W. Y., H. Ohmura, S. K. Kulp, and Y. C. Lin. 1993. 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Vendola K., J. Zhou, O. O. Adesanya, S. J. Weil, and C. A. Bondy. growth factor I (IGF-I) system in sows after weaning. J. Reprod. 1998. Androgens stimulate early stages of follicular growth in Fertil. 112:175–184. primate ovary. J. Clin. Investig. 101:2622–2629. Wilson, J. D. 1975. Metabolism of testicular androgens. In: R. O. Vendola K., J. Zhou, J. Wang, O. A. Famuyiwa, M. Bievre, and C. Greep and E. B. Astwood (ed.) Handbook of Physiology. Sect 7: A. Bondy. 1999. Androgens promote oocyte insulin-like growth Endocrinology. vol 5. Male Reproductive System. pp 491–508. factor I expression and initiation of follicle development in the American Physiological Society, Washington, DC. Yuan, W., M. C. Lucy, and M. F. Smith. 1996. Messenger ribonucleic primate ovary. Biol. Reprod. 61:353–357. acid for insulin-like growth factors-I and -II, insulin-like growth Weil, S., K. Vendola, J. Zhou, and C. A. Bondy. 1999. Androgen factor-binding protein-2, gonadotropin receptors and steroido- and follicle stimulating hormone interactions in primate ovarian genic enzymes in porcine follicles. Biol. Reprod. 55:1045–1054. follicle development. J. Clin. Endocrinol. Metab. 48:2951–2956. Zak, L. J., J. R. Cosgrove, F. X. Aherne, and G. R. Foxcroft. 1997. Wehling, M. 1997. Specific, nongenomic actions of steroids hormones. Pattern of feed intake and associated metabolic and endocrine Annu. Rev. Physiol. 59:365–393. changes differentially affect postweaning fertility in primipa- Wheaton, J. E., R. L. Meyer, R. H. Jones, and A. J. Kramer. 1998. rous lactating sows. J. Anim. Sci. 75:208–216. Effects of passive immunization using antibody against an α- Zak, L. J., I. H. Williams, G. R. Foxcroft, J. R. Pluske, A. C. Cegielski, inhibin peptide on follicle-stimulating hormone concentrations E. J. Clowes, and F. X. Aherne. 1998. Feeding lactating primipa- and litter size in sows. Theriogenology 49:813–822. rous sows to establish three divergent metabolic states: I. Associ- Whitley, N. C., D. B. Payne, H. Zhang, and N. M. Cox. 1998a. The ated endocrine changes and postweaning reproductive perfor- mance. J. Anim. Sci. 76:1145–1153. influence of insulin administration after weaning the first litter Zamboni, L. 1974. Fine morphology of the follicle wall and follicle on ovulation rate and embryo survival in sows. Theriogenology cell-oocyte association. Biol. Reprod. 10:125–149. 50:479–485. Zeleznik, A. J., and C. J. Kubik. 1986. Ovarian responses in macaques Whitley, N. C., M. N. Quirk-Thomas, J. O. Skelton, A. B. Moore, J. to pulsatile infusion of follicle-stimulating hormone (FSH) and Purvis, Y. Qiu, and N. M. Cox. 1998b. Influence of insulin on luteinizing hormone: increasing sensitivity of the maturing folli- follicular development and the intrafollicular insulin-like cle to FSH. Endocrinology 119:2025–2032. Early embryonic survival in the pig: Can it be improved?1

R. D. Geisert*2 and R. A. M. Schmitt†

*Department of Animal Science, Oklahoma State University, Stillwater 74078 and †Seaboard Farms Inc., Guymon, OK 73942

ABSTRACT: A major limitation for increasing litter ment can result in loss of embryos before d 10 of gesta- size in swine is embryonic loss that occurs during the tion. Competitive acquisition of adequate uterine space 2nd to 3rd wk of gestation. High ovulation rates of between littermate embryos, essential for blood flow modern sows have more than supplied the potential delivery of nutrients needed for survival to term, is number of embryos necessary to improve litter size. The established during conceptus elongation on d 12 of ges- current challenge is determining how early conceptus tation. Progressive changes in the uterine microenvi- development affects the ability to maintain the viability ronment between d 10 and 16 of gestation play a major through the remaining 90 d of gestation to maximize role in embryonic survival following trophoblast elonga- farrowing house production. To achieve this, it is neces- tion and placental attachment. In current production sary to identify and understand the possible causes of systems, there can still be sufficient numbers of em- embryonic death. Because fertilization rates are gener- bryos present after d 30 of gestation to provide improve- ally high in swine, early embryonic loss during the first ment in average litter size at farrowing. However, pro- 20 d of gestation is considered to critically effect poten- ducers are still faced with the challenge of maximizing tial litter size. There are three periods during which fetal survival to term. Therefore, fully understanding early embryonic loss can occur: 1) pre-elongation devel- the biological controls of follicle ovulation rate, syn- opment, 2) trophoblastic elongation, and 3) placental chrony of ovulation, embryonic developmental rate, uni- attachment. The first two periods are related to time formity of conceptus elongation, uterine horn capacity, of fertilization and subsequent developmental rate for uterine glandular and vascular development, and pla- each individual embryo within the litter. Asynchrony cental vascularization could provide possible clues to in embryonic development relative to uterine develop- improving embryo quality.

Key Words: Embryo, Pigs, Pregnancy, Uterus

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E54–E65

Introduction ovary do not survive through gestation (Pope, 1994). Fertilization of ovulated swine ova is generally greater Improving litter size would have a substantial impact than 95%; therefore, the loss of potential piglets is pre- on the efficiency of swine production; Tomes and Niel- dominately the result of early embryonic (d 10 to 30) son (1982) and Rothschild (1996) have indicated that and fetal (d 31 to 70) deaths. A previous review by Pope overall profitability increased as the number of pigs (1994) indicated that although ovulation rate estab- per sow per year increased. Although it has long been lishes potential piglet number, an increase in ovulation recognized that increasing litter size would increase rate by itself would not result in a marked improvement the efficiency of production, significant improvements in litter size. Ovulation rate responds to direct selection have not been achieved. There is potential for average in swine, but the returns for improving litter size have litter sizes of 14 or more pigs if all ovulated ova develop been minimal (Cunningham et al., 1979; Lamberson et into live pigs, but the average number of pigs born per al., 1991). Eleven generations of selection for uterine litter in the United States is only 10.5 (USDA, 2001); capacity using the unilateral-hysterectomy-ovariec- approximately 30 to 50% of the ova released from the tomy (UHOX) model has also failed to significantly im- prove overall litter size of gilts (Christenson and Ley- master, 2000). However, index selection for ovulation rate and embryonic survival, designed to maximize re- 1 Approved for publication by Director, Oklahoma Agric. Exp. Sta. sponse in litter size, has been effective in increasing 2Correspondence: phone: 405-744-6077; fax: 405-744-7390; E-mail: [email protected]. litter size after 14 generations of selection (Johnson et Received April 5, 2001. al., 1999). Therefore, future improvements in litter size Accepted August 15, 2001. may be realized if we gain a clearer understanding

E54 Early embryonic survival in the pig E55 of the balance of follicular maturation and timing of jority of the uterine surface, the perimplantation em- ovulation, as well as embryonic, uterine, and placental bryos migrate and space themselves throughout both factors associated with conceptus development and uterine horns between d 8 to 11 of gestation (Pope et survival. al., 1982a; Dzuik, 1985). Following migration within and between the uterine horns, conceptuses undergo a Day 10 to 30 Is a Critical Stage rapid differentiation and expansion of their trophoblas- in Conceptus Development tic membranes between d 11 and 12 of gestation (Ander- son, 1978; Geisert et al., 1982a,b). Conceptuses change Establishment and maintenance of pregnancy in- in diameter and morphology as they develop from a 1- volves a number of closely integrated signals between to 2-mm sphere to a 3- to 8-mm ovoid and a 9- to 20- the ovary, uterus, and conceptus in most mammalian mm tubular shape on d 10 to 11 (Geisert et al., 1982b). species (Bazer et al., 1982; Roberts et al., 1993). Alter- Once reaching approximately 10 mm in diameter, con- ations in the synchronous development of the uterine ceptuses rapidly elongate (2 to 3 h) to a thin, filamen- environment with the growing and differentiating con- tous (> 100 mm) form and initiate attachment to the ceptus during early pregnancy can result in a failure uterine surface epithelium (Dantzer, 1985; Blair et al., to establish pregnancy (Geisert and Yelich, 1997). In 1991; Burghardt et al., 1997). Rapid expansion of the monotocous domestic species, failure of the embryo to trophoblast, which is linked to an increase in estrogen develop and survive results in termination of the preg- synthesis and release, is followed by a continuation of nancy, but in the polytocous pig, failure of individual placental growth (Ka et al., 2001), and the trophoblast embryo survival within the uterus is a norm that does can exceed1minlength by d 16 of gestation (Perry not necessarily affect continuation of the pregnancy. and Rowlands, 1962). Expansion of the conceptus However, total loss of conceptuses early in development throughout the uterus of the pig is essential, because would result in a failure to maintain functional corpora early pregnancy is not maintained if a substantial por- lutea beyond d 15 of gestation (Bazer et al., 1984). tion of each horn is not occupied by conceptuses (see The majority of embryonic mortality usually occurs Dzuik, 1987; Geisert et al., 1990). The morphological prior to d 20 to 30 of gestation (Pope, 1994). Data for change in the elongating conceptus is not the result of the gilt, excluding females in which total fertilization cellular hyperplasia, but rather of cellular remodeling failure occurs, have indicated that little embryonic loss (Geisert et al., 1982a; Mattson et al., 1990; Pusateri et takes place before 7 d after the onset of estrus (Polge, al., 1990). 1982). Estimates suggesting that the majority of embry- Rapid transition in conceptus morphology followed by onic mortality occurs between d 10 and 30 of gestation initiation of placental attachment to the uterine surface are not surprising given the critical events that occur represents one of the periods of greatest embryonic loss during this period. The reasons for which early embry- in the pig (Pope and First, 1985). Conceptus secretion onic mortality occurs can be quite extensive and com- of estrogen and development from a spherical to fila- plex (see Dzuik, 1987). Improper timing of conceptus mentous morphology occur concurrently with the period and uterine development, which involves synthesis of of pregnancy establishment in the gilt (see Geisert et nutrients and attachment factors, failure of conceptus al., 1990) and are temporally related to changes in endo- signaling, competition of embryos (uterine crowding), metrial estrogen (Geisert et al., 1993) and progesterone and genetic factors all contribute to early conceptus (Geisert et al., 1994) receptors. The decline in uterine loss. The following review will attempt to bring together epithelial progesterone receptors on d 12 of pregnancy the current information available on factors that are and the presence of epithelial estrogen receptors in the proposed to be involved with conceptus development, uterine endometrium during d 10 to 18 relates to the factors leading to embryonic mortality, and the many effects of estrogen on the uterine epithelial glycocalyx, approaches taken to improve embryonic survival in conceptus attachment, and alterations in endometrial the pig. secretory products (Dantzer, 1985; Blair et al., 1993; Geisert and Yelich, 1997; Ka et al., 2000). Following Conceptus Development conceptus elongation and the initiation of estrogen re- lease, trophoblastic attachment to the epithelial lining To understand the cause for loss of apparently of the endometrium occurs between d 13 and 18 of gesta- healthy embryos during the first third of pregnancy, one tion (King et al., 1982; Burghardt et al., 1997). The first needs to understand how pig conceptuses interact embryo develops from the inner cell mass and the allan- with the uterine environment to modulate endometrial toic membrane expands to form the placental vascular function and placental attachment for nutrient ex- network for transport of nutrients and fluid into the change from the mother (reviewed by Stroband and Van allantoic cavity (Bazer et al., 1981; King et al., 1982). der Lende, 1990). In the pig, conceptus synthesis and Physiological events during early pregnancy are con- release of estrogen on d 11 to 12 and 15 to 20 of gestation trolled by ovarian progesterone secretion, conceptus es- provides the signal to maintain luteal production of trogen synthesis, and the cellular localization of endo- progesterone throughout gestation (Geisert et al., metrial steroid receptors (see Geisert and Yelich, 1997; 1990). In order to deliver estrogen throughout the ma- Burghardt et al., 1997). E56 Geisert and Schmitt It is evident that three events, rapid trophoblastic also confirmed that less-developed embryos within a elongation, conceptus estrogen synthesis, and placental litter are just as capable of surviving when not placed attachment to the uterine epithelial surface, are critical in competition with more-developed littermates. for early porcine embryonic survival. Early trophoblast Possible reasons for the diversity of conceptus devel- expansion most likely regulates and limits the final opment within a litter have been suggested. Consider- size of each embryo’s placental surface area throughout able information has indicated that the Ped (preimplan- gestation. The synthesis and release of growth factors tation embryo development) gene influences the time and cytokines by the developing conceptus during the to cleavage and thus the rate of early development in preimplantation period of embryonic development are mice (Warner et al., 1998; Wu et al., 1999). The Ped considered to play potentially critical roles in cellular gene product is the major histocompatibility complex growth, differentiation, and remodeling of the embryo (MHC) class Ib, Qa-2 protein that is expressed on the and extraembryonic tissues. Both embryonic and endo- cell surface, and it seems to function through a cross- metrial production of various growth factors, as well as linking mechanism to influence rate of cleavage in early interactions between the maternal endometrium and mouse embryo development (McElhinny and Warner, developing conceptus, are essential for providing the 2000; McElhinny et al., 2000). The lack of inbred lines synergistic environment for endocrinological and im- to discern the presence of a similar relationship in the munological signals for establishment of pregnancy (see swine leucocyte antigen (SLA) MHC has limited our review by Geisert and Yelich, 1997; Ka et al., 2000). knowledge of the Ped gene in pigs. There has been some Developmental events that influence embryonic differ- indication for haplotype differences in ovulation rate entiation and growth are most likely regulated through and number of embryos in inbred miniature pigs (Con- the programmed expression of growth factors and their ley et al., 1988), but its effect on embryo development specific receptors. Both the conceptuses and endome- has not been established. A search for the human homo- trium must actively participate in balancing the pres- logue to the Ped gene is currently underway (Warner ence of many proteolytic enzymes and their inhibitors et al., 1998); if it is identified this may assist in locating to mediate the epitheliochorial type of placental attach- the Ped gene in swine. ment and growth in the pig (see Geisert and Yelich, Although the data for pigs are extremely limited, we 1997). must also not overlook the possible paternal contribu- The previous information provides a base to under- tions to early conceptus development (Dzuik, 1987). The stand the many temporal interactions that occur be- fertilizing sperm can influence variation in fertility; Eid tween the conceptuses and the uterine environment et al. (1994) indicated that the rate and time of entry during establishment of pregnancy and placental at- of the oocyte into the S-phase of the cell cycle is different tachment. The following sections will now try to provide with sperm from high- vs low-fertility bulls. The transi- some insight into the cause(s) of early conceptus loss tion from maternal to embryonic genome regulation of and attempts to alleviate mortality. cell function occurs at the four-cell stage of development in the pig (Prather, 1993). The classic experiments of Litter Embryonic Diversity Surani et al. (1986) first demonstrated the critical func- tion of both the maternal and paternal genome in modu- One of the easiest times to observe embryonic devel- lating normal embryonic and placental growth in mice. opmental diversity within individual litters is to recover Genomic imprinting (see John and Surani, 2000) has embryos from the uterus between d 11 and 12 of preg- been demonstrated to have major effects on placental nancy. Perry and Rowlands (1962) and Anderson (1978) development and fetal survival in mice (Georgiades et demonstrated the size and morphological differences al., 2001). Insulin-like growth factor-2 and insulin-like that can occur within a litter prior to and during the growth factor-2 receptor are two of the genes proposed period of trophoblastic elongation. Geisert et al. (1982b) to be imprinted during early embryonic development indicated that the conceptuses initiate elongation upon (see Moore and Reik, 1996). The presence of insulin- reaching an approximate spherical diameter of 10 mm like growth factors in the porcine uterine lumen and on d 11 to 12. However, although these conceptuses can insulin-like growth factor receptors in the developing elongate, littermates that are 7 to 8 mm or smaller conceptus (Geisert and Yelich, 1997) certainly suggest may not elongate for at least another 4 to 8 h. This that if imprinting occurs similarly to that in the mouse, transitional lapse in development can obviously create the paternal genome could contribute to the rate of a problem in gaining appropriate uterine territory for conceptus growth and differentiation in the pig. survival of the less-developed embryos to term if uterine Asynchronous oocyte maturation and variation in space is limiting. The classic studies of William Pope time of ovulation for a cohort of developing antral folli- clearly illustrated that competition between embryos cles have been linked to the diversity of embryos within can occur within the uterus. Through embryo transfer, a litter (Pope et al., 1990; Pope, 1994). Utilizing it was demonstrated that asynchrony in development transrectal utlrasonography to evaluate time of ovula- of healthy porcine conceptuses leads to a competitive tion in pigs, Soede et al. (1992) determined that ovula- advantage of the more developmentally advanced lit- tion of follicles occurred within 1 to 4 h and was not termates (Pope et al., 1982b, 1986b). Wilde et al. (1988) related to embryo diversity. However, a number of stud- Early embryonic survival in the pig E57 ies have reported that the duration of ovulation can The second period of uterine selectivity of embryonic vary from 1 to 9 h between individual animals (see survival occurs with conceptus elongation and estrogen Pope, 1994). Although the majority of follicles ovulate synthesis on d 11 to 12 of pregnancy. Vallet et al. (1998) over a short period of time, a few (one to four) ovulate documented that changes in uterine secretion on d 11 over a more protracted period. Pope et al. (1988) demon- to 12 of pregnancy are related to progressive alterations strated that elimination of the late-ovulating follicles in protein secretion induced by progesterone rather on d 1 of the estrous cycle removed the diversity in than being an event completely triggered by the concep- embryo development at d 11 of gestation. Furthermore, tus. Precise timing for the changes in uterine secretion Xie et al. (1990a,b) demonstrated that the later-ovulat- most likely occurs with progesterone-induced down-reg- ing follicles become the less-developed embryos in the ulation of uterine epithelial progesterone receptors on litter. These later-ovulating oocytes clearly contribute d 10 of the estrous cycle and pregnancy (Geisert et al., to the pool of embryos that are less-developed on d 1994). Loss of progesterone receptor is specific to the 11 of gestation and, therefore, maybe the first to be endometrial surface and glandular epithelium, because eliminated from the litter when uterine space is limited. receptor levels are maintained in the stroma. The ele- Whether it is time of fertilization or genes that control gant study of Cunha et al. (1983) first demonstrated cleavage rate of the embryos, some major and minor the interactions between developing epithelium and differences in embryo development within a litter do mesenchyme. It is clear that epithelial-mesenchyme in- exist. teractions are involved with hormonal responsiveness of the uterus (Cooke et al., 1998; Kurita et al., 1998). Many effects of progesterone on uterine epithelial func- How Can Embryonic Diversity Determine Survival? tion can be attributed to progesterone activation of stro- It is clear that embryonic diversity can affect survival mal progesterone receptors and the stimulated release of littermate embryos (Wilmut et al., 1985; Pope, 1994). of progestamedins such as keratinocyte growth factor (KGF) (Ka et al., 2000). A switch from epithelial proges- When does selection of embryos begin? There are two terone regulation to stromal control would not only periods of uterine selectivity during early embryonic allow an alteration in uterine protein and enzyme re- development in the pig. The first period of embryonic lease into the uterine lumen, but also induce changes loss (d 5 to 10) is not related to competition between in epithelial apical glycocalyx permissive to placental embryos but to individual embryo asynchrony with its attachment (Dantzer, 1985; Burghardt et al., 1997; uterine environment. A portion of early selection could Geisert and Yelich, 1997). Uterine release of tissue kal- be attributed to meiotically immature or genetically likrein, a serine protease, on d 12 of the estrous cycle abnormal ovulated oocytes that fail to develop at all or in the pig may function to alter the uterine epithelial at a normal rate in pace with uterine development. glycocalyx for attachment (Vonnahme et al., 1999) and Indeed, Hunter (2000) recently reviewed the factors activate many growth factors for conceptus growth involved in oocyte maturation and follicle heterogene- (Geisert et al., 2001). These studies indicate that the ity, which if suboptimal could compromise subsequent uterus dictates the period of rapid conceptus growth embryo development. Koenig and Stormshak (1993) in- and attachment following hatching from the zona pellu- dicated that immature and chromosomally abnormal cida. However, there are also select changes in uterine ova may represent as many as 30 to 40% of the oocytes release of ions, growth factors, and enzymes that are ovulated in gilts, and this could account for the lower stimulated by the elongating conceptus (Geisert and embryo survival in gilts bred on first vs third estrus. Yelich, 1997; Ka et al., 2000). Several studies have indi- These data suggest that the quality of the oocytes ovu- cated that estrogen from the tubular and elongating lated may play an important role in subsequent devel- conceptuses can alter the uterine environment, which opment and survival. Asynchronous embryo transfer may influence the ability of lesser-developed lit- studies have indicated that porcine embryos become termates to either elongate or obtain sufficient uterine highly sensitive to the changing uterine environment space to survive (Pope et al., 1990; Xie et al., 1990b; during the blastocyst stage of development. The uterine Geisert et al., 1991). Estrogen release from elongating environment seems to support various stages of devel- conceptuses may induce a uterine secretory environ- opment up to d 6 to 7 of gestation. However, Polge ment that is not conducive to later elongation of less- (1982) demonstrated that pregnancy rates decrease developed littermates, resulting in their degeneration, with transfer of embryos 24 h behind in development as has been proposed by Pope et al. (1990). Certainly, with the recipient uterus and decrease to nearly zero the uterine environment is changed with the synthesis if they are 48 h behind. Rapid embryo deterioration of estrogen and elongation of the conceptuses through- within 24 h occured in d-6 embryos transferred to a d- out the uterine horn (see Geisert and Yelich, 1997). 8 uterine environment (Geisert et al., 1991). Thus, if Although this seems to be a very plausible hypothesis, embryos are greatly behind in development, the uterine a degree of caution must be taken because data have environment established through corpus luteum pro- indicated that administration of estrogen to pregnant gesterone stimulation can have a negative influence gilts prior to conceptus elongation does not interfere on survival. with development on d 11 and 12 of pregnancy (Morgan E58 Geisert and Schmitt et al., 1987a,b; Geisert et al., 1991; Cardenas et al., rich source for growth factors such as insulin-like 1997). However, administration of estrogen prior to con- growth factor (IGF)-I and -II, epidermal growth factor, ceptus elongation does act as an endocrine disruptor leukemia inhibitory factor, KGF, and connective tissue and affects subsequent survival of embryos after d 16 growth factor (see Geisert and Yelich, 1997; Ball et al., of pregnancy (Long and Diekman, 1986; Pope et al., 1998; Ka et al., 2000), most of for which their ligand 1986b; Morgan et al., 1987a). The loss of embryos seems receptor is expressed in the developing pig conceptus to be an effect of estrogen on the uterine environment (Geisert and Yelich, 1997; Ka et al., 2001). The reported and epithelial glycocalyx rather than a direct effect on decrease in uterine protein secretion in Meishan com- the conceptus (Blair et al., 1991); estrogen treatment pared to commercial pigs (Youngs et al., 1994; Vallet on d 12 of pregnancy does not interfere with embryo et al., 1998) would support a uterine role in modulating survival (Geisert et al., 1991). conceptus growth. However, the reciprocal conceptus Although uniformity in embryonic development is an transfer study of Youngs et al. (1994) indicated that, attractive model, it does not seem to be the sole basis in addition to the uterine environment, genotype of the for increased prolificacy of the Chinese Meishan pig conceptus is also involved with preimplantation (Wilmut et al., 1992; Ford, 1997). The advantage of the growth, which links back to the previous conceptus Ped Meishan embryos in litter size seems to be achieved gene discussion. Increasing our understanding of the through modulating placental length that can be gener- factors modulating conceptus trophoblast elongation ated following the initial elongation of the conceptuses and placental angiogenesis following placental attach- coupled with an increase in placental vascularity (see ment to the uterine surface will become increasingly Ford, 1997; Biensen et al., 1999). Meishan embryos important for future attempts to improve competition have similar numbers of inner cell mass cells but con- among embryos within the pig uterus. tain fewer trophectoderm cells and produce smaller Progress has been made in determining changes in amounts of estrogen compared to contemporary York- gene expression during the process of conceptus elonga- shire embryos (Anderson et al., 1993; Rivera et al., tion (Yelich et al., 1997a,b; Wilson et al., 2000); how- 1996; Kaminski et al., 1997). These two key factors ever, the key(s) to triggering the process or understand- would effectively restrict the surface area occupied by ing the pathway is far from established at this time. each conceptus following expansion of the trophoblast Recent studies have suggested the presence of a positive and reduce the effects of estrogen on neighboring lit- feedback loop for conceptus growth (Green et al., 1995); termates. The high concentrations of estrogen from the the conceptus seems to stimulate endometrial release developing Yorkshire gilts is not necessary to support of IGF and KGF into the uterine lumen during the development; a 57% reduction in conceptus estrogen developmental period preceding conceptus elongation synthesis with aromatase inhibitors did not affect early (Geisert et al., 2001; Ka et al., 2001). Treatment of gilts conceptus development (O’Neill et al., 1991). So, does with estrogen near the time of trophoblast elongation the lower estrogen synthesis by the developing concep- increases uterine protein and can advance the initiation tuses lead to greater survival? Although exogenous es- of the cellular remodeling process (Cardenas et al., trogen seems to have an obvious effect on embryo sur- 1997). Ka et al. (2001) have demonstrated that concep- vival, there is a question as to whether even the higher tus estrogen release during pregnancy increases uter- amounts of estrogen produced by Yorkshire conceptuses ine epithelial KGF expression, which in turn stimulates can have more than a local effect on the uterine micro- proliferation and differentiation of conceptus trophecto- environment that it contacts. derm. The known role of KGF in stimulating epithelial The inability of the sow to maintain a unilateral preg- proliferation, migration, and cellular differentiation nancy early in pregnancy (Anderson, 1966) and the (Rubin et al., 1995) makes the interactions between need for at least two embryos in each uterine horn IGF, estrogen, and KGF an attractive model for concep- to establish pregnancy in the pig (Polge et al., 1966) tus growth and remodeling. Interrelationships between suggests that estrogen does not diffuse throughout the these factors can explain conceptus growth regulation uterine horn easily. The presence of sulphotransferase by the uterus and the conceptus itself, as previously in the porcine endometrium effectively conjugates and discussed. These data would also lead to the suggestion inactivates estrogens moving through the endometrium that the more developmentally advanced, estrogenic (Pack and Brooks, 1974). Thus, although effects of estro- conceptuses gain an even more competitive advantage gen cannot be discounted, the Meishan embryos’ slowed in enhancing trophectoderm growth. rate of development (Youngs et al., 1993) and uterine inhibitory effect on embryonic development (Youngs et Is Embryonic Uniformity the al., 1994) have provided possible clues for survival Model for Improving Litter Size? through restriction of conceptus elongation and devel- opment of a more vascular placenta, as proposed by Given all the information on conceptus growth and Ford (1997). Determination of how the decrease in development, one must ask whether conceptus unifor- trophectoderm cell number is regulated either in the mity is an important factor contributing to litter size conceptus or through uterine secretion needs to be eval- in commercial swine herds. Theoretically, in the perfect uated. On d 10 to 12 of gestation, the pig uterus is a pregnancy, embryo uniformity would be the most desir- Early embryonic survival in the pig E59 able situation for the pig. This would require that the horn following fertilization, the restricted conceptuses number of ovulations did not exceed uterine capacity, reach the tubular stage of development but become de- all oocytes ovulated were at the same maturity and formed and do not elongate (J. P. Harney and F. W. viability, fertilized synchronously, and had the same Bazer, personnel communication). Thus, during the genetic potential for rate of development, and equidis- process of trophoblast elongation, some conceptuses tant uterine spacing occurred. Asynchrony with the that are lodged between two elongated embryos most uterine environment would not be a problem because likely will not undergo elongation and subsequently the time of ovulation regulates when the initial increase become deformed in development. Wu et al. (1988) indi- in luteal progesterone secretion occurs to stimulate the cated that an embryo requires more than 20 cm of uter- uterine secretory program for conceptus development. ine horn length to survive after the 7th wk of gestation. In this paradigm, ovulation rate should not exceed uter- Therefore, although there will be viable embryos that ine capacity because this could in theory lead to uterine develop to d 30 of gestation, they may not have sufficient crowding and loss of the entire litter if no mechanism placental surface area to survive to term. for having a competitive advantage between embryos So how are consistent large litter sizes achieved? Be- existed. It is difficult enough to control any one of the cause some embryonic loss is inevitable, the ovulation aforementioned factors, which explains our inability to rate needs to be high enough to overcome the loss of rapidly improve litter size. With this thought in mind, early defective embryos but provide a developing cohort it would be remiss not to acknowledge the thoughtful of 15 to 16 embryos that can elongate relatively synchro- and insightful paper Dzuik (1987) published 14 yr ago nously. Total synchrony of ovulation is not important addressing the same question. Dzuik (1987) champi- as long as cohorts of 15 to 16 quality oocytes are present oned that asynchrony in conceptus development is actu- at fertilization (see Pope et al., 1990). The majority of ally a survival advantage in the pig and that we may the three to four lesser-developed embryos would either pay too much attention to embryonic loss rather than be eliminated (fail to elongate) or, depending on individ- focusing attention on why some embryos survive. A ual positioning of embryos within the horn, fill in space limited amount and degree of nonuniformity between if available. Positioning of conceptuses in relation to groups of developing porcine embryos is actually an one another within the uterus is still an important issue attractive model. When ovulation rate does not exceed to survival (Dzuik, 1987). Embryo survival in this fe- uterine capacity, the majority of embryos that are devel- male would depend on the ovulation rate, number and opmentally competent to survive through d 8 will elon- quality of embryos developing in the uniform cohort, gate despite some variation in development rate. Even and the uterine capacity of the dam, which leads back if these embryos are somewhat variable in develop- to the traits of Meishan embryos. What are the traits ment, they should have less difficulty in acquiring suf- of the embryos from more contemporary sows developed ficient uterine space for placental development. The with the index selection for ovulation rate and embry- number of conceptuses present on d 12, not conceptus onic survival (Johnson et al., 1999) that have increased uniformity, directly affects litter size in this female. If litter size? Is there balance in ovulation of a cohort of ovulation rate exceeds uterine capacity in a female, quality embryos and(or) increase in placental effi- then there will be a number of possible scenarios for the ciency? Although a larger, more complete study is potential outcome in litter size. There could be greater needed, Wilson et al. (1999) suggested that selection early (before d 8) loss of embryos through ovulation of based on placental size and efficiency could improve immature and(or) defective oocytes and late-ovulating litter size at term. If placental size and vascularity follicles that places a number of developing embryos in are the important modulators of improved efficiency, jeopardy of being asynchronous with the uterine envi- research is needed to determine how to effectively de- ronment. Loss of these early embryos could normalize tect sows with increased placental efficiency or deter- the space available for the surviving conceptuses de- mine methods to regulate conceptus and uterine devel- pending on the number developing to d 12 and set in opment in pigs. motion the competition related to rate of growth and elongation. This female would be limited by both uter- Attempts to Improve Embryonic Survival ine capacity and the number of conceptuses initiating elongation, which makes for a variable range in possible The multitude of studies attempting to improve litter outcomes with litter size. For example, if 18 viable em- size through selective time of breeding, regulation of bryos were present on d 12 and they all elongated at ovulation, feed intake, steroid therapy, and nutritional the same time, crowding would cause fetal survival supplementation have been previously reviewed (Chris- problems after d 30 of gestation, depending on the indi- tenson, 1986; Dzuik, 1987; Ashworth, 1994; Pope, 1994; vidual placental surface area providing nutrients for Foxcroft, 1997). For the most part, studies evaluating each fetus. If 10 to 12 embryos elongated first, they nutritional supplementation have suffered from the would have space available to develop but could restrict lack of sufficient animal numbers and have failed to lesser-developed littermates from either elongating or either clearly demonstrate improved fertility or have having sufficient placental space to develop to term. If had no effect (Foxcroft, 1997). Certainly the presence a ligation is placed 15 cm from the tip of the uterine and important role of uterine factors such as riboflavin, E60 Geisert and Schmitt uteroferrin, retinol-binding protein, and folate-binding It should seem obvious that if we want more uterine protein (Malathy et al., 1990; Vallet et al., 1996, 1999) space, we need to select for pigs with longer horns. fueled the numerous nutritional supplementation trials Uterine length seems to be a limiting factor to litter attempting to increase embryonic survival (see Dzuik, size when ovulation rate increases (Wu et al., 1987). 1987; Ashworth, 1994; Foxcroft, 1997). Given that the However, although variation in uterine horn length ex- uterine environment of pigs on a nutritionally balanced ists, a difference between commercial breeds and Meis- diet is not significantly altered by additional nutrients han pigs is not apparent (see Ford, 1997). Gama and or vitamins and that it is difficult to affect conceptus Johnson (1993) also indicated that there was no signifi- developmental rate, it is not surprising that studies cant change in uterine dimensions of cyclic gilts follow- have failed to demonstrate consistent, if any, beneficial ing eight generations of selection for litter size. If the effects. Indeed, embryo survival is not significantly af- data do not indicate variation in length of the uterine fected in gilts subjected to inanition for the first 40 horns as a major factor in embryo survival, what uterine d of gestation (Anderson, 1975). Problems in growth, parameter(s) would affect placental surface area neces- survival, and maintenance of pregnancy occur if the sary for embryo survival? The level of endometrial fold- period of inanition is increased to greater than 40 d, ing, number of uterine glands, and uterine capillary but pregnancy is maintained if the females are supple- bed density within the endometrium are likely candi- mented with daily treatments of progesterone and es- dates that cannot be easily evaluated in the live animal. trogen. Porcine endometrium is not a flat, smooth surface but Improvements in embryo survival have been noted contains many macroscopic folds that branch off into when gilts are treated with retinyl-palmitate prior to numerous primary and secondary ridges to increase the time of ovulation (Coffey and Britt, 1993; Whaley surface for the placenta (Bjorkman et al., 1981; Dan- et al., 1997). An increase in embryo survival was attrib- tzer, 1984, 1985; Keys and King, 1990). Density of endo- uted to advancing resumption of meiosis and improving metrial folding cannot be accounted for by only measur- embryo quality rather than to any direct effect through ing horn length because this folding, along with uterine altering uterine function (Whaley et al., 2000). Feeding growth, provides for the expansion of the uterus after above maintenance requirement before mating also im- d 18 of gestation (Wu et al., 1988). A key component for uterine survival in all species, especially true for proved blastocyst cell numbers and reduced size vari- epitheliochorial-type placentation in the pig, is nutrient ability within litters (Ashworth et al., 1999). Cardenas passage to the placenta established through capillary et al. (1997) indicated that treatment of gilts with tes- blood flow within the endometrium (Ford, 1995). Alter- tosterone from d 13 of the estrous cycle until the follow- ation of blood flow to the uterus is evident at the time ing estrus improved blastocyst survival, possibly of conceptus elongation and during expansion of the through improved oocyte quality. These data are consis- allantois to fill the uterine horn between d 15 and 30 tent with the previous report of Hunter et al. (1993), of gestation (Ford, 1995). The conceptus induces an who indicated that oocytes in Meishan gilts are more inflammatory response to enhance nutrient flow into mature and uniform than Large-White gilts. Thus, in- the uterine lumen (Keys and King, 1988, 1995). The formation points to the importance of increasing the presence of kinin β2 receptors in the endometrium and number of developmentally mature oocytes before we the conceptus-induced release of kinins (Allen et al., even progress to the uterine environmental modulation 2002), potent regulators of blood pressure, indicates of conceptus development. the active participation of the conceptus in modulating So what can be done to improve the interaction be- maternal blood flow to the porcine placenta. Although tween conceptus and uterine environment? Obviously, from a different species and type of placentation, vascu- understanding how placental length and vascularity lar casts of the uterus and maternal cotyledons (Figure are modulated will help direct us to studies that may 1) in Florida native ewes compared to the more prolific alter early development of the conceptus. Development Blackfaced ewes (two to three lambs) illustrates the of differential-display PCR, PCR-based cDNA subtrac- alteration in vascular density that can be developed to tion, and the forthcoming production of microarrays support multiple fetuses in the prolific breed of sheep will accelerate identification of genes involved with con- (F. W. Bazer, F. F. Bartol, and D. H. Barron, unpub- ceptus and uterine development. Information concern- lished data). The vascular networks of the maternal ing genes involved with conceptus and uterine develop- uterus and fetal placenta in the pig are complementary ment may improve our search for and use of genetic to one another and become more complex as gestation markers for reproductive efficiency as the pig genome proceeds (Lesier and Danzter, 1988). Many angiogenic map is completed (Rothschild et al., 1997; Linville et factors are involved with regulating endometrial and al., 2001). Technologies in transgenesis and cloning are placental vascular growth (see Reynolds and Redmer, attractive methods to develop increased reproductive 1995). However, we currently do not have a clear under- efficiency in swine but have been rather difficult to standing of the regulation of angiogenesis during preg- develop, and the low heritability of litter size will still nancy in the pig. not provide the rapid increase that most might expect The superficial attachment and placentation of the (Haley et al., 1988). pig dictates the critical need for endometrial generation Early embryonic survival in the pig E61

Figure 1. Vascular casts of the maternal cotyledons from a single-lamb-bearing Florida native ewe (B, C) and a prolific, multilamb-bearing Blackface ewe (A, D). of uterine secretions. The importance of glandular sup- is involved in the endometrial gland remodeling and port for the developing porcine conceptus has been secretory function in the pregnant ewe. Uterine infu- made even more evident by the failure of early ovine sion of growth hormone and placental lactogen in- conceptuses to elongate and attach in uterine gland creases uterine glandular development and secretory knockout ewes (Gray et al., 2001a,b). Prolonged neona- function in steroid-treated, ovariectomized ewes. It is tal exposure of ewes to progesterone ablates uterine also possible to treat neonatal lambs with prolactin and gland morphogenesis and absence of glands in the adult increase the number of uterine glands in the adult (T. (Bartol et al., 1999). Thus, it is clear that neonatal E. Spencer, personal communication). The presence of exposure to steroids or other endocrine disruptors can prolactin receptors in the porcine endometrium (Young alter uterine function in the adult. In the pig, uterine et al., 1990) suggests that it may be possible to influence glands are absent at birth and uterine adenogenesis in the number of uterine glands in the pig. With the known the pig occurs from birth to 21 d postnatally (Bartol et effects of estrogen and prolactin on uterine and glandu- al., 1993; Tarleton et al., 1999). Tarleton et al. (1999) lar development in females, uterine alterations and lit- demonstrated that estrogen stimulates but treatment ter capacity in pigs need to be evaluated following neo- with the estrogen antagonist ICI 182,780 inhibits aden- natal treatment with estrogen and prolactin. ogenesis in the neonatal uterus. Therefore, it may be Over the past 40 yr of probing the complexity of em- possible to alter neonatal uterine glandular morpho- bryonic development and loss, we have developed more genesis and development through steroidal and(or) lac- answers for why embryonic loss occurs but provided togenic hormone administration. Spencer et al. (1999) little in the solution for rapidly improving litter size in proposed that sequential exposure of the endometrium the pig. Improvements are not likely to result from to steroids and lactogenic and somatogenic hormones any nutritional or steroid therapies aimed at changing E62 Geisert and Schmitt uterine function during gestation but could result foundation for subsequent fertility. J. Reprod. Fertil. Suppl. through better control of ovulation and oocyte quality 53:284–300. Bartol, F. F., A. A. Wiley, T. E. Spencer, J. L. Vallet, and R. K. or changes in the uterine vascular and secretory capac- Christenson. 1993. Early uterine development in pigs. J. 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Wu, L., F. Honghui, and C. M. Warner. 1999. Identification of two transforming growth factors during rapid trophoblastic elonga- major histocompatibility complex class Ib genes, Q7 and Q9, tion in the porcine conceptus. Biol. Reprod. 57:126–134. as the Ped gene in the mouse. Biol. Reprod. 60:1114–1119. Yelich, J. V., D. Pomp, and R. D. Geisert. 1997b. Ontogeny of elonga- Wu, M. C., M. D. Hentzel, and P. J. Dzuik. 1987. Relationships tion and gene expression in the early developing porcine concep- between uterine length and number of fetuses and prenatal tuses. Biol. Reprod. 57:1256–1264. mortality in pigs. J. Anim. Sci. 65:762–770. Young, K. H., R. R. Kraeling, and F. W. Bazer. 1990. Effect of Wu, M. C., W. J. Shin, and P. J. Dzuik. 1988. Influence of pig embryos pregnancy on exogenous ovarian steroids on endometrial pro- lactin receptor ontogeny and uterine secretory response in pigs. on uterine growth. J. Anim. Sci., 66:1721–1726. Biol. Reprod. 43:592–599. Xie, S., D. M. Broermann, K. P. Nephew, M. D. Bishop, and W. Youngs, C. R., L. K. Christenson, and S. P. Ford. 1994. Investigations F. Pope. 1990a. Relationship between oocyte maturation and into the control of litter size in swine: III. A reciprocal embryo fertilization on zygotic diversity in swine. J. Anim. Sci. transfer study of early conceptus development. J. Anim. Sci. 68:2027–2033. 72:725–731. Xie, S., D. M. Broermann, K. P. Nephew, R. D. Geisert, and W. F. Youngs, C. R., S. P. Ford, L. K. McGinnis, and L. H. Anderson. Pope. 1990b. Ovulation and early embryogenesis in swine. Biol. 1993. Investigations into the control of litter size in swine: I. Reprod. 43:236–240. Comparative studies on in vitro development of Meishan and Yelich, J. V., D. Pomp, and R. D. Geisert. 1997a. Detection of tran- Yorkshire preimplantation embryos. J. Anim. Sci. 71:1561– scripts for retinoic acid receptors, retinol binding protein, and 1565. Uterine capacity in the pig reflects a combination of uterine environment and conceptus genotype effects1,2

S. P. Ford3,5, K. A. Vonnahme3, and M. E. Wilson4

Department of Animal Science, Iowa State University, Ames 50011

ABSTRACT: Prenatal losses in U.S. pig breeds range selection for either longer uterine horns or for a reduced from 30 to 50%, of which greater than 75% occurs before conceptus size should potentially increase litter size d 30 of gestation and is thought to result from littermate in the pig. Researchers have evaluated the impact of asynchrony. Numbers of embryos can be experimen- differences in prepubertal uterine horn length on subse- tally increased to d 30 using superovulation (excess ova quent uterine capacity using a unilateral hysterectomy- shed) and superinduction (transfer of embryos to an ovariectomy model but have had modest and variable already pregnant uterus); however, these females far- success in increasing litter size at farrowing. In con- row the same number of pigs as untreated controls. trast, results from our laboratory suggest that placental These data demonstrate that between d 30 and parturi- size is moderately heritable and results in consistent tion there are significant additional periods of concep- increases in litter size of two to three pigs in the York- tus loss, which has led to the conclusion that uterine shire breed with little impact on pig birth weight or capacity (i.e., the number of conceptuses a sow uterus neonatal viability. This selection of pigs for smaller and can accommodate) is the major limitation to litter size relatively more efficient placentae (i.e., the number of in the pig. The special importance of uterine capacity grams of fetus that can be supported by1gofplacenta) in the pig may result from the noninvasive epithelio- seems to provide a useful method for increasing litter chorial type of placentation in this species, making the size in the pig. A careful evaluation of the physiologic surface area of attachment between the placenta and and genetic differences of conceptuses with differing endometrium a limiting factor. In devising selection placental efficiencies is necessary if we are to determine schemes for this trait, one could logically conclude the specific factors affecting litter size in the pig.

Key Words: Litter Size, Placenta

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E66–E73

Introduction produce and secrete significant amounts of estrogen into the uterine lumen, which results in endometrial Uterine capacity is defined as the number of concep- secretion of a myriad of growth factors such as IGF- tuses that the pig uterus can successfully carry to I, which could affect subsequent uterine as well as term. Data suggest that limitations of uterine capacity conceptus growth and development (Wilson and Ford, are set early in gestation, possibly around the time of 1997). At this time, littermate conceptuses space conceptus elongation on d 11 through 13 (Figure 1). evenly throughout the uterus (Dhindsa et al., 1967; It is during this time that the conceptus begins to Dziuk, 1968, 1985) and expand asynchronously from 1-cm spherical blastocysts to filamentous forms of variable length (Geisert et al., 1982), in association with a 20 to 30% conceptus loss in U.S. and European 1Journal paper no. J-19264 of the Iowa Agric. and Home Econ. Exp. pig breeds (Dziuk, 1968, 1987; Pope, 1994). Uterine Sta., Ames, Iowa, project Nos. 0239, 3505, and 3900, and supported by capacity begins to measurably affect litter size after Hatch Act and State of Iowa funds. 2This reasearch was also supported in part by USDA-NRICGP d 30 of gestation in the pig (Fenton et al., 1972; Pope awards no. 97-35205-5077 and no. 00-02156-5966. et al., ‘971; Huang et al., 1987), when a real competi- 3Current address: Dept. of Ani. Sci., Univ. of Wyoming, Box 3684, tion for limited uterine space and nutrients by lit- Laramie, WY 82071-3684. termates becomes increasingly critical. 4Current address: Division of Anim. and Vet. Sci., West Virginia Uterine capacity can be defined in terms of the rela- Univ., Morgantown, WV 26506. tive surface area of placental endometrial attachment 5Correspondence: Rm 113 Animal Science Building (phone: 307- 766-2709; fax: 307-766-2355; E-mail: [email protected]). required to support the nutrient requirements of an Received April 5, 2001. individual fetus throughout gestation. Recent evi- Accepted August 15, 2001. dence suggests that placental efficiency (PE; fetal wt/

E66 Conceptus-uterine interactions E67 trol line. The authors speculated that as ovulation rate increased in the selection line uterine capacity increasingly became the limiting factor in prenatal survival after d 50 of gestation. The authors went on to speculate that selection on component traits of litter size would have been more effective if in addition to ovulation rate a measure of uterine capacity had also been included, as previously suggested by Bennett and Leymaster (1989). Cassady et al. (2001) attempted to identify chromo- somal regions harboring QTL for reproductive traits in pigs using a reference population created by Rathje et al. (1997) at the University of Nebraska. Grandpar- ents of this population had been selected for ovulation rate and embryonic survival. While reporting evidence for QTL for a number of reproductive traits (including ovulation rate, age at puberty, number of teats, and number of fully formed pigs) he concluded that few, if any, major genes for reproductive traits were segregat- ing in the population. He went on to conclude that little evidence exists to encourage marker-assisted selection at the present time. This conclusion is consistent with results recently reported by Linville et al. (2001), who used the candi- date gene approach to determine whether specific loci explain the observed differences in litter size. They found no associations between six polymorphic mark- Figure 1. A time line demonstrating the time periods ers in two lines of pigs, again originating from the of conceptus loss in the pig. selected and control lines described by Johnson et al. (1999). In fact, for several of these markers, the esti- mated effect of purported favorable alleles was nega- placental wt) is an individual conceptus trait and is tive. They reported that these selection lines exceeded highly variable within a litter (Wilson et al., 1999). the control line by 20 to 50% in both ovulation rate Impacts of selection for uterine length (Christenson and litter size. The authors went on to explain the et al., 1990), ovulation rate (Johnson et al., 1999), and rationale for the selection of the six specific markers placental efficiency (Wilson et al., 1999) have shown chosen, on an individual basis, but, as Cassady et al. variable success in increasing litter size. In this paper, (2001) concluded for this same population, selection we will attempt to summarize the various approaches for ovulation rate, embryonic survival, and litter size used to increase uterine capacity, both from the mater- seems to have acted on multiple loci, each of which nal and fetal sides, and the impacts they have had on exerts only a modest effect (i.e., these are polygenic litter size. traits). Although there are many possible reasons for this lack of association reported by Linville et al. Genetic Selection (2001) between selected markers and litter size, a fail- ure to understand what specific physiologic differences Johnson and co-workers at the University of Ne- were selected for in this population suggests that little braska have used genetic selection in an attempt to progress should be expected using only this approach. increase litter size in the pig, but with limited success. In the next section, we will discuss the findings of Johnson et al. (1999) conducted 11 generations of se- several researchers who are attempting to understand lection (from a Large White and Landrace genetic the necessary interactions between the conceptus and base) for increased index of ovulation rate followed by the uterus that promote optimal conceptus survival three generations of selection for litter size. By the through alteration of uterine capacity. 11th generation of selection they verified a selection advantage of 7.4 ovulations and 3.3 fetuses on d 50 Conceptus-Uterine Interaction but only realized an advantage of 1.1 live pigs at far- rowing. The response to selection at generation 14 was Evidence suggesting that uterine capacity is lim- an increase of 1.4 live pigs/litter; however, the number iting to litter size after d 30 of gestation is based of stillborn pigs increased and pig birth weights de- largely on studies using superovulation (excess num- creased. As a result, the number of pigs weaned actu- ber of ova shed), superinduction (transfer of embryos ally declined in the selected line compared to the con- to an already pregnant uterus), and unilateral hyster- E68 Ford et al. ectomy-ovariectomy (UHO) models. Superovulation 1948). From d 35 to 70 of gestation, the surface area and superinduction increase the number of concep- of the chorioallantoic membrane increases rapidly tuses surviving to d 30 (percentage of embryo survival (Knight et al., 1977). This results from the prolific is similar); however, these females do not farrow lit- folding of the placental surface into permanent folds ters larger than controls (Fenton et al., 1972; Monk in the endometrium, followed by the development of and Erb, 1974; Webel and Dziuk, 1974). Females sub- an interlocking network of placental and endometrial jected to UHO ovulate normal numbers of ova, but microvilli, which function to further expand the pla- only farrow approximately one-half the number of pigs cental:endometrial surface area of exchange (Friess et of intact controls (Christenson et al., 1987). The UHO al., 1980; Bjørkman and Dantzer, 1987). From d 70 model has been used to estimate uterine capacity per through approximately d 90 there is little noticeable uterine horn in the absence of any possible limitation change in placental surface area (Knight et al., 1977), in ovulation rate (Christenson et al., 1987). In these but from d 90 of gestation through term as much as studies, it was concluded that UHO gilts that had the a doubling of placental surface area is seen (Wigmore largest litters (> 1/2 of control gilts) at farrowing may and Strickland, 1985; Biensen et al., 1998). provide the most optimal uterine environment (i.e., As previously stated, the limitations of uterine ca- increased uterine capacity. Following this rationale, pacity begin to affect conceptus survival after d 30 and Gama and Johnson (1993) used UHO in lines of gilts are associated on d 35 to 40 with a marked and abrupt at the University of Nebraska that had been selected increase in the surface area of placental exchange due for differences in ovulation rate, embryo survival to d to the development of microscopic interdigitations be- 50, and litter size. Pregnant gilts were slaughtered tween the placenta and uterine luminal surface re- on d 93 through 100 of gestation and ovulation rate, ferred to as primary rugae (Bjørkman and Dantzer, number of fully formed pigs, and number of mummied 1987; Leiser and Dantzer, 1988; Leiser and Dantzer, pigs were determined. As previously reported by 1994). During the last third of gestation, when another Christenson et al. (1987), uterine capacity was esti- period of observed fetal loss occurs (Christenson et al., mated as twice the number of fully formed fetuses in 1987; Johnson et al., 1999), the functional surface area the remaining horn. When this was accomplished, the for nutrient and waste product exchange is again ac- estimated difference between individuals selected for celerated in association with rapid fetal growth by the increased litter size and the randomly selected con- development of secondary rugae, an additional tier of trols was 0.66 ± 1.28 pig. Although this increase was interdigitation along the primary rugae (Friess et al., not significant, the authors suggested that these data 1980; Bjørkman and Dantzer, 1987). indicate that additive genetic variance for uterine ca- In addition to the progressive increases in the func- pacity does exist in swine. tional placental:endometrial surface area allowing fe- In the pig, uterine capacity may specifically relate tal-maternal exchange, the vascularity of placental to epitheliochorial and diffuse placentation, whereby and adjacent endometrial tissues also changes with the chorionic epithelium is in direct contact with the the advancement of gestation, in conjunction with a uterine luminal epithelium (Grosser, 1933; Benirchke, progressive decrease in the diffusion distance between 1983; MacDonald and Bosma, 1985). This placental the placental and fetal capillaries. Vonnahme et al. type, which is appositional rather than invasive and (2001) demonstrated that in the white composite relies largely on simple diffusion, requires an ade- breeds, density of placental blood vessels per unit sur- quate surface area for nutrient exchange from the ma- face area tissue (vascular density) actually decreased ternal to fetal blood streams. Between d 20 and 30 of during the first rapid expansion of the placenta from gestation, there is a rapid increase in placental length d 25 through 44, then increased progressively from d that slows asymptotically, reaching a plateau around 44 through 90 before reaching a plateau until term d 70 before increasing again after d 100 (Pomeroy, (Figure 2). Of interest is the fact that vascular density 1960; Knight et al., 1977). From d 30 to 40, the pla- did not increase during either the early or late periods centa progresses from a very simply organized struc- of increased fetal death. Further, Vallet et al. (1996) ture, consisting of a thin membrane, hyaluronic acid and Pearson et al. (1998) have suggested that fetal gel, and a rapidly proliferating system of blood vessels erythropoiesis is negatively affected by intrauterine to a more complex structure (Perry, 1981; Bazer, crowding and may play a role in fetal loss. Vonnahme 1989). Also between d 30 and 35 of gestation, the yolk et al. (2001) also reported that placental vascular en- sac rapidly regresses, and the responsibility for the dothelial growth factor (VEGF), a potent angiogenic absorption of oxygen and nutrients and the removal and permeability-enhancing factor, followed the same of carbon dioxide and waste is shifted to the highly pattern as that of placental vascular density through- vascularized chorioallantoic membrane (Bjørkman, out gestation and was positively correlated with both 1973; Tiedemann and Minuth, 1980; Dantzer, 1985). placental (r = 0.34; P < 0.05) and adjacent endometrial Placental capillaries ramify throughout the entire (r = 0.34; P < 0.05) vascular density changes of individ- surface of the placenta in close contact with the chori- ual placentae (see Vonnahme et al., 2001 for technical onic epithelium, except for the ends, which are nonvas- details). Increased permeability at the placental:endo- cular and are referred to as necrotic tips (Patten, metrial interface with advancing gestation was sug- Conceptus-uterine interactions E69 dependent on an adequate surface area of functional contact between the placenta and uterine wall if the fetus is to exhibit optimal growth and development.

Placental Efficiency

To avoid the confounding effects of differences in ovulation rate and uterine length, both of which have been reported to have impacts on litter size, we chose to compare and contrast the reproductive physiology of the prolific Chinese Meishan gilt (two to five postpu- bertal estrous cycles) with Yorkshire gilts of similar reproductive age. Although Meishan and Yorkshire gilts at this reproductive age exhibit similar ovulation rates (16.5 ± 5 and 16.3 ± 0.4, respectively) and uterine lengths, the litter size of Meishan gilts was markedly greater (12.3 ± 0.4 vs 8.9 ± 0.4 live pigs/litter; Ford, 1997). Further, the Meishan and Yorkshire gilts in our herds ovulated at the same time after the initiation of estrous activity, as evidenced by the recovery of embryos at similar stages 48 to 54 h after estrus onset (one to eight-cell embryos). From d 5 to 12 of gestation, however, Meishan conceptuses are smaller than York- shire conceptuses due to a selective reduction in the number of trophectoderm cells, the cell type con- taining the rate-limiting enzyme for conceptus estro- gen synthesis (Youngs et al., 1993; Rivera et al., 1996; Wilson and Ford, 1997). Further, we observed that Figure 2. (a) Changes in vascular endothelial growth although both Meishan and Yorkshire conceptuses ini- factor mRNA and (b) the relative number of blood vessels tiated estrogen secretion and elongated on the same per unit area of placental tissue throughout gestation in days of gestation (d 11 to 12), Meishan conceptuses the pig. Means and SEM with different superscripts secreted less estrogen into uterine luminal fluid and < within a measurement differ (P 0.05) (reproduced with were smaller (Anderson et al., 1993). The decreased permission from Vonnahme et al., 2001). uterine luminal estrogen concentrations in Meishan vs Yorkshire females on d 11 to 12 was associated with reduced levels of total protein, calcium, oxytocin, and gested by Freiss et al. (1980, 1982), who demonstrated IGF-I in the same fluid (Ford and Youngs, 1993; Wil- that the functional maternal:fetal intercapillary dis- son and Ford, 1997; Vallet et al., 1998). This is consis- tance decreased from 15 to 20 ␮m to approximately 2 tent with the role of IGF-I in increasing trophectoderm ␮m with the advancement of gestation. mitotic rate in pig conceptuses (Lewis et al., 1992). In the pig, placental transfer of freely diffusable As a result, Meishan filamentous conceptuses were nutrients seems to rely on a vascular anatomy that is shorter and contained fewer cells than Yorkshire con- either concurrent or possibly crosscurrent (Friess et ceptuses on d 14 (Wilson et al., 1995). Further, Meis- al., 1982). Transported solutes such as glucose, amino han conceptuses exhibit markedly smaller placentae acids, ions such as sodium, potassium, calcium, and on d 30, 50, 70, 90, and 110 and at term (Hunter et. protons require transporter protein-mediated transfer al., 1994; Biensen et al., 1998; Wilson et al., 1998) (Sibley et al., 1997). In the pig, this transporter-medi- than do conceptuses of a variety of less-prolific pig ated transfer seems to be concentrated in the troughs breeds. This reduced placental size of the Meishan of the chorionic folds, which are adjacent to the ridges regardless of the number of conceptuses present seems of endometrial folds, and is generally not tied directly to be a direct result of a reduced preimplantation uter- to the rate of blood flow (Friess et al., 1980; Poston, ine exposure to estrogen. Injections of estrogen into 1997; Sibley et al., 1997). In addition to the transfer pregnant Meishan females around the time of concep- that occurs across the uterine lumenal epithelial-cho- tus elongation was found to markedly increase placen- rionic epithelial interface, there is also transfer of tal size at term (Wilson and Ford, 2000). large macromolecules such as uteroferrin, which are The real breakthrough in our understanding of how produced by the uterine glands and then absorbed the Meishan fetus survived on such a tiny placenta intact by the placental areolae (Perry, 1981; Freiss et came when both Meishan and Yorkshire embryos were al., 1982; Roberts et al., 1986; Leiser and Dantzer, placed in the uterus of a Yorkshire recipient female 1994). All of these transfer mechanisms are, however, on d 2 and allowed to compete for survival (Wilson et E70 Ford et al. In a subsequent study (Wilson et al., 1999) using our Yorkshire population, pigs were matched to their placentae as described above, and we found that there was significant (threefold) variation in placental weights across females, with as much as twofold differ- ences exhibited within a single litter. From these lit- ters we then selected boars and gilts with birth weights ≥ 1,250 g that had higher than average PE and those that had lower than average PE. Although pig birth weights of the high-PE group were similar to those of the low-PE group, their placentae were markedly smaller and more vascular. We then allowed these pigs to reach puberty and at the second estrus we bred gilts of each group (high PE or low PE) to boars of the same group and collected farrowing data from parities 1 and 2. In both parities, the high-PE females farrowed more (P < 0.05) live pigs than the Figure 3. Similar-sized Meishan and Yorkshire lit- low-PE group (12.5 ± 0.7vs9.6± 0.5). Although the termates farrowed by a Yorkshire sow. high-PE group farrowed pigs that were about 20% lighter than the low-PE group (1.2 ± 0.1 vs 1.5 ± 0.1 kg, P < 0.05), their placentae were 40% lighter (250 ± al., 1998). At farrowing, the umbilical cord of each pig 10 vs 347 ± 15 g, P < 0.01), resulting in an increased PE was double-ligated and tagged so that all pigs could for pigs farrowed in the high-PE group. Interestingly, be matched with their placentae when they were ex- Vonnahme and Ford (2001) recently reported that se- pelled en masse after all pigs were farrowed. To our lection of Yorkshire pigs for high PE markedly in- = = surprise, Meishan (n 7) and Yorkshire (n 7) pigs creased placental VEGF mRNA compared to unse- ± exhibited similar birth weights (1,308 76 and 1,585 lected controls. Also of interest was the fact that num- ± 95 g, respectively), as a result of marked increases in bers of stillborn pigs and postfarrowing death losses Meishan pig birth weight compared to those farrowed were low and similar for both groups, suggesting no normally by straightbred Meishan females, which av- negative impact on pig viability. Further, when the erage ∼800 to 900 g (Figure 3). Further, all Meishan association between a pig’s PE at birth was compared pigs were gestated on very small and highly vascular to subsequent production traits in a commercial herd placentae, and all their Yorkshire littermates were of Yorkshire and Landrace females, no effect of PE gestated on much larger and less-vascular placentae was seen on 21-d weight, days to 105 kg, backfat depth, (209 ± 17 and 472 ± 46 g, respectively). At this point or loin muscle area (Biensen et al., 1999). These data we began employing PE as describing the grams of suggest that the selection for PE might be a valuable fetus that could be supported by a gram of placenta, tool for increasing litter size in commercially relevant as previously used by others (Molton et al., 1978; Kurz pig breeds without decreasing periparturient pig via- et al., 1999). Placental efficiency of Meishan pigs in bility. the litter averaged 6.25, and the PE of their Yorkshire littermates averaged 3.36. These PE differences result Validation of the Impacts of Ovulation Rate, from a 56% difference in placental weights and a 17% Uterine Size, and PE in a Commercial Swine Herd difference in pig weight. These data confirm that the PE is an individual conceptus trait independent of In a recent study (Wilson et al., 2000), we used a uterine environmental control. In subsequent studies, very large, highly productive (average litter size 10.4 we determined that changes in PE were a result of pigs/litter), and healthy herd in Iowa (Dayton Pork, changes in placental weight, not pig weight. Further, Dayton, Iowa; a portion of Swine Graphics Enter- there was much less variation in the weights and vas- prises, Inc., Webster City, IA) to gain insight into the cularity of the Meishan placentae within a litter than factors that limit litter size in commercial swine pro- exhibited by their littermate Yorkshire placentae (S. duction. For too long, researchers have used a few P. Ford, unpublished observations). Thus, we specu- university herds for their studies on litter size without lated that during the domestication of the Meishan comparing physiological differences in reproductive breed, which encompassed as many as 7,000 yr (Yun, function (i.e., ovulation rate, uterine length, placental 1988), an intense selection for litter size may have size, etc.) of their herds to those used by the swine resulted in an indirect selection for small, highly vas- industry, or for that matter other university herds. cular, and relatively uniform placentae. If this was Thus, many of the conclusions drawn from these stud- indeed the case, then we felt that there might be sig- ies may have limited relevance to today’s swine indus- nificant variation in this trait in our Yorkshire popu- try. In our attempt to understand the factors limiting lation. litter size, researchers have focused on three main Conceptus-uterine interactions E71 traits: 1) ovulation rate, 2) uterine size, and, more Based on the data reported here for this highly produc- recently, 3) placental efficiency. Although all three tive commercial herd, it is imperative that we define seem moderately heritable, selection for increases in the reproductive phenotypes of our research herds to ovulation rate and uterine length have resulted in make sure that the conclusions we draw are relevant very limited and highly variable increases in litter to the industry. This is especially critical in the use size, whereas recent and limited studies using selec- of the candidate gene approach, which is being increas- tion for placental efficiency have shown a much ingly promoted for the determination of whether spe- greater effect. cific loci contribute to an increased litter size. For this study (Wilson et al., 2000), a total of 190 Camborough line 6-02 (PIC) sows representing parit- Implications ies 1 to 14 were slaughtered on d 25 (n = 63), 36 (n = = 60), or 44 (n 67) of gestation, and the gravid uteri Data presented in this review suggest that more and associated ovaries were recovered. Uterine length than adequate numbers of viable embryos enter the and the number of ovulations were not different among uterus on d 2 or 3 of gestation, but due to an asynch- the three groups, averaging 434 cm and 26.6, respec- rony of development, 20 to 30% of conceptuses are lost tively. Further, there was no correlation between uter- by d 18. Additional periods of conceptus loss occur ine length or ovulation rate and the parity of the sow. between d 30 and 40 (15 to 20% loss) and during the Viable conceptus number decreased from 15.8 on d 25 last third of gestation (5 to 10%) as the competition (before the limitations of uterine capacity) to 12.9 on for limited uterine space becomes critical (i.e., uterine d 36 (after the impact of uterine capacity), then re- capacity). Uterine capacity seems to be increased by mained relatively constant through d 44 (12.1 fetuses). increasing uterine size (maternal effect), or alterna- This reduction in the number of viable conceptuses tively by decreasing placental size, with a correspond- under the limitation of uterine capacity was reflected ing increase in placental efficiency (individual concep- by a marked decrease in conceptus survival (number tus effect). If an optimal litter size is to be achieved, of conceptuses/number of corpora lutea), which de- future research must concentrate on the selection for creased from 60.2% on d 25 to 50.1% on d 36. reproductive phenotypes that is based on a thorough Although ovulation rate was highly and positively understanding of the physiological factors mediating correlated (P < 0.01) with the numbers of conceptuses successful conceptus-uterine interactions during present on d 25 (r =+0.50) before the limitations of these three critical periods of conceptus loss. uterine capacity were seen, by d 36 this association was lost (r =+0.02; P > 0.10). In contrast, length of uterine horns was not associated with the numbers of Literature Cited conceptuses found on d 25 (r =−0.03; P > 0.10), but these traits were positively correlated (P < 0.01) there- Bazer, F. W. 1989. Allantoic fluid: Regulation of volume and compo- after (d 36, r =+0.36; d 44, r =+0.40). Interestingly, sition. In: R. A. Brace, M. G. Ross, and J. E. Robillard (ed.) Reproductive and Perinatal Medicine. pp 135–155. Perinatol- although fetal weight was not associated with concep- ogy Press, Ithaca, NY. tus number on any of the days examined, placental Benirschke, K. 1983. Placentation. J. Exp. Zool. 228:385–389. weight was negatively correlated (r =−0.32; P < 0.01) Bennett, G. L., and K. A. Leymaster. 1989. Integration of ovulation with the number of viable conceptuses on all 3 d exam- rate, potential embryonic viability and uterine capacity into a ined. We have previously shown (Biensen et al., 1998) model of litter size in swine. J. Anim. Sci. 67:1230–1241. =+ Biensen, N. J., M. F. Haussmann, D. C. Lay, Jr., L. L. Christian, that placental weight is highly correlated (r 0.82; and S. P. Ford. 1999. The relationships between placental and P < 0.001) with placental surface area, regardless of pig birth weights and growth traits. Anim. Sci. 68:709–715. breed, sex, or day of gestation, and therefore is a good Biensen, N. J., M. E. Wilson, and S. P. Ford. 1996. Differential indicator of placental size. mechanisms regulating Meishan (M) and Yorkshire (Y) fetal These data suggest that larger litters on d 25, 36, growth. J. Anim. Sci. 74(Suppl. 1):215 (Abstr.). Biensen, N. J., M. E. Wilson, and S. P. Ford. 1998. The impact of and 44 are composed of conceptuses exhibiting fetuses either a Meishan or Yorkshire uterus on Meishan or Yorkshire of similar size, but with smaller placentae and thus fetal and placental development to days 70, 90, and 110 of exhibiting an increased PE. Further, the ovulation gestation. J. Anim. Sci. 76:2169–2176. rate of sows from this commercial herd was extremely Bjørkman, N. 1973. Fine structure of the fetal-maternal area of high compared to that reported by investigators for exchange in the epitheliochorial and endotheliochorial type of their research herds (Christenson, 1993; Ford, 1997; placentation. Acta Anat. 86(Suppl 1):1–22. Bjørkman, N. and V. Dantzer. 1987. Placentation. In: H.-D. Dell- Johnson et al., 1999) but was similar to that reported mann and E. M. Brown (ed.) Textbook of Veterinary Histology. by Foxcroft (1997) for another line of commercial sows. (3rd ed.) pp 340–360. Lea & Febiger, Philadelphia. Additionally, although conceptus number was posi- Cassady, J. P., R. K. Johnson, D. Pomp, G. A. Rohrer, L. D. Van tively related to ovulation rate on d 25, by d 36 the Vleck, E. K. Spiegel, and K. M. Gilson. 2001. Identification of limitations of uterine size began to reduce conceptus quantitative trait loci affecting reproduction in pigs. J. Anim. Sci. 79:623–633. number regardless of ovulation rate. These data sug- Christenson, R. K. 1993. 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W. L. Flowers2

Department of Animal Science, North Carolina State University, Raleigh 27695-7621

ABSTRACT: The influence of the number and qual- of spermatozoa from different boars to fertilize ova. A ity of spermatozoa inseminated on litter size in swine number of semen quality tests have been developed to is examined in this paper. There is evidence to support estimate the fertility of semen. Several of these have the following observations. Litter size varies among documented that increases in estimates in sperm qual- boars when insemination doses contain the same num- ity are associated with increases in litter size. However, bers of spermatozoa. Increasing the number of sperm the relative effectiveness of each of these for determin- inseminated generally has a positive effect on the num- ing the optimal number of spermatozoa that should be ber of pigs born alive, especially between the range of included in insemination doses remains to be eluci- 1to3× 109 cells. The manner in which litter size re- dated. In summary, increasing the fertilization rate of sponds to increasing the number of spermatozoa insem- boars should be possible by improving semen quality, inated varies among boars. These relationships be- increasing the number of spermatozoa inseminated, tween the number of sperm inseminated and the re- and adjusting using estimates of sperm quality to ad- sulting litter size provide credence to the idea that boars just number of sperm inseminated. However, the mag- exhibit unique fertility patterns. These divergent fertil- nitude of changes in litter size resulting from these ity patterns probably reflect variability in the ability strategies is likely to vary considerably among boars. Key Words: Boars, Litter Size, Spermatozoa

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E47–E53

Introduction is physiologically reasonable to assume that there are two basic characteristics that are directly responsible A spermatozoon that ultimately results in the birth for a boar’s influence on litter size: the number of sper- of a live piglet is transported from the cervix through matozoa inseminated and the proportion of these that the uterus and enters the uterotubal junction (Hunter, can successfully engage ova. The latter of these is often 1990). At this point it attaches to an oviductal cell until referred to as the quality of spermatozoa and can be it is released and, eventually, encounters an ovum after estimated in a variety of different ways, including the ovulation (Sirard et al., 1993). During interaction with monitoring of several physical and biochemical traits the ovum, a spermatozoon must bind to and then move that allow spermatozoa to fertilize ova. Whether or not increases in the number of spermato- through the zona pellucida (Miller, 2000). This allows zoa can compensate for decreases in sperm quality is it to enter the perivitelline space and fuse with the a question that is often posed but has yet to be resolved. plasma membrane, which eventually leads to the union This is due to the fact that most studies have examined of its genetic information with that of the ovum. the effect of these variables on litter size independently Presumably, a boar associated with a high farrowing or have used a limited range of sperm numbers or qual- rate and large litters consistently produces insemina- ity ratings (Flowers, 1997; Xu et al., 1998). As a result, tions that contain sufficient numbers of spermatozoa information about the effect of their interactions on capable of completing all of these tasks. Therefore, it litter size is limited. The purpose of this paper is to discuss recent information examining the relative im- portance of the number and quality of spermatozoa in- seminated in terms of the manner in which they affect 1Supported by funding from North Carolina Agricultural Research the ability of boars to produce live pigs. Service, National Pork Producers Council, North Carolina Pork Coun- cil, and USDA (USDA 98-35203). Litter Size Based on Numbers 2Correspondence: Box 7621 (phone: 919-515-4003; E-mail: of Spermatozoa Inseminated william_fl[email protected]). Received April 5, 2001. Salisbury and Vandermark (1961) proposed a theo- Accepted August 9, 2001. retical relationship between male fertility and semen

E47 E48 Flowers characteristics that resembles a positive, asymptotic was between 8.8 and 12.2 pigs. These data provide evi- equation. Initially, when the number of spermatozoa is dence that boar fertility, as measured by litter size, can low, male fertility is poor. Increasing the number of differ considerably among individuals when identical sperm in the insemination dose results in a positive numbers of spermatozoa are inseminated. It is interest- increase in fertility. The magnitude of this response ing to note that estimates used to evaluate sperm qual- gradually diminishes as the number of sperm cells in- ity in these studies were high. The percentages of sperm seminated is increased until a plateau is reached. At cells that exhibited progressive forward motility and this point, insemination of additional numbers of sper- had normal morphology were greater than 71 and 91%, matozoa does not affect male fertility. Salisbury and respectively. Consequently, it is unlikely that the ob- Vandermark (1961) referred to this relationship as the served differences in litter size were due to individual male fertility pattern or curve. variations in these characteristics. The physiological rationale for male fertility patterns In contrast, Xu et al. (1998) failed to observe differ- is based on the concept that a finite number of compe- ences among boars but did show a positive effect of the tent spermatozoa must be present in the oviduct to number of spermatozoa inseminated on litter size. In successfully fertilize ova. Improvements in fertility are their study, insemination doses of 2 or 3 × 109 spermato- observed until this threshold level is achieved. After zoa were produced from ejaculates collected from six this point, there is not a concomitant increase in fertility boars and used to inseminate sows three times during when the number of sperm cells inseminated is in- a 2-d estrus. A total of 444 sows housed on a single creased because the population of spermatozoa in the farm were used. The range in mean litter size was be- oviduct needed to optimize fertilization has already tween 10.2 and 11.5 pigs when the insemination dose been achieved. contained 3 × 109 spermatozoa and between 9.1 and It has been difficult to demonstrate the existence of 10.1 pigs when 2 × 109 sperm cells were used. There fertility patterns for boars. Some of the problems en- was a main effect of insemination dose on litter size, countered are physiological in nature, whereas others but no interaction between insemination dose and boar. are associated with constraints inherent to the normal Collectively, these studies provide evidence that several management systems in which boar fertility is as- of the assumptions involving relationships between the sessed. Physiological limitations include the inability number of sperm inseminated and litter size central to to store boar semen for long periods without significant the fertility pattern concept of Salisbury and Vanderm- reductions in its fertilizing capacity (Johnson et al., ark (1961) are fulfilled. 2000) and the need to use large numbers of sperm cells However, what is lacking is the demonstration that in insemination doses compared with other livestock the manner in which litter size changes in response to species (Flowers, 1998a). Both of these effectively limit increasing sperm numbers differs among boars. Results the number of sows that can be inseminated from a from a study conducted with boars in a commercial stud single ejaculate. This, in turn, presents challenges in provide evidence for this (Flowers, 2002). In this study, separating the effect of the boar on litter size from 40 to 45 ejaculates were collected from 200 crossbred those of the sow, the production environment, and their boars (Duroc × Pietran × Large White, 2 to 4 yr of age) interaction. Constraints associated with management over a period of 2 yr. The sperm-rich fraction of each systems include the high replacement rate for boars and ejaculate was used to make insemination doses con- the diverse nature of mating regimens used (Flowers, sisting of 1, 3, 5, 7, or 9 × 109 total spermatozoa in 80 1998b). These also make estimation of boar fertility mL of Androhep (Minitube of America, Verona, WI) difficult for reasons similar to those mentioned pre- semen extender. Only ejaculates with greater than 70% viously. motility were extended and processed for delivery to Nevertheless, there is evidence to support the con- farms. At least two crossbred sows (Landrace × Large cepts that boars exhibit fertility patterns based on the White × Yorkshire) were bred with each insemination number of spermatozoa inseminated and that these dif- dose from each ejaculate. This resulted in a minimum fer among individuals. Two assumptions central to the of 75 sows bred with each insemination dose from each existence of fertility patterns are that males differ in boar. Insemination doses were stored between 16 and their fertility when the same number of sperm are in- 18°C and used within 48 h of collection during the study. seminated and that increasing the number of spermato- Sows received one insemination each day of estrus. zoa inseminated increases fertility within some portion It is important to note that sows used in the study were of the fertility curve. Johnson et al. (1981, 1982) con- housed on four different farms that were located within ducted two comprehensive studies that clearly demon- a 40-km radius of one another. Each farm consisted of strated that the fertility of boars is different when in- two 4,000-sow units under the same management. As semination doses contain equal numbers of sperm. In a result, sows on a single farm were inseminated with both of these studies, insemination doses of 3 × 109 semen from only 40 of the 200 boars during individual spermatozoa were used to inseminate sows once be- collection periods. Insemination doses were allocated tween 12 and 24 h after estrus. Collectively, there were to farms in a manner such that boars were used on the 3,300 sows used on at least 36 different farms. The same farm and in combination with one another an range in mean litter sizes among the 24 boars evaluated equal number of times during the duration of the experi- Boar effects on litter size E49 ment. Each farm had similar standard operating proce- dures, farrowing rates (84.7 ± 5.1%), and number of pigs born alive (11.2 ± 0.6) before and during the study. Due to the experimental design of this study, boar and farm effects on litter size were partially confounded during any individual collection period. However, due to the allocation process used for boars, it was possible to estimate the effects of boars and farms. This was done with a repeated measures analysis within an in- complete block design using mixed model methodolo- gies (Cochran and Cox, 1957; Littell et al., 1996). The incomplete block in the model consisted of the combina- tions of boars and farms that occurred during the study due to the allocation procedures. The statistical model included block, farm, boar, number of sperm insemi- nated, week, and appropriate interactions. When sig- nificant interactions between boar and number of sperm inseminated were present, differences among the num- ber of sperm inseminated within boars were de- termined. Analyses of litter size data from this study revealed several different fertility patterns as the numbers of sperm inseminated were increased from 1 to 9 × 109. One pattern was similar to the asymptotic relationship proposed by Salisbury and Vandermark (1961) in which there was an increase in litter size initially before a plateau was reached. This type of relationship was pres- ent in 133 of 200 boars. In contrast, a second fertility pattern was linear. This type of pattern occurred in 32 of 200 boars. It is possible that the linear pattern observed for some boars was simply a function of the experimental design. Their fertility pattern may have × 9 Figure 1. Representative examples of different relation- actually reached a plateau if more than 9 10 sperma- ± tozoa were inseminated. If this speculation is correct, ships between number of pigs born alive (mean SEM) then the range from 1 to 9 × 109 sperm cells may repre- and number of spermatozoa inseminated for boars in a sent the portion of their fertility curve in which litter commercial stud. The top panel illustrates examples of size increases as the number of sperm inseminated in- variation among boars with fertility patterns that reached creases. Finally, there were 35 boars for which a sig- a plateau. The lower panel illustrates examples of varia- nificant farm × boar interaction was observed for the tion among boars with fertility patterns that were linear. relationship between litter size and number of sperma- tozoa inseminated. The interaction occurred because the fertility pattern for these boars reached a plateau of spermatozoa are inseminated. Second, increasing the on some farms but remained linear on others. Because number of sperm cells in the insemination dose results the farms on which this occurred differed among boars, in an increase in litter size for some boars. Finally, the explanations for this interaction were not apparent. magnitude of the response in litter size to an increase In addition, there were distinct differences among in sperm numbers is not the same for all boars. From a boars in the shape of their individual fertility patterns. practical perspective, these observations provide some For those that reached a plateau, individual boar varia- important opportunities for improving litter size via tions occurred in both the insemination dose at which boar management on commercial swine operations. the plateau occurred and the mean litter size that re- Due to the variation observed among boars, it is reason- sulted. Similarly, the slope of the linear responses dif- able to speculate that development of practical and cost- fered among boars. Selected examples of the variation effective methods for determining the optimal number in the relationship between litter size and number of of spermatozoa for insemination doses would have a spermatozoa inseminated are illustrated in Figure 1. positive effect on litter size. For example, it would be In summary, based on the results from these four interesting to know whether spermatozoa from boars studies, three general conclusions concerning the rela- that exhibited a linear fertility pattern were different tionships among litter size, individual boars, and the from those with a pattern that reached a plateau in number of sperm inseminated seem warranted. First, terms of key morphological and biochemical character- litter size can differ among boars when equal numbers istics that are related to their ability to successfully E50 Flowers Table 1. Summary of selected measurements used to estimate sperm quality in boarsa

Measurement Rationale References

Florescent stains Some florescent dyes (Hoechst 33258) Johnson et al., 1996 enter if the membrane is damaged and stain only dead cells. Others (SYBR-14) enter cells with a membrane potential and stain only live cells. Macroscopic morphology Proportion of sperm with visual Xu et al., 1998 morphological defects is inversely related to ability to fertilize eggs in vitro. Hypoosmotic swelling test When exposed to hypoosmotic Vazquez et al., 1997 conditions the head of sperm with damaged membranes increase in size (begin to swell). Proportion of these sperm in an ejaculate is inversely related to fertility. Computer-assisted Number and characteristics of motile Holt and Medrano, 1997 motility analyses sperm using computer-assisted semen analysis (CASA) are correlated with fertility. Hemizona binding assay Oocytes are bisected and the number of Fazeli et al., 1995 sperm from different boars that bind to each half is correlated with their fertility. Sperm plasma membrane Proportion of three plasma membrane Ash et al., 1994 protein profile proteins present on sperm cells is Berger et al., 1996 profile positively correlated with fertility. Sperm chromatin When exposed to acidic conditions and Evenson et al., 1994 structure stained with metachromatic dyes, double-stranded DNA emit a green florescence, but single-stranded DNA (damaged) emit a red florescence. Ratio of green to red sperm is correlated to fertility of a boar. aAdapted from Flowers (1997). fertilize ova. If these differences are present and can phology (Xu et al., 1998), sperm membrane swelling be identified prospectively, then appropriate adjust- (Vazquez et al., 1997), viability (Johnson et al., 1996), ments in the number of spermatozoa in insemination binding to oocytes (Fazeli et al., 1995), and sperm chro- doses on an individual basis could be performed. matin structure (Evenson et al., 1994). Two of these tests, normal morphology and sperm Estimates of Semen Quality and Their chromatin structure, merit additional consideration be- Relationship to Litter Size cause their usefulness for determining the effect of indi- vidual boars on litter size has been examined. The pro- A number of procedures have been used to assess portion of spermatozoa with normal morphology ex- semen quality. Selected examples of these are summa- plained a large part (R2 = 0.59) of the variance in litter rized in Table 1 along with a brief synopsis of the ratio- size in the study of Xu et al. (1998). As mentioned ear- nale for each one. For the purpose of understanding lier, this study involved semen from six boars that was how these tests might be used to identify boars with used to breed 444 sows on a single farm. What is impres- superior fertility, it is useful to divide them into two sive about this study is that a single measure of semen categories. One category includes tests that focus on quality accounted for more than one-half of the varia- determining the proportion of spermatozoa in an ejacu- tion in litter size in a commercial setting. Consequently, late that possess certain characteristics that have been these results should be encouraging for others at- shown to be involved with a sperm cell’s ability to fertil- tempting to develop prospective methodologies for esti- ize ova. In essence, for these tests, the assumption is mating boar fertility. made that the proportion of spermatozoa with the prop- Studies evaluating the influence of sperm chromatin erty being measured is either positively or negatively structure used heterospermic inseminations composed related, depending on the characteristic, to the number of equal amounts of spermatozoa from two boars (Even- of pigs born alive when the boar is used for breeding. son et al., 1994). One of the boars used in the mixture The majority of techniques outlined in Table 1 fit into historically produced a high number of pigs born alive, this classification, including evaluations of normal mor- and his counterpart routinely sired small litters. The Boar effects on litter size E51 ratio of sperm with normal DNA to those with damaged reasonable to speculate that spermatozoa judged to be DNA was more than 90% effective in predicting the nonviable by fluorescent stains probably also are non- paternity of the pigs resulting from the heterospermic motile. In this situation, viability estimates should have inseminations. Traditionally, heterospermic insemina- a high positive correlation with motility and results tions have been viewed as a way to minimize the influ- from either procedure should be able to identify boars ence of the sow and the production environment on that produce small or large litters. In contrast, it is estimates of male fertility (Dziuk, 1996) because sper- conceivable that sperm cells with excellent morphology matozoa from different individuals literally compete could be deficient in the protein composition of their with one another simultaneously during fertilization. plasma membranes. If this were to occur, then a fertility However, it is important to acknowledge that fertility test based on normal morphology would predict the studies conducted in this way are comparative, or quali- production of large litters, whereas its counterpart us- tative, in nature. Their results can be used to rank ing sperm binding assays would not. In order to address boars in terms of fertility, but it is difficult to translate these possibilities, studies that examine the effective- the rankings into a mean value for the number of pigs ness of using several of these tests in sequence are born alive or other measures of fertility. As a result, needed. additional work is necessary in order to equate the proportion of spermatozoa with normal DNA in an ejac- Interactions Between Number and Quality ulate with actual litter size data resulting from homo- of Spermatozoa Inseminated spermic inseminations. The second category for semen quality tests involves As mentioned previously, whether or not changes in procedures that quantify the degree to which individual the number of spermatozoa inseminated can compen- spermatozoa exhibit a given characteristic, which, also, sate for variability in sperm quality is an interesting is involved with fertilization. For these tests, the as- question that has important implications for increasing sumption is made that sperm cells express varying lev- litter size in pigs. If one accepts the argument that a els of the characteristic. Boars whose spermatozoa ex- critical number of competent spermatozoa in the ovi- hibit increased amounts produce larger litters com- duct are required to optimize fertility, then it seems pared with individuals whose sperm cells have low reasonable that this critical number could be achieved values. Computer-assisted motility analyses (Holt and via various combinations of the quality and number of Medrano, 1997) and the protein composition of sperm spermatozoa inseminated. However, in order for this membranes (Berger et al., 1996) are examples of these to be realistic physiologically, three things must occur: types of semen quality tests. In a series of studies con- 1) increases in the number of spermatozoa inseminated ducted on commercial swine farms, changes in the ve- should increase the number of sperm entering the ovi- locity of the motion of spermatozoa during an in vitro duct, 2) entry into and retention of spermatozoa in the incubation period explained 20% of the normal varia- oviducts need to differ among boars, and 3) transport tion in litter size (Holt et al., 1997). Furthermore, boars and entry of spermatozoa into the oviduct should be whose spermatozoa exhibited increased straight-line independent of sperm quality. velocity and track linearity were associated with large There is evidence for the existence of each of these litter sizes. Similarly, concentrations of three proteins events. First, Baker and coworkers (1968) inseminated isolated from the plasma membrane were positively gilts with 1, 5, or 10 × 109 spermatozoa and determined correlated (r = 0.38 to 0.53) with the ability of sperm the number remaining in the oviducts 12 to 16 h later. from individual boars to bind to egg membranes (Ash Even though the recovery rate was low, a positive rela- et al., 1994) and the proportion of pigs they sired when tionship between the number of spermatozoa insemi- heterospermic inseminations were used (Berger et al., nated and the number of sperm cells recovered from 1996). Consequently, both of these measures of semen the oviducts was present. Second, significant effects of quality seem to be good candidates for explaining some boars on the number of spermatozoa recovered from of the variation in fertility patterns among boars. the lower isthmus of the oviduct around ovulation have However, what is difficult to decipher, at the present been observed (Mburu et al., 1996). Differences among time, is which of the characteristics outlined in Table 1 boars varied from 461 to 1,972 spermatozoa. Finally, a simply reflect shifts along an individual fertility pattern series of experiments by First et al. (1968) demon- compared with those that are involved with changing strated that dead spermatozoa are transported and en- the level at which the plateau occurs or the slope of ter the oviduct with efficiencies similar tho those of live a linear response. Shifts along a given fertility curve sperm. Consequently, based on what is known about optimize litter size for individual boars. In contrast, the dynamics of spermatozoa in the oviduct, adjusting increasing the litter size at which the maximal response sperm numbers at insemination based on some assess- is achieved has the potential, in theory, to improve the ment of their quality seems to be a physiologically plau- fertility of boars regardless of their inherent pattern. sible way to increase litter size for some boars. Another question that awaits resolution is the degree Even though critical evaluations of this strategy in- of dependency inherent among the characteristics mea- volving the birth of live pigs are lacking, results from sured by each of these procedures. For example, it is a field study conducted by Johnson et al. (1988) are E52 Flowers encouraging. In this study, insemination doses con- insemination, increasing litter size probably can occur sisting of 3 × 109 spermatozoa were used to breed sows in several different ways, including improving semen between 0 and 72 h after collection. In contrast, if the quality, increasing the number of spermatozoa insemi- collection-to-insemination interval was greater than 72 nated, and using estimates of sperm quality to adjust h, 6 × 109 spermatozoa were inseminated. No effect number of sperm inseminated. However, the magnitude of semen age on litter size was observed. Although a of changes in litter size resulting from these strategies contemporary treatment consisting of insemination is likely to vary considerably among boars. doses with 3 × 109, aged (> 72 h) spermatozoa was not included in the design, it is commonly accepted that Literature Cited quality estimates and fertility of fresh semen decrease significantly as storage time increases. This is particu- Ash, K. L., T. Berger, C. M. Horner, and T. R. Famula. 1994. Boar larly relevant considering that the semen extenders sperm plasma membrane protein profile: correlation with the zona-free hamster ova assay. Theriogenology 42:1217–1226. used in their study, BTS and MR-A, are classified as Baker, R. D., P. J. Dziuk, and H. W. Norton. 1968. Effect of volume short-term extenders because fertility of semen is re- of semen, number of sperm and drugs on transport of sperm in duced significantly after 3 d of storage (Johnson et al., artificially inseminated gilts. J. Anim. Sci. 27:88–93. 2000). Consequently, the speculation that sperm num- Berger, T., D. L. Anderson, and M. C. T. Penedo. 1996. Porcine sperm bers can be adjusted to compensate for reduced semen fertilizing potential in relationship to sperm functional capacit- ies. Anim. Reprod. Sci. 44:231–239. quality seems to be supported indirectly by the results Cochran, W. G., and G. M. Cox. 1957. Experimental Designs. 2nd from field studies. ed. John Wiley & Sons, New York. However, there are several caveats associated with Dziuk, P. J. 1996. Factors that influence the proportion of offspring this strategy that should be considered. The best esti- sired by a male following heterospermic insemination. Anim. mates of sperm quality upon which adjustments should Reprod. Sci. 43:65–88. Evenson, D. P., L. Thompson, and L. Jost. 1994. Flow cytometric be made are not known. Nevertheless, it is logical to evaluation of boar semen by the sperm chromatin structure speculate that tests that are categorical in nature, such assay as related to cryopreservation and fertility. Theriogenol- as the percentage of spermatozoa with normal morphol- ogy 41:637–651. ogy, probably are the best candidates to study initially. Fazeli, A. R., C. Holt, W. Steeneg, M. M. Bevers, W. V. Holt, and B. Based on the studies discussed previously, insemina- Colenbrander. 1995. Development of a sperm hemizona binding assay for boar semen. Theriogenology 44:17–27. tion of additional numbers of normal sperm should re- First, N. L., R. E. Short, J. B. Peters, and F. W. Stratman. 1968. sult in additional normal sperm in the oviduct. How- Transport and loss of boar spermatozoa in the reproductive tract ever, as mentioned previously, estimates based on a of the sow. J. Anim. Sci. 27:1037–1040. sequence of several different tests probably deserve Flowers, W. L. 1997. Management of boars for efficient semen produc- equal consideration. tion. J. Reprod. Fertil. Suppl. 52:67–78. Flowers, W. L. 1998a. Artificial insemination in animals. In: E. Knobil In addition, it is unlikely that quality adjustments and J. D. Neill (ed.) Encyclopedia of Reproduction. p 296. Aca- for the number of spermatozoa inseminated would have demic Press, San Diego, CA. similar effects for all boars in terms of increasing litter Flowers, W. L. 1998b. Insemination programs for swine to increase size. For example, boars whose fertility pattern reached fertility. J. Anim. Sci. 76(Suppl. 3):39–46. a plateau at 3 × 109 spermatozoa would not be expected Flowers, W. L. 2002. Fertility patterns of boars in a commercial stud. Anim. Reprod. Sci. (In press). to elicit the same response as their counterparts with Holt, C., W. V. Holt, H. D. M. Moore, H. C. B. Reed, and R. M. linear patterns or those with patterns that reached a Curnock. 1997. Objectively measured boar sperm motility pa- plateau at 7 × 109 spermatozoa. Presumably, the level rameters correlate with the outcomes of on-farm inseminations. at which the plateau occurs reflects differences among Results of two fertility trials. J. Androl. 18:20–31. boars in terms of the critical number of competent sper- Holt, W. V., and A. Medrano. 1997. Assessment of boar sperm function in relation to freezing and storage. J. Reprod. Fertil. Suppl. matozoa in the oviduct needed for optimal fertilization. 52:213–222. As a result, it is reasonable to speculate that this strat- Hunter, R. H. F. 1990. Fertilization of pig eggs in vivo and in vitro. egy for improving litter size is better suited for boars J. Reprod. Fertil. Suppl. 40:211–226. with high rather than low requirements. Johnson, L. A., J. G. Aalbers, C. M. T. Willems, and W. Sybesma. 1981. Use of boar spermatozoa for artificial insemination. I. Fertilizing capacity of fresh and frozen spermatozoa in sows on Implications 36 farms. J. Anim. Sci. 52:1130–1136. Johnson, L. A., J. G. Aalbers, C. M. T. Willems, J. H. M. Rademaker, The number and quality of spermatozoa inseminated and C. E. Rexroad, Jr. 1982. Use of boar spermatozoa for artificial determine the boar’s impact on litter size. The relation- insemination III. Fecundity of boar spermatozoa stored in belts- ship between these traits and litter size tends to be ville liquid and kiev extenders for three days at 18 C. 1982. J. Anim. Sci. 54:132–136. unique for boars and is best described as a fertility Johnson, L. A., J. G. Aalbers, and H. J. G. Grooten. 1988. Artificial pattern or curve. Increasing litter size on operations insemination in swine: Fecundity of boar semen stored in Belts- using natural service is most likely to result from im- ville TS (BTS), Modified Modena (MM) or MR-A and inseminated provements in the quality of spermatozoa. This is due on one, three and four days after collection. Zuchthygiene (Berl.) 23:49–55. to the fact that most boars contain sufficient spermato- Johnson, L. A., W. M. C. Maxwell, J. R. Dobrinsky, and G. R. Welch. zoa in their ejaculate to maximize sperm numbers in 1996. Staining sperm for viability assessment. Reprod. Domest. the oviduct. In contrast, on operations using artificial Anim. 31:37–47. Boar effects on litter size E53

Johnson, L. A., K. F. Weitze, P. Fiser, and W. M. C. Maxwell. 2000. Sirard, M. A., A. Dubuc, D. Bolamba, Y. Zheng, and K. Coenen. 1993. Storage of boar semen. Anim. Reprod. Sci. 62:143–172. Follicle-oocyte-sperm interactions in vivo and in vitro in pigs. Littell, R. C., G. A. Milliken, W. W. Stroup, and R. D. Wolfinger. J. Reprod. Fertil. Suppl. 48:3–16. 1996. SAS௡ System for Mixed Models. SAS Inst. Inc., Cary, NC. Vazquez, J. M., E. A. Martinex, P. Martinez, C. Garcia-Artiga, and Mburu, J. N., S. Einarsson, N. Lundheim, and H. Martinez Rodriguez. J. Roca. 1997. Hypoosmotic swelling of boar spermatozoa com- 1996. Distribution and membrane integrity of spermatozoa in pared to other methods for analyzing sperm membrane. Therio- pig oviduct in relation to ovulation. Reprod. Domest. Anim. genology 47:913–922. 31:57–68. Xu, X., S. Pommier, T. Arbov, B. Hutchings, W. Sotto, and G. R. Miller, D. J. 2000. A sperm’s perspective of fertilization. Proc. Am. Soc. Anim. Sci., 1999. Available at http://www.asas.org/jas/symposia/ Foxcroft. 1998. In vitro maturation and fertilization techniques proceedings/0918.pdf. Accessed Aug. 1, 2001. for assessment of semen quality and boar fertility. J. Anim. Sci. Salisbury, G. W., and N. L. Vandermark. 1961. Physiology of Repro- 76:3079–3089. duction and Artificial Insemination in Cattle. Freeman and Co., San Francisco, CA. Can farm animal welfare be understood without taking into account the issues of emotion and cognition?

R. Dantzer

Integrative Neurobiology, INRA-INSERM U394, 33077 Bordeaux Cedex, France

ABSTRACT: Although the concept of welfare makes species, to animals of different ages, which are likely reference to feelings of individual animals, the exact to have varying degrees of emotional and cognitive ca- nature of these feelings and their relationship to emo- pacity. In the past, disagreements about exact defini- tions and cognitive abilities of the animals under con- tions of emotion and cognition have blurred the matter sideration are never detailed. Based on the concepts and hampered research. However, this should no longer of stress and coping, an extensive list of indicators of be the case; the study of emotions and feelings has physical health, production, behavior, and physiology emerged as a field of active research in psychology and has been set up for the purpose of recognizing good from neuroscience over the last two decades. It is now possi- bad welfare. However, these indicators do not allow us ble to go over philosophical discussions on the nature to make any inference about mental states. This issue of feelings and to set up a research agenda on emotion is important because welfare considerations apply to and cognition in farm animals that should help us to farm animals of different species and, within a given understand their welfare requirements.

Key Words: Cognitive Development, Domestic Animals, Emotions, Neurology, Stress

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E1–E9

Introduction of positive and negative emotional states. There are obvious reasons for that, the main one being the reluc- Statements about the nature of animal welfare make tance of ethologists to engage in a thorough study of explicit or implicit reference to mental states. As a typi- emotions in animals. There is still the biased prejudice cal and influential example, Broom (1986) defines the that research on emotion and cognition is unscientific welfare of an animal as “its state as regards its attempts and should be carried out only on human beings, who to cope with its environment.” In the report of the Euro- can express what they feel via verbal reports. The pur- pean Scientific Veterinary Committee on the welfare pose of this theoretical review paper is to show that of calves (European Commission, 1995), this definition there are ways out of this conservatory attitude and it sets the scene and is followed by the statement that is possible to engage in objective studies of emotions in “the state of the animal, as referred to in the definition, farm animals and how they relate to cognition. includes the feelings of the individual animals as its physical state. Suffering is one of the most important Emotion and Cognition aspects of poor welfare and we should investigate the existence of good or bad feelings wherever possible The most elementary component of an emotional when trying to assess welfare.” However, the problem is state is its affective dimension. According to Cacioppo that what these mental states termed “feelings” exactly and Gardner (1999), affective categorization and re- consist of, and how they relate to coping, is never clari- sponses have been shaped by evolution to allow differ- fied. As pointed out by Dawkins (2001), research on entiation between hostile and hospitable stimuli. This the psychological health of farm animals has mainly differentiation is so critical that organisms have rudi- concentrated on establishing checklists of biochemical, mentary reflexes for categorizing and approaching or physiological, and behavioral indicators without taking withdrawing from certain classes of stimuli and for pro- the necessary steps to relate them to the occurrence viding metabolic support for these actions. An addi- tional adaptive advantage is conferred to species whose individual members have the capacity to learn based 1Correspondence: Rue Camille Saint-Saens (phone: 33 5 57 57 37 25; fax: 33 5 56 98 90 29; E-mail: [email protected]). on the unique environmental contingencies to which Received April 5, 2001. they are exposed, to represent and predict events in Accepted August 27, 2001. their environment, to manipulate and plan based on

E1 E2 Dantzer representations, and to exert some control over their and tonic immobility. All these behavioral responses attentional and cognitive resources. are usually accompanied by changes in physiology that The affective component of an emotional state can be reflect activation of the hypothalamic-pituitary-adrenal simply assessed by determining whether the animal (HPA) axis and the orthosympathetic branch of the approaches or avoids the eliciting situation either spon- autonomic nervous system. These physiological taneously or after appropriate classic or instrumental changes do not necessarily differ according to the emo- learning. An object is described as pleasurable, and the tion that is experienced (Dantzer, 1989). Their relation animal is assumed to experience pleasure, if this object to the subjective component of emotions is quantitative is approached and eventually consumed or if the animal rather than qualitative, in the sense that the magnitude remains in close contact with it. Conversely, an object of the physiological arousal modulates the intensity of is said to be aversive, and an animal exposed to this emotion. An emotion requires both a physiological object is assumed to experience a negative emotion, if arousal and the perception of a meaningful, contextual- this animal escapes it or subsequently makes every environmental cognition (Schachter and Singer, 1962). effort to avoid it. Preference tests in applied ethology There has been much debate on the relative importance make use of this basic distinction to determine what is of the visceral and the cognitive processes in the genesis good and what is bad for the welfare of an animal. of emotion, and the order in which they take place (i.e., Typically, if a pig that is given the choice among two does the visceral arousal precede cognition, or vice- types of floor stays longer on a given floor type (e.g., a versa?). In the present case, it is not necessary to enter straw-bedded floor) and hardly walks or lies down on the details of this controversy, and it is sufficient to the other floor type, (e.g., concrete slats), its state of state that modern theories of emotion agree on the role welfare will be reputed to be good or bad depending of the environmental context in determining the specific on what type of floor it is normally exposed to. More quality of the emotional response (Leventhal, 2000). sophisticated methods have been developed to find out This allows us to understand why behavioral and physi- how much animals value resources by measuring how ological indicators of emotion in animals are mostly high a price, in terms of time and energy, they will pay used for describing emotionality and its variation ac- to access or avoid them (Dawkins, 1983). The rewarding cording to genotype and previous experience, whereas nature of a single resource such as food can be estimated contextual cues are used to specify which type of emo- by the use of a progressive ratio schedule that consists tion is experienced. of requiring an animal put into an operant conditioning Whether emotions are defined on a bidirectional axis chamber to press a panel once to get the reward, then (pleasure vs displeasure) or in terms of discrete emo- twice, then four times in a row, and so on, until the tional states, the assumption remains the same. Ani- reward is no longer worth the effort (the so-called break- mals that display behavioral and physiological signs of ing point) (Dantzer, 1976). Several resources can be emotion are assumed to experience the corresponding compared by use of closed-economy set-ups in which emotional state. Many scientists choose to stay at this animals have permanent access to several resources level when it comes to defining emotions. A typical ex- providing they pay the appropriate cost (e.g., by push- ample is Panksepp (1988), who started his career with ing through weighted doors) (Mason et al., 2001). the description of stimulus-bound behavior in rats, i.e. It is possible to go one step further and qualify the behavioral responses elicited by electrical stimulation specific emotional state that is experienced by an ani- of specific brain sites. Panksepp (1982) claimed that mal. Primary emotions refer to a set of six to eight basic the primary emotions animals are endowed with (expec- emotional reactions (e.g., fear, joy, disgust, sadness, tancy, rage, fear, and panic; joy was added later) arise surprise, and anger) that are innate and found in hu- from the operation of hard-wired neural circuits in the man and non-human animals (Leventhal, 2000). The visceral-limbic brain that facilitate diverse and adap- usual approach for determining experimentally tive behavioral and physiological responses to environ- whether an animal can express a given emotion is to mental challenges. The mere existence of the appro- expose this animal to an emotional stimulus that is priate behavioral and physiological signs of emotion assumed to be at the source of the emotion under consid- allows us to infer that the corresponding emotional eration and to assess whether it presents the behavioral state exists in the animal under consideration. Al- and physiological changes that are congruent with the though the Panksepp (1988) description is limited to emotional nature of the stimulus. A chicken is described the mammalian brain, it is certainly possible to find as frustrated if it responds to a situation in which it is equivalents of such circuits in the reptilian brain that prevented from performing a motivated behavior by birds are equipped with, especially so because Pank- showing signs of behavioral agitation (e.g., wing flap- sepp also worked with chicks. ping, aggression toward conspecifics, spot pecking, or The problem with the Panksepp (1988) description pacing). In the same way, a chicken is assumed to be of emotional states is that it does not allow us to differ- frightened if it responds to a potentially threatening entiate fixed response patterns that are hard-wired and stimulus by behavioral signs of fear that vary according do not necessarily require a central nervous system to the distance from the threat and the ability to avoid from more plastic, adaptive responses. As aptly pointed it. Fear responses in chickens include flight, fighting, out by Staddon (1983), relatively simple built-in re- Emotion, cognition, and farm animal welfare E3 sponses such as tropisms enable animals to display Damasio (1998) has added to this description the so- a large set of very well-adapted behaviors, including matic marker hypothesis, which allows us to relate vis- avoiding bad things and approaching good things. Rolls ceral responses to feelings. Those brain areas in which (1999) claims that such adaptive mechanisms that re- emotions are represented in the form of motor programs quire little or no dependence on past history do not need to be able to process visceral information and to involve any intervening emotional state. Emotions re-actualize this processing according to the way emo- come into play when adaptive behavior depends more tions are expressed both behaviorally and physiologi- and more on the animal’s past. Emotions, therefore, cally. In the brain, the mechanisms of emotions are require the ability to establish temporal and instrumen- part of a survival kit, just above the basic motivations. tal contingencies. Learning a temporal contingency re- Feelings come into action to allow anticipation of what fers to the ability to relate the occurrence of one stimu- can happen or prediction of what is going on during the lus to another one when both occur in succession, and course of an emotional situation that emotional re- to make the first one a possible causal factor for (or a sponses are already dealing with. Consciousness of an predictor of) the second one. Learning an instrumental emotion is a form of a meta-knowledge because it corres- contingency refers to the ability to assess the conse- ponds to the knowledge of one’s ability to experience quences of one’s response on the environment (e.g., to emotions (Damasio, 1998). learn that a given response is followed by a set of specific In more general terms, the issue of the relationship consequences). In this context, emotions provide the between emotion and cognition has already received a common currency with which animals can balance con- lot of attention in psychology. Contrasting theories have flicting demands of avoiding bad things and ap- been put forward, some proposing that there cannot be proaching good things and evaluate the priority to be any emotion without some form of cognition, others given to one over the other. pointing out that emotion and cognition are distinct The main problem with all of that which precedes is processes. Without getting into the details of the contro- that it describes the way an emotional system in the versy that is mainly based on empirical work carried brain is operating and the minimum mental operations out in human subjects, it is important to know what is it requires, but it says nothing about the subjective at stake here because it has significant implications for aspects of emotions, or feelings. As pointed out by Le the conceptualization of emotions in animals. A few Doux (1996), “a subjective emotional experience, like scientists, such as Zajonc (1980), claim that emotions, the feeling of being afraid, results when we become actually confounded with preferences, are more primi- consciously aware that an emotion system of the brain, tive than cognition, represented by mental operations like the defense system, is active.” To have a defense such as object recognition, and therefore can occur in system that gives rise to appropriate behavioral and the absence of any cognitive operation. The vast major- physiological responses in the face of a threatening situ- ity of psychologists, however, agree that an emotion ation is not sufficient to feel afraid; we need to have involves some form of cognition about an emotional the key elements of consciousness that allow us to be situation. For Lazarus (1984), an emotional state re- aware of the activity of this defense system. Duncan quires a primary appraisal of the emotional stimulus. and Petherick (1991) stand on the same ground when This primary appraisal is responsible for the setting they claim that the extent to which animals are aware and execution of action plans, and it is followed by of their internal state while performing behavior known a secondary appraisal of the consequences of action. to be indicative of so-called states of suffering, such as Contemporary psychologists have tried to describe fear, frustration, and pain, will determine how much more precisely the exact dimensions of the eliciting situ- they are actually suffering. Of course, emotions have ation that are evaluated and give rise to an emotion. evolved as brain states and bodily responses, and not Although their description usually tells us nothing or necessarily as subjective feelings conveyed linguisti- very little about the contribution of physiological activa- cally; otherwise, Darwin (1872) would not have been tion to this appraisal process, the dimensions that have able to study their phylogeny. However, these basic been identified are certainly useful for trying to describe elements are not sufficient to make up a full emotion. in very objective terms the types of emotion that are Some elements of consciousness are still necessary. For accessible to different animals and the way they can LeDoux, there is nothing special in the conscious opera- differ according to age and species. A very good example tions that allow us to be aware of our emotional states. of this type of contribution is that provided by Scherer The mental representation of a flower and the mental (1997), with his theory of the emotion-antecedent ap- representation of a threat make use of the same build- praisal process. In contrast to most of his colleagues ing blocks. What is specific is the way the sensory input working on appraisal, he chose not to rely on verbal is processed to give rise to a mental representation. reports to assess the way appraisal is carried out; in- Emotional stimuli are given a special treatment in the stead, he concentrated on the objective antecedents or sense that most of them escape the sensory cortical eliciting conditions for emotional reactions. The major analyzer to be deciphered at the subcortical level, via criteria he listed for an emotional object are its novelty, a shortcut, a neural pathway directly connecting the intrinsic pleasantness, goal significance, coping poten- sensory thalamus to the amygdala. tial, and relevance to external and internal standards. E4 Dantzer Table 1. Appraisal patterns (listed in rows) for some primary emotions (listed in columns)a

Item Digust Sadness Fear Rage Happiness

Novelty Suddenness Open Low High High Low Familiarity Low Low Low Low Open Predictability Low Open Low Low Medium Pleasantness Very low Open Low Open High Goal significance Outcome probability Very high Very high High Very high Very high Urgency Medium Low Very high High Very low Coping potential Open Very low Open High Open aAdapted from Scherer 1988. Note that goal significance includes only outcome probability and urgency, and coping potential refers only to controllability. The status of a given antecedent varies between low and high. Low familiarity, for instance, corresponds to novelty; high urgency refers to the obligation of doing something very quickly in order to deal with the situation. The status is said to be open when its level is not relevant; for instance, disgust can be induced by sudden as well as by slowly developing taste stimuli.

With the help of these criteria, he was able to distin- creased pituitary-adrenal activity, as already described guish most primary and secondary emotions, and even in pigs that have been trained to work for a food reward to find emotional states for which there is still no verbal by pushing a panel with their snouts in an operant emotion label (Table 1). conditioning cage, and which, on the test day, are put into the cage with the feeder turned off (Dantzer et A Strategy for Studying Emotion and al., 1980). These behavioral and physiological reactions Cognition in Farm Animals form a basic vocabulary of primitive emotions that is suggestive, and only suggestive, of the presence of an Because applied ethology deals with a wide range of emotional state and its associated emotional expe- species, some standardization is obviously in order to rience. describe the range of emotional states that are accessi- Although behavioral agitation and physiological ble to farm animals. It is necessary to overcome the arousal are the most visible components of the pig’s previously described limitations of the behavioral ap- automatically elicited emotional state of frustration, proaches that have been used so far for studying emo- they are short-lived and not necessarily accompanied tional responses in animals. In order to incorporate in by a feeling of frustration. Such a feeling is most likely this task an appropriate evaluation of cognitive abilities to arise if the pig is able to elaborate on what has of farm animals, the reference framework has to rely happened to it and how it has reacted to it. This is where the schematic level of processing comes in. The as far as possible on observable elements rather than minimum requirement here is the ability to form what on linguistic labels. The appraisal model elaborated by cognitivists call schemata (i.e., neural representations Scherer (1997) can be proposed for this purpose. It is of the situation and the organism’s response to this relatively easy to plan similar emotional situations for situation). If a pig that is repeatedly exposed to a stimu- different animal species based on the elementary ap- lus that signals the cessation of delivery of food learns praisal dimensions that are delineated in the Scherer to respond to it by emitting a response that terminates (1997) theory (novelty, pleasantness, coping potential, the signal so that it can now be rewarded, then there etc.) and to investigate the details of the appraisal pro- is some reason to believe that it is able to feel frustrated. cess rather than debating the extent to which it is cogni- Note that schemata operate automatically and are not tive or not. Based on Leventhal’s theorizing, emotions easily controlled by conscious reasoning or voluntary can be seen as developing from relatively simple, reflex- effort. The fact that mammalian farm animals can learn like forms (wired in innate sensory-motor processes) temporal and instrumental contingencies is a good in- into complex cognitive-emotional processes that result dex of their ability to form schemata. A pig that has from the participation of at least two distinct levels of learned to avoid impending electric shocks by jumping memory and information processing, a schematic and from one side of a two-compartment cage to the other a conceptual level (Leventhal and Scherer, 1987) (Fig- will respond to a fear signal (i.e., a tone previously ure 1). associated with electric shock) by increasing its re- Imagine an emotional state with the following ante- sponse rate (Dantzer and Morme`de, 1976) (Figure 2). cedents: suddenness, low familiarity, low predictability, Conversely, it will react to a safety signal (i.e., a tone very low intrinsic pleasantness, high goal significance, signaling the absence of electric shock) by decreasing and low coping potential; in all likelihood, this emo- its response rate. In other words, pigs make use of their tional state corresponds to frustration. This emotional past experience to predict what is likely to happen in state is characterized by behavioral agitation and in- the situation they are exposed to. Emotion, cognition, and farm animal welfare E5

Figure 1. Multilevels of emotion processing (adapted from Leventhal and Scherer, 1987).

The synchrony between behavioral and physiological results in physiological de-activation (Dantzer, 1989, responses that is characteristic of an emotional re- 1993). Conversely, the inability to control the situation sponse takes place not only at the sensory-motor level, is a strong inducer of physiological arousal. The way but also at the schematic level that involves situational animals respond to the frustrating situation is deter- and response expectations. The ability to control the mined by their expectation of the food reward, which emotional situation by a suitable behavioral response itself influences their blood levels of cortisol. This syn-

Figure 2. Behavioral responses of pigs with a continuous avoidance history to a fear signal (CS+) and to a safety signal (CS−). Pigs were trained to avoid painful electric shocks by crossing a barrier in a two-compartment cage according to a continuous avoidance procedure (shock-shock interval = 10 s, response-shock interval = 30 s). After stabilization of performance, they were presented with a 5-s tone always (CS+) or never (CS−) followed by shock, 10 times for each tone, in four conditioning sessions. On the next day following the last discriminative fear conditioning session, they were tested in the two-compartment cage in extinction conditions (i.e., without any shock), and CS+ and CS− were presented in random order nine times each. Half the pigs were pretreated with saline (n = 4) or with 1 mg/kg diazepam (n = 4) 30 min before the session. Each graph shows the mean number of responses (mean ± SEM) in 5-s successive intervals just before the CS (−5 s), during it (0 s), and after (5 to 30 s). Note that presentation of the fear signal increased response rate whereas that of the safety signal had the reverse effect. Diazepam injected at an anxiolytic dose had no effect. Adapted from Dantzer and Morme`de, 1976). E6 Dantzer with the situation (e.g., high cortisol levels are associ- ated with a higher probability of engaging in passive coping, whereas high catecholamine levels are associ- ated with a higher probability of engaging in active coping; Dantzer and Morme`de, 1983). They also allow brain memory systems to differentially process for fur- ther use sensory aspects of the emotional situation and outcomes of emotional behavior (Morme`de and Dan- tzer, 1978). A full account of emotional reactions in farm animals requires at least the assessment of the extent to which the necessary synchronization between behavioral and physiological responses takes place. This type of re- search is unfortunately still relatively rare. In the case of chickens, for example, frustration has been defined at the behavioral (Duncan, 1970) but not at the physio- logical level, so that it is not yet known whether the operating characteristics of the emotional system at Figure 3. Relationship between plasma cortisol levels the sensory motor level are the same in mammals and and behavioral response to frustration in pigs. Pigs that in birds. had learned to get a food reward by pressing a panel In Leventhal’s theory, the third level of processing is with their snouts were put into the operant conditioning the conceptual (Leventhal and Scherer, 1987). At this cage on the day of test with the food dispenser turned level, it becomes possible to reflect upon, abstract, and off. Open circles show for each pig the initial plasma draw conclusions about the environment and one’s emo- cortisol levels, measured just before the start of the ses- tional responses to it. This is analogous to the meta- sion, and closed circles the plasma cortisol levels mea- knowledge discussed before (Damasio, 1998), and it can sured 30 min after completion of the frustration session. be safely assumed that it needs a minimum form of Note that the number of responses emitted during the abstraction and self-consciousness to be achieved. session was correlated negatively to initial (dotted line) Therefore, the probability that this level is accessible but not to final (interrupted line) plasma cortisol levels. to farm animals is unlikely. Adapted from Dantzer et al. (1980). Application of the hierarchical model of emotion to farm animals allows us to determine more precisely the type of emotional experience that is accessible to them. chrony between physiological activation and behavior The case of sows in maternity stalls illustrates this can be seen from the positive correlation between blood issue very well. The nervousness and stereotypes preg- cortisol levels at the start of the frustration session nant sows develop just before parturition are often in- and the number of times pigs continue presenting the terpreted as indices of the frustration these animals previously rewarded response (Dantzer et al., 1980) are experiencing. It is said that sows feel frustrated (Figure 3). because they cannot engage in the nest-building behav- Because of this synchronization process, changes in ior they would normally present in the wild. However, circulating hormones can be used a posteriori to assess based on the multilevel theory of emotion, frustration whether the behavioral response to the situation is can develop only if sows are able to form schemata, in truly indicative of an emotional state. This possibility the present case a mental representation of what they has been very elegantly made use of by Mason et al. are missing (e.g., adequate material for nest building). (2001) to show that the way mink value different re- The path is open here for a logical trap. By definition, sources in a closed-economy set-up does not necessarily frustration can only occur in individuals that are miss- imply that the lack of preferred resources is detrimental ing something they have experienced earlier. Sows that to welfare. In their experiment, urinary cortisol levels are confined in gestation stalls are obviously likely to increased only when mink were prevented from getting miss several features they have already experienced, access to a water pool, but not when they were unable such as the ability to move around and to interact with to visit an alternative nest site or an empty cage, two conspecifics. This can be a source of frustration, and resources with lower incentive value than the water the level of frustration confined sows experience is a pool. Elevations in urinary cortisol of mink denied ac- function of the relative importance of the items they cess to the water pool were of the same magnitude as are missing. In order to assess this relative importance, those observed in mink deprived of food. Activation of the same type of procedure as that used by Mason et the autonomic nervous and neuroendocrine systems al. (2001) for mink can be put to work. However, once serves to fuel the motor components of the emotional more, the positive results obtained with such a proce- response. However, it is not its sole role. By their action dure will not demonstrate the existence of a frustration on the brain, hormones influence the way animals cope in a situation other than the one that has been set up Emotion, cognition, and farm animal welfare E7 for the experiment. The mink that were studied in the the triggering factor would be the restlessness and en- Mason et al. experiment had no previous experience hanced arousal associated with the beginning of partu- with the swimming pool. In order to learn to push open rition, and the resulting emotional state would be that a door so as to get access to the swimming pool, they of the frustration already experienced during confine- obviously had to develop some representation of what ment in gestation stalls. There are several direct and a swimming pool is and what it means to their welfare. indirect ways of testing this possibility. An obvious ap- The fact that subsequent blockade of access to the swim- proach is to verify that the postulated emotional state ming pool door caused behavioral agitation and in- develops less easily in those animals that are exposed creased urinary cortisol levels was clearly indicative of to the situation but have had limited opportunity to a state of frustration. However, these findings do not previously experience the same emotional state than imply at all that mink that are raised as fur animals in those animals that have already had ample opportu- and have never seen a swimming pool are frustrated nity to experience it. However, such an experiment is when there is no swimming pool in their cage. Such a not necessarily easy to set up. An indirect approach conclusion is condemned to be logically flawed as long would be to search for evidence of the plastic changes as mink have not been shown independently to be able in the brain that underlie the expression of such a phe- to form mental images of something they have never ex- nomenon. At the neurobiological level, the process that perienced. allows a full emotion to develop in response to a weak The appropriate vocabulary here is therefore that of stimulus that normally has no effect on its own is known motivation, or need, not that of emotion. To come back as sensitization. When the initial stressor is sufficiently to sows in maternity stalls, they are likely to be in intense, the neural circuitry that mediates the corres- the same state, in relation to the hormonally induced ponding emotion may undergo a long-term increase in changes in their internal milieu, as are hungry animals sensitivity (Antelman et al., 2000). The cellular and in relation to their energy status. Hunger is a motiva- molecular basis of this sensitization process has been tion, not an emotion. The fulfillment of a need takes elucidated in several neurochemical systems in vivo place at the level of sensory-motor processing. It obvi- (Bremner et al., 1996; Post and Weiss, 1997), and even ously has an affective dimension that is generally posi- in simpler animal models such as the gill-withdrawal tive, but such an affective state is independent of the reflex of aplysia (Cohen et al., 1997). In neurobiological ability to form schemata. Being hungry is certainly terms, however, what is relevant for emotion is not aversive in most situations and will therefore be ac- the nature of the biochemical process per se but where tively avoided. However, being hungry is a necessary exactly it takes place in the brain. but not a sufficient condition for frustration because The previously delineated description of emotional frustration does not occur just because of the lack of states has been carried out hardware-free (i.e., without food but because of the lack of expected food. paying any attention to the neural basis of emotion). This type of reasoning must not be pushed too far, This attitude was certainly very popular until two de- however; otherwise it leads to another trap, that of cades ago. However, it has been superseded now by dogmatism. Engrained in the multi-level theory of emo- the recent developments of research on emotion and tion developed by Leventhal (Leventhal and Scherer, cognition in neuroscience. The reductionist power of 1987) is the possibility that when an emotion takes neuroscience has made clear that mental states are place in a system that has the capacity of forming mem- ultimately activities of the nervous system and are not ories, the brain in that case, it will leave memory traces formed at the cellular level, as was wrongly claimed by that, under certain conditions, can be reactivated by early neurobiologists, but rather at a certain level of only part of the conditions that initially triggered the organization of this system, that of the connectivity emotion. In other words, processing of information at between different brain structures. There is already one level of the multilevel process that is at the source evidence that the basic elements of an emotional behav- of emotion, even when this level is the most elementary ior are organized in the brain stem. This is the case, one, may trigger the full-blown emotion. As a typical for instance, for the oro-motor patterns of taste-induced example, agoraphobic patients can develop a panic at- disgust and pleasure that can even occur in anence- tack when exposed to a place in which they feel trapped. phalic newborns (Grill and Norgren, 1978). However, However, the same pathological emotion will also de- the effect of experience enlarges the range of stimuli velop when they are exposed to alkalosis, whether pro- that are able to trigger these basic responses, and this duced by hyperventilation or by sodium lactate infusion process takes place in the visceral limbic brain (Yama- (Maddock, 2001). The explanation here is that under moto et al., 1994). It is even possible to assess which the effect of these treatments, panic patients experience neural structure is involved in processing environmen- visceral sensations that are reminiscent of those associ- tal cues associated with specific emotional states. In ated with the panic episode. It can be imagined that a the case of a conditioned taste aversion that requires similar process takes place in animals that have been the learning of an association between a novel taste repeatedly exposed to strong or repeated emotional ex- stimulus (the conditioned stimulus, or CS) and visceral perience, especially in unpredictable and uncontrolla- malaise (the unconditioned stimulus, or US), the CS- ble conditions. In the case of sows in maternity stalls, US association seems to take place in the parabrachial E8 Dantzer nucleus, whereas the gustatory cortex modifies the death. However, what all these indicators mean in strength of this association depending on the novelty terms of well-being will remain elusive as long as there of the taste stimulus, and the amygdala is indispens- is little or no research on the basic components of well- able for the expression of conditioned taste aversion being: perceptions, representations, and feelings. It is (Yamamoto et al., 1994). In the case of conditioned fear, certainly time for agricultural research to address this the amygdala seems to be necessary for forming repre- question directly, especially because suitable strategies sentation of discrete emotional stimuli, whereas the are now available as a result of recent advances in hippocampus plays a key role when it is necessary to psychology and neuroscience. form a representation of the context in which the emo- tional response has taken place (LeDoux, 1996). Literature Cited Needless to say, this type of research is still in its infancy in farm animals, with the noticeable exception Antelman, S. M., J. Levine, and S. Gershon. 2000. Time-dependent of the neural basis of the mental representation of social sensitization: The odyssey of a scientific heresy from the labora- stimuli in the ovine brain (Kendrick, 1991; Kendrick et tory to the door of the clinic. Mol. Psychiatry 5:350. Bolles, R. C., and M. S. Fanselow. 1982. Endorphins and behavior. al., 1997). 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S. Held*2, M. Mendl*, K. Laughlin*3, R. W. Byrne†

*Centre for Behavioral Biology, Department of Clinical Veterinary Science, University of Bristol, Langford, Bristol BS40 5DU, U.K. and †Scottish Primate Research Group, School of Psychology, University of St. Andrews, Fife, KY16 9AJ, U.K.

ABSTRACT: The purpose of this paper is twofold. aggression in groups of pigs. Several studies indicate First, it discusses in general terms how animal produc- that some social assessment and recognition take place tion can be affected by the animals’ cognitive abilities; between individuals, allowing them to judge each oth- second, it aims to introduce our work on pig cognition. er’s aggressiveness and to avoid fighting once a domi- We suggest that livestock cognition does not only affect nance hierarchy has been established. However, the production indirectly through its effects on livestock regrouping of previously familiar pigs can also lead to welfare, but also that cognition can have direct effects. high aggression levels. This suggests that pigs may be Direct effects are evident when cognitive abilities limit able to form only short-term social memories, or that some aspects of their social memory are disrupted be- feed intake, for example, or in the recognition of fore regrouping. Our work shows that pigs have well- groupmates or offspring. We illustrate such direct ef- developed spatial memory abilities, which can be dis- fects with two case studies from pig production: volun- rupted by common management procedures. If this tary feed intake after weaning and production losses were also the case for social memory, it could help ex- associated with aggression in groups. Voluntary feed plain increased aggression levels in previously familiar intake after weaning is affected by preweaning experi- pigs after routine procedures. We also show that pigs ences, weaning age, and postweaning practices. Some are able to adjust their foraging behavior depending on studies suggest a link between early environment and the presence or absence of a subordinate, exploitable cognitive development in piglets, as has previously been co-forager that knows where food is. This ability has demonstrated in other species. We suggest this as a potential implications for the way feed is best dispensed possible contributing factor to low feed intake immedi- to pigs such that all group members can maximize ately after weaning. The other case study centers on their intake.

Key Words: Animal Behavior, Livestock Farming, Pigs

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E10–E17

Introduction 2001). Similarly, at weaning under commercial condi- tions without the provision of creep-feed, piglets have In modern production systems, farm animals are con- to learn instantly how to acquire food from unfamiliar stantly and inadvertently presented with many learn- sources. This is in stark contrast to natural conditions, ing and memory challenges. Consider as an example in which the weaning process is prolonged and gradual, chickens housed in large groups in free-range or barn allowing the piglets to explore sources of solid food over systems. We rely on their learning and spatial memory many weeks before they have to rely exclusively on abilities to locate important resources such as food, wa- them (Held and Mendl, 2001). In single-space feeder ter, nesting areas, or familiar subgroups (Mendl et al., systems, the learning challenge can be even greater. The weaners may have to work out how to operate the mechanism that releases the feed before they receive 1The Biotechnology and Biological Sciences Research Council of their first postweaning meal (e.g., Pluske and Williams, Great Britain funded our work. 1996). In addition to these spatial memory and operant 2Correspondence: phone: +44 117 9289571; fax: +44 117 9289582; conditioning abilities, the recognition of conspecifics as E-mail: [email protected]. familiar or unfamiliar, as dominant or subordinate to 3Present address: 1230B Anthony Hall, Dept. of Anim. Sci., Michi- gan State Univ., East Lansing, MI 48824. oneself, or as own or other offspring (e.g., Kendrick et Received May 1, 2001. al., 1997) is also likely to be important if animals are Accepted November 8, 2001. to thrive in modern production systems.

E10 Livestock cognition and production: Pig studies E11 Our paper has two main aims. First, it discusses how also realize that it does. To have control over its environ- livestock productivity can be affected by the animals’ ment, the animal must learn and understand how the cognitive abilities. Postweaning growth-check in wean- environment works and how the environment can be ers and production losses linked to aggression in groups changed to its benefit. And to this end, the animal must of pigs serve as case studies. Second, the paper briefly be able to perceive, store, and use appropriately the reviews some of our recent work on cognition in pigs. pertinent environmental information. We want to emphasize again that in our view welfare Livestock Cognition and Implications is crucially linked to suffering, which implies at least for Production some subjective awareness (see Dawkins, 1990), though the study of cognition as we refer to it here does not Until recently, the word cognitive was predominantly concern itself with the subjective emotional experiences used to describe mental processes that require at least of the animals. However, knowledge of how much farm some form of mental representation of the processed animals understand about their surroundings, how information (Shettleworth, 1998). Or, put differently, they use information about their physical environment it referred to mental abilities that involve information and about the behavior and intentions of their manipulation and storage beyond making simple stim- groupmates can help us understand how they are nega- ulus-response connections. For example, considerable tively or positively affected by their surroundings. For effort has been devoted to investigating whether ani- example, social stress induced by common husbandry mals, as humans, store information about the spatial procedures such as mixing of unfamiliar animals or layout of their environment as mental spatial represen- social isolation from familiar individuals is one of the tations or so-called cognitive maps (Tolman, 1948), or most potent sources of stress in farm animals (Zayan whether they attribute mental states to others (review and Dantzer, 1990). The extent to which animals suffer on primates by Heyes, 1998). in social environments is likely to be affected by their We use a broader definition of cognitive here to in- “social cognitive” abilities, that is, their ability to assess, clude all mental processes animals use to acquire infor- monitor, and predict the behavior and intentions of mation about their environment, to store and recall it, their groupmates (Wiepkema and Schouten, 1990). and to use it in their decision making (cf. Shettleworth, As another example, one might consider animals that 2001). This encompasses learning and memory abili- are capable of forming only short-term memories. They ties, but also what may be called “higher” cognitive would be unlikely to suffer from memories of negative abilities, such as forming cognitive maps, which are experiences in the more distant past. Similarly, if ani- indicated by unlearned, complex, and flexible behavior mals were unable to form expectations based on events patterns (e.g., finding new routes home from an unfa- in the past in the absence of present external and inter- miliar release site). nal (for example physiological) stimuli, it would be un- How, then, can livestock cognition affect production? likely that they suffered because their expectations are We suggest that it does so not only indirectly by the not met (Nicol, 1996). The cognitive abilities of livestock positive and negative effects it has on livestock welfare, can thus inform us about the sorts of environments, or and thus productivity, but that it can also have direct situations, in which the animals might suffer and in effects. In our distinction, “indirect” effects occur when which their welfare may be compromised. cognition, or its failure, leads to some form of stress Direct effects are evident where cognitive abilities response, which may be associated with suffering and limit feed intake, for example, or recognition of compromised welfare and may negatively affect produc- groupmates or own offspring. In the following, we illus- tivity. “Direct” cognitive effects, on the other hand, re- trate such direct effects with two case studies from sult in the animals’ not functioning appropriately with pig production. direct consequences for their productivity (e.g., failure to find food). They may also have welfare-related impli- Postweaning Growth-Check in Piglets cations for productivity, but do not necessarily or always. The first is an experiment by Pluske and Williams The relationship between the cognitive abilities of (1996) on the effects of feed and feeder type on post- farm animals and welfare has already been discussed weaning feed intake. Piglets were weaned at about 30 (see most recently Dantzer, 2001). We know that lack d and put into weaner groups of nine piglets each. Their of predictability of and control over the variable aspects feed intake and growth rates were measured for 4 wk of the animals’ environment increases the physiological after weaning. Feed was dispensed either from single- symptoms of stress (Weiss, 1971; Wiepkema and space feeders wet and dry, or only dry or from Schouten, 1990) and associated loss of productivity such multispace feeders. To operate single-space feeders, the as decreased daily weight gains (e.g., de Jonge et al., piglets had to operate a latch mechanism with their 1996) or reproductive disorders (Varley and Stedman, snouts to make feed drop into a trough. Piglets in the 1994). For the environment to be predictable to an ani- “wet and dry” treatment group had to coordinate the mal in the first place, not only must the environment flow of water from a drinker inside the bowl with the have some predictable properties, but the animal must delivery of food into the bowl. Multispace feeders were E12 Held et al. conventional troughs with enough feeding spaces for accompanied by increasing experience of new sources all nine weaners to feed at the same time. The expecta- of food and gradual nutritional changes. tion was that voluntary feed intake would be greater One common practice thought to decrease the post- in single-space feeders, because fighting at the feeders weaning growth-check by facilitating a more gradual would be reduced, because potential attackers could change in diet and increasing preweaning experience not reach the feeding piglet’s head or ears (Pluske and of new food sources is the provision of supplementary Williams, 1996). Pluske and Williams also predicted creep feed during lactation. Piglets typically start to that adding water to the feed would increase voluntary eat it when they are 2 to 3 wk old (e.g., Pajor et al., feed intake even further. However, against expectation, 1991). However, there is large within- and between- there was no difference in feed intake or weight gain litter variation in the amount of feed consumed before over the 4 wk after weaning. In the 1st wk after wean- weaning, and the effect of providing creep feed on post- ing, the feed intake was actually 27% higher in the weaning feeding behavior and growth rates is not clear- multispace treatment than in the two single-feeder cut (Pluske et al., 1995; Held and Mendl, 2001). Wean- treatments. Weight gain in the 1st wk after weaning ing age also affects voluntary feed intake immediately was significantly lower in the single-space feeder treat- after weaning. Generally, all weaners tend to spend ment with wet and dry feed (Pluske and Williams, less time feeding on the day after weaning than on 1996). As mentioned above, weaners had to learn how subsequent days regardless of weaning age. However, to operate the single-space feeders, and with the “wet the younger the piglets are at weaning, the stronger is and dry” feeders they also had to learn how to coordi- this effect (Held and Mendl, 2001). Efficient feed intake nate water flow and food delivery. Pluske and Williams seems to develop gradually with age and experience, observed that it took the piglets about a week to learn with the younger piglets taking longer to pick up feed- how to use the single-space feeders. They suggest that ing on solids (see also Appleby et al., 1991; Fraser et this led to the initial postweaning growth-check and al., 1995). We suggest that not only age-dependent gut concluded that any potential increase in feed intake maturity, but also cognitive maturity, may contribute caused by lower aggression levels at the single-space to this age effect. feeders was overridden by the piglets’ problems with Both preweaning experiences and weaning age affect the ability of piglets to cope with the weaning challenge. learning how to operate these feeders (Pluske and Wil- In general, weaning practices that mimic certain as- liams, 1996). pects of the natural weaning process ameliorate some of Young commercial weaners often show a characteris- the behavioral problems in young commercial weaners tic growth-check in the 1st wk after weaning, which is such as low feed intake in the first few days after wean- thought to be associated with abrupt weaning at ages ing (Fraser et al., 1995; Held and Mendl, 2001). Early when piglets would still be nutritionally dependent on indications are that piglets reared outdoors up to wean- the sow under natural conditions (cf. Pajor et al., 1991). ing, for example, spend more time feeding at weaning One of the main factors limiting growth immediately (and mixing) into straw yards than indoor-reared pig- after commercial weaning is thought to be voluntary lets (Webster and Dawkins, 2000; Cox and Cooper, feed intake (Pluske et al., 1995). Voluntary feed intake, 2001). Cox and Cooper (2001) also observed that out- in turn, may be determined by cognitive processes such door-reared piglets fought less at weaning and mixing. as learning, as suggested by Pluske and Williams’ The suggestion is that outdoor piglets have better-de- (1996) study. It might also be affected by the disruption veloped social and general cognitive skills than conven- of proper cognitive functioning through separation tionally reared piglets. Similarly, de Jonge et al. (1996) stress or neophobia (cf. Mendl, 1999). Under natural found that female piglets reared under conventional conditions, the number of sucklings begins to fall after indoor conditions up to weaning were more aggressive the 1st wk after birth and continues to decline gradually to each other when weaned into littermate pairs than (Jensen, 1988; Jensen and Rec´en, 1989). The sow begins piglets from enriched farrowing pens. These differences to terminate more and more sucklings (Newberry and carried over into the later postweaning period and pu- Wood-Gush, 1986; Jensen and Rec´en, 1989), and she berty, with the subordinates in pairs originating from spends increasing amounts of time away from the nest conventional conditions (only) showing symptoms of to forage and feed (Jensen and Redbo, 1987; Stangel and chronic social stress (de Jonge et al., 1996). De Jonge Jensen, 1991). The frequency of sucklings continues to et al. (1996) suggest as one possible explanation that decrease steadily, with the steepest fall during the first rearing piglets in relatively barren conditions nega- 4 wk (Jensen, 1988). Feeding on solids by the piglets tively affects the development of their social skills, lead- is well established when they are 4 wk old and increases ing to increased aggression levels. More direct indica- considerably between the 6th and 10th wk after birth tions for a link between rearing environment and cogni- (Newberry and Wood-Gush, 1985; Jensen, 1995). In the tive development in pigs come from a study by de Jong Petersen et al. (1989) study, for example, piglets started et al. (2000). They found that pigs reared in standard to graze when they were between 24 and 36 d old and farrowing and finishing pens had impaired long-term to feed on pelleted feed between 28 and 39 d of age. spatial memory abilities compared to pigs reared in For the piglets, the natural weaning process is thus larger pens with straw (de Jong et al,. 2000). In rats, Livestock cognition and production: Pig studies E13 humans, and nonhuman primates, a strong positive of social assessment, recognition, memory, or other so- relationship has been established between social and cial cognitive functions might help explain why aggres- physical environmental complexity early in life and cog- sion can occur between previously familiar animals nitive development and level of cognitive abilities even- (Mendl, 1999). tually attained (Rosenzweig et al., 1996). It has also been possible to link these effects to changes in brain Cognition Studies with Pigs anatomy and neurotransmitter expression (e.g., Ickes et al., 2000). It might, therefore, be fruitful for wider Spatial Memory application to pig welfare and production to further examine the developmental cognitive mechanisms un- Our first experiment on pig cognition has centered derlying the observed effects of preweaning environ- on such disruption of cognitive processes and on how ment on postweaning feeding (and social) behavior in sophisticated the social cognitive abilities of pigs really pigs. are. Mendl et al. (1997) investigated whether pigs re- membered the location of food hidden in one of 10 possi- Production Losses Caused by Aggression ble areas. The task required the pigs to search for and in Groups of Pigs locate the food in their first visit to the arena (the search trial), to remember the location when returned to their A large proportion of production loss in weaners and home pens (retention interval), and to relocate it during group-housed, nonlactating sows can be attributed to a second visit (the relocation trial). As such it tested aggression (e.g., Tan et al., 1991; Stookey and Gonyou, the pigs’ spatial memory abilities. Pigs did well in relo- 1994). Some of the aggression observed directly relates cation trials. They found the food, making fewer errors to competition over access to resources such as drinkers than would be expected if they had been searching ran- or feeder spaces (cf. Gonyou, 2001). Another source is domly. Pigs remembered food locations over retention the establishment and re-establishment of dominance intervals of 10 min and 2 h. However, how well they hierarchies after mixing and remixing (Meese and Ew- remembered depended on what happened to them dur- bank, 1973; Puppe and Tuchscherer, 1994). At weaning, ing the retention interval. It has long been known that for example, when piglets are mixed into groups of non- stressors can have deleterious effects on memory forma- littermates, an initial intensive period of aggression tion and recall (recent review in Mendl, 1999). Mendl lasting approximately 2 h is commonly observed (e.g., et al. (1997) therefore tested whether this applied to Meese and Ewbank, 1973). This is associated with in- spatial memory in pigs by presenting them with various vestigation of unfamiliar individuals by nosing the face, treatments or “disturbances” in the retention interval. ano-genital region, and belly. The animal that eventu- These were chosen to act as mild stressors or at least ally becomes top-ranking deals out most of the aggres- to lead to changes in arousal, which are also known to sion. Skirmishes and fights can continue for 24 to 48 affect cognitive function (e.g., Kavaliers and Colwell, h. The frequency of aggressive interactions falls at the 1995; in pigs e.g., van Rooijen and Metz, 1987). “Distur- same time as a stable social hierarchy emerges. The bances” also mimicked common husbandry events: pigs decrease in aggression and continued low levels are were confined in a holding pen on the way back to their likely to be the result of the piglets’ recognition and home pen for 3 min at the beginning of the retention memory of each other’s relative social status. During interval, or for 3 min at the end when they were on fighting, pigs may be able to assess each other’s relative their way back into the arena for the relocation trial; competitive abilities (Rushen, 1988; Mendl and Erhard, or they were placed into an unfamiliar isolation pen for 1997), and their efficiency at this social assessing may the whole of the retention interval; or they received determine the frequency and duration of fights. food in the isolation pen after the search trial and were Various factors influence the speed with which a so- then moved on to their home pen for the remainder of cial hierarchy is established and aggression during mix- the retention interval; or they were allowed to explore ing decreases. Weight asymmetry in weaner groups fa- an unfamiliar room for the duration of the retention cilitates social assessment. Rushen (1987), for example, interval. All disturbances negatively affected the per- showed that fights during the first 2 h after mixing formance of the pigs in the relocation trials. Pigs re- lasted longer in groups that consisted of evenly-sized quired more area visits, that is, they made more errors, piglets than in groups in which weights differed more. before relocating the food than in control trials in which Similarly, Mendl and Erhard (1997) showed that it is they had not been disturbed during the retention inter- possible to split weaner pigs into “fast” and “slow” at- val. The treatments, however, did not totally block or tackers according to their individual aggressive charac- destroy memory of the food location. In “disturbance” teristics. When pigs of the same aggressiveness (fast trials, pigs still did better than they would had they or slow) were mixed, the number of escalated fights was been searching totally randomly (Mendl et al., 1997). higher than when pigs of different aggressiveness were Laughlin et al. (1999) built on this to investigate mixed. Again, this implies that some form of social as- whether a more complex spatial memory task might be sessment took place between individuals that allowed more susceptible to interference from environmental them to judge each other’s aggressiveness. Disruption stressors. Rather than requiring the pigs to remember E14 Held et al. and relocate 1 out of 10 locations, they were tested in their advantage, that is, having taken in information a win-shift task in a radial eight-arm maze. Four arms about others, do they change their behavior accordingly were baited with food, the pig was introduced into the to their own benefit? Do they, for example, understand maze, and it searched at random until it had found all what others can and cannot see? four baited arms. It was then returned to its home pen The first experiment addressed the question of for a 10-min retention interval before being allowed whether pigs use the behavior of others in a competitive back into the maze for a relocation trial. Pigs were foraging situation to forage more efficiently (Held et trained to a win-shift strategy, that is, they were re- al., 2000). We adapted the Informed Forager paradigm warded for visiting the four arms that had not been of Menzel (1974) to test the social tactics of two pigs baited in the search trial. Pigs, thus, had to remember foraging together for a hidden food bait. We formed four out of eight rather than 1 out of 10 locations. When eight pairs of one light and one heavy juvenile pig. the pigs reached criterion level of performance on the Lighter pigs were subordinate in a food competition relocation task, tests proper started. There were four test, heavier pigs dominant. Pairs were housed together disturbance treatments, applied during the retention in their home pens. Every morning they were allowed interval. Again, some were chosen to mimic common to forage for food hidden in an arena. Initially, during husbandry events that are thought to be potential stres- training, they foraged alone to learn their respective sors. In the “social encounter” treatment, the test pig tasks. Then, during testing, they foraged together. Both spent the retention interval in a pen where it had con- pigs learned to expect food in one location (bucket) out tact with an unfamiliar pig through a barred gate; in of eight possible ones in an open arena with wooden the “novel” treatment, the pig was led to an unfamiliar screens providing visual barriers between the food loca- outdoor area where it spent the 10-min retention inter- tions. In training trials, they searched the arena indi- val; in the “maze” treatment, the pig was confined in vidually. A different bucket was baited with food at the central part of the radial maze with all arms closed each trial. The subordinate pig in each pair visited the off; and in the “weigh” treatment, the pig was retained arena twice. The same bucket was baited in its second in weigh crate for 10 min then released back into the visit as in its first. In training, the subordinate member maze for its relocation trial. All except the “weigh” treat- of each pair thus learned to search for the food in its ments significantly increased the number of arm visits first visit to the arena (search trial), then relocate the in relocation trials compared to control trials without food location in its second visit (relocation trial). The disturbance (Laughlin et al., 1999). These disruptive dominant pig visited the arena only once and searched effects were stronger than in the study of Mendl et at random for the one bucket of the eight that was al. (1997), as the performance of pigs in the study by baited. The purpose of training, thus, was to turn the Laughlin et al. dropped to that expected of a randomly subordinate pigs into informed foragers or “I-pigs,” but searching animal. One possible explanation for this dif- keep the dominants naı¨ve, or uninformed. When the I- ference is that more complex memory tasks such as pigs had reached criterion level in their relocation task remembering multiple sites rather than just one may we tested the two pigs of each pair together. The I-pig be more susceptible to interference from environmental still got its solitary search trial, but for the relocation stressors (Laughlin et al., 1999). it was now paired with its uninformed, but heavier, These two studies suggest that even common hus- partner. In pair trials, then, the subordinate knew bandry events may act as mild environmental stressors where the food was hidden, the dominant did not. that can have deleterious effects on spatial memory What would the uninformed partner do in the pair performance in pigs and may affect how efficiently they trials: continue searching at random as in solitary trials deal with their environment. If these effects extended during training, or use the knowledge of the subordi- to social memory, this would provide some explanation nates to lead them to the food? We found that unin- for the observation that even in previously familiar pigs formed pigs abandoned searching for food for them- aggression levels sometimes increase after temporary selves and learned to follow their informed partners to removal of individuals for routine management proce- the baited bucket (Held et al., 2000). Once at the baited dures (Luescher et al., 1990; Mount and Seabrook, bucket, they displaced the subordinates and monopo- 1993). Such procedures may be at least temporarily lized the food bait. Following the informed subordinates stressful enough to disrupt aspects of social memory dramatically increased the foraging efficiency of the and resultant social recognition necessitating the re- dominants. In solitary trials, the dominants searched establishment of relative social status when the animal the arena randomly, that is, they visited as many un- is put back into the group (cf. Mendl, 1999). baited locations before finding the baited one as ex- pected of a randomly searching animal. When they fol- Social Cognition lowed the subordinates in pair trials, however, they visited significantly fewer locations before they reached Our work on social cognition has started by looking the baited one (Held et al., 2000). This shows that pigs at how sophisticated the social cognitive abilities of pigs change their behavior in the presence of a companion really are. How much do they understand about the that is a better forager and exploitable. Dominant pigs behavior and intentions of others? Do they use this to must have had some means of assessing relative forag- Livestock cognition and production: Pig studies E15 ing efficiency, probably using their co-forager’s behavior corridor that they had seen the trainer enter. During as the source of information. this training phase, the pigs were also given the experi- These skills are useful if one considers the natural ence of not being able to see the baiting event when a behavior of pigs. Studies on feral and domestic pigs visual barrier was placed in front of their start box. kept in seminatural conditions have shown that domes- Once they had reached criterion level on the training tic pigs retain the social and foraging behavior of their task they were tested in an unrewarded probe trial with wild ancestors (Graves, 1984; Wood-Gush et al., 1990). two companion pigs in the start boxes on either side. They forage for patchily distributed food in family Companions were trained to always enter one particu- groups in large home ranges (Krosniunas, 1979; Janeau lar corridor regardless of the movements of the baiting and Spitz 1995). These family groups consist of one trainer, and also regardless of whether their view of or several sows and their offspring with differentiated the arena was blocked by a visual barrier or not. In rank relationships between them (Mauget, 1981; Pet- tests, a visual barrier was then placed in front of the ersen et al., 1989; Mendl, 1995). Optimality models pre- test pig in the middle and in front of one companion, dict that under such conditions individuals would bene- the “blind” companion. The other, “seeing,” companion fit from the ability to adjust their foraging behavior had full view of the arena and corridor entrances. The flexibly to the presence and behavior of other members test pig in the middle could not see the arena, but it of their group. The optimal tactics in terms of foraging could see the “blind” and “seeing” companions on either returns for each individual depend on its social rank side, and it could see that one had a visual barrier in or relative competitive ability, and the resource distri- front, the same as itself, and the other did not. While bution (Barta and Giraldeau 1998). Generally, when the visual barriers were in place, the trainer crossed food is clumped dominant group members should spe- the arena rattling the bucket noisily to indicate that cialize as scroungers by exploiting subordinates as food- baiting of a corridor was taking place, then exited the finders or producers (Caraco et al., 1989; Vickery et arena quietly to the left or right. After the visual barri- al., 1991). We would, therefore, expect domestic pigs to ers were removed the two companions were released have evolved flexible social tactics, and the cognitive and entered the two corridors that they had been abilities that facilitate them, so as to maximize their trained to enter. Once they had entered, the test pig individual foraging success. Good spatial memory abili- was released. Where would it go? ties allow individuals to store for future use information The hypothesis under test was that test pigs will about the location of profitable food patches. The ability discriminate between “seeing” companions that have to use to their own advantage the pertinent information visual access to the arena and “blind” ones that do not, held by another group member (knowledge exploita- if they were able to take the companions’ visual perspec- tion) allows pigs access not only to food sources they tives. And the resultant prediction was that test pigs found themselves, but also to those located by subordi- would follow “seeing” companions more often because nate others. they understood that only these would lead them to the The next experiment again looked at whether and food. A positive result would show the pigs’ understand- how pigs use social information in a foraging task. Here ing of the behavior of their conspecifics to be a lot more we investigated whether pigs understand what another complex than previously assumed. As such it would can or cannot see. That is, we tested the visual perspec- drastically broaden the range of social situation pigs tive-taking ability of pigs. A detailed analysis of the might perceive as stressful. In situations in which they results is still outstanding, and we can therefore not cannot see, for example, they might use “seeing” others draw any conclusions as yet. However, we have in- as their source of information with resultant expecta- cluded the study here to introduce the type of experi- tions of their environment, which may subsequently mental methodology that allows investigation of more be thwarted. complex social cognitive abilities in farm animals. We adapted a classic primate experiment (Povinelli et al., Implications 1990) to test whether pigs base their foraging choice on the visual perspective of a companion. Visual per- Modern production systems rely on the ability of live- spective-taking experiments work on the assumption stock to learn and remember how to use unfamiliar that animals with perspective-taking abilities will dis- equipment, where to find important resources, and so criminate between individuals that can and cannot see on. That is, they rely on the animals’ cognitive abilities. some critical object or event (Heyes, 1998). We predicted Cognition affects production indirectly through its asso- that when pigs cannot see for themselves where food ciation with subjective awareness, stress, and welfare. is being hidden they should follow an individual that Production may also be directly affected when the cogni- had been able to see where the food was rather than tive faculties of livestock are underdeveloped or dis- follow one that had not. rupted. Our work shows that pigs have good spatial Subject pigs initially learned that they had to watch memory abilities that can be disrupted by common and see where a human trainer went to know which of management procedures. If this extended to social four corridors contained food. Over differing numbers memory, it could help explain increased aggression lev- of training trials, the test pigs learned to enter the els in previously familiar pigs after routine procedures. E16 Held et al. We have also found that pigs adjust their foraging be- Kendrick, K. M., A. P. C. 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D. H. Beermann1

Department of Animal Science, University of Nebraska, Lincoln 68583

ABSTRACT: Mechanisms by which ractopamine and metabolic effects resulting from continuous systemic other beta-adrenergic agonists stimulate skeletal mus- infusion has been conducted in relatively few experi- cle growth are discussed. Oral administration dose-re- ments. Detailed blood flow and hind limb net flux data sponse studies in surgically altered laboratory animals are available for a single beta-agonist, cimaterol. Kinet- provide evidence that indirect endocrine-mediated ef- ics studies and close arterial infusion of cimaterol in the fects are not an essential component of efficacy. Results hind limb of growing cattle demonstrate large transient from age-comparison studies in laboratory animals and increases in amino acid extraction from the circulation livestock species provide evidence that metabolic matu- and similar patterns of net uptake when compared with rity of skeletal muscle may be a critical factor with the contralateral control saline-infused hind limb. Pre- regard to efficacy, suggesting that receptor presence dictions of differential net effects on protein accretion and density are important. Temporal studies demon- using integration of essential amino acid net flux mea- strate the rapidity of responses associated with protein surements are corroborated by quantitative documen- and lipid metabolism changes, and that progressive de- tation of protein mass differences in individual muscles cline in rate of anabolic response in skeletal muscle from treated and control hind limbs. Definitive descrip- results from chronic administration. Associated results tions of specific pathway mechanism(s) of action for that demonstrate progressive beta-adrenergic receptor increasing protein synthesis have not as yet been re- density reductions are observed and suggest, likewise, ported. Therefore, additional research is required for that protein accretion rate and muscle growth rate re- elucidation of cellular and intracellular components of sponses are receptor-mediated. Measurement of in vivo mechanism(s) of action. Key Words: Beta-Adrenergic Agonists, Growth, Muscles

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E18–E23

Introduction without altering organ or bone mass, they are also re- ferred to as repartitioning agents. These repartitioning Rapid preferential increase in skeletal muscle protein phenethanolamines are classified as beta-adrenergic mass is the most striking response to oral administra- receptor (βAR) agonists because they share structural tion of select phenethanolamines in laboratory and similarities and pharmacological properties with the farm animal species. Dose-dependent increases in endogenous catecholamines epinephrine and norepi- weight of individual muscles have been observed in nephine, and many of their metabolic or physiological rapidly growing rats, mice, rabbits, lambs, cattle, pigs, responses can be reduced or blocked by βAR antagonists chickens, and turkeys (Anderson et al., 1991; Moloney (Barnes, 1995). The objectives of this paper are to high- et al., 1991). Although up to 40% greater weight of light our current state of knowledge regarding the muscles has been observed in treated sheep and cattle, mechanisms underlying βAR agonist effects on skeletal magnitude of response varies greatly among phenetha- muscle growth and to identify key areas in which new nolamines studied and is influenced by age, species, research efforts are needed. Results from studies in sex, diet, breed, time to reach plateau, and other factors several species provide clarification of effects on muscle (Beermann, 1993; Mersmann, 1998). Percentage of fiber histology and histochemistry and patterns of RNA, weight or protein mass increase also varies somewhat DNA, and protein accretion. Inferences are drawn from from muscle to muscle, and in a few instances a signifi- these results for effects on cellularity and cellular meta- cant response was not observed in a small number of bolic changes. Influences on aspects of protein synthesis muscles. Because these compounds reduce body fat and degradation, as they pertain to protein accretion in muscle growth, are also described. Indirect vs Direct Actions 1Correspondence: E-mail: [email protected]. Received July 11, 2001. The complexity of hormonal influences on skeletal Accepted November 1, 2001. muscle growth make it necessary to separate possible

E18 Beta-agonists and skeletal muscle growth E19 indirect, endocrine-mediated actions of βAR agonists total weight and protein content of hind limb muscles from direct, βAR-dependent mechanisms of action. In- in lambs fed 10 ppm cimaterol for 3 wk (O’Connor et direct modes of action of βAR agonists include possible al., 1991b). During this same time interval DNA concen- perturbations in circulating concentrations, sensitivity, tration was reduced by 42% and total DNA content of and(or) responsiveness to hormones known to influence the same muscles was unchanged. After 6 wk of treat- skeletal muscle growth. Surgically altered animal mod- ment, differences in RNA concentration and content els that exhibit lack of specific metabolic horomones disappeared, but muscle weight and protein content have provided additional insight. Assessment of direct remained 25% higher in treated lamb muscle. DNA βAR-mediated effects on skeletal muscle includes close concentration remained 25% less. These results suggest (direct) arterial infusion of βAR agonists and saline in that stimulation of satellite cell proliferation and incor- contralateral hind limbs, pretreatment or cotreatment poration into growing muscle fibers is not an essential with general and βAR subtype-specific receptor antago- component of fiber hypertrophy. Although ractopamine nists in vivo, and in vitro incubation of agonists and has been shown to elicit a 30% increase in proliferation antagonists in myoblast, satellite cell, and myotube cul- rate, and in protein and DNA concentrations in cultured tures. Variation in results between in vitro and in vivo mouse C2C12 myoblasts (Shappell et al., 2000), this in studies has provided equivocal results, however. Sev- vitro effect is lost after successive passages of the cells, eral lines of evidence support involvement of βAR in and ractopamine failed to increase DNA or protein in mediation of muscle growth responses. Selectivity and myotubes derived from the C2C12 cells. Grant et al. binding kinetics of phenethanolamines for βAR sub- (1990) observed an increase in proliferation, but not types have been demonstrated (Colbert et al., 1991; fusion, of chick breast muscle satellite cells incubated Spurlock et al., 1994; Smith et al., 1990). Temporal with ractopamine. These results were confirmed in changes in down-regulation or desensitization of beta- studies using turkey satellite cells (McFarland et al., adrenergic receptors is observed with chronic βAR ago- 1995; Shappell et al., 2000) and porcine satellite cells nist administration (Smith, 1989). Myoblast and satel- (Cook et al., 1995). Shappell et al. (2000) concluded lite cell culture studies provide demonstration of activa- “that in turkeys, ractopamine may not work directly tion of βAR signal transduction pathways (Shappell et through the β-adrenergic receptor of muscle cells (and al., 2000), and blockage of responses with nonselective c-AMP) to stimulate changes in muscle carcass traits, and selective antagonists implicate AR dependence β but possibly through extramuscular effects, such as in- (Reeds et al., 1988; MacLennan and Edwards, 1989). creased blood flow and amino acid uptake by muscles.” Use of more recent technologies to create knockout and In summary, in vitro studies fail to provide substantial receptor-modified animal models provides strong evi- evidence in support of βAR-mediated stimulation of sat- dence for receptor-mediated mechanisms. ellite cells as a means by which muscle hypertrophy is initiated by feeding βAR agonists in growing animals. Effects on Muscle Cellularity

The anabolic effects of βAR agonists on muscle in- Indirect Effects of βAR Agonists clude muscle fiber hypertrophy, changes in muscle fiber type frequency, and differential rates of muscle RNA, Stimulation of skeletal muscle growth by βAR ago- DNA, and protein accretion. No evidence of muscle fiber nists may involve modulation of normal endocrine in- splitting or increase in fiber number has been reported. fluences on growth and metabolism. Many studies were Percentage increases in average fiber cross-sectional conducted to investigate these possibilities. Feeding ci- area are similar to percentage increases in muscle materol to growing lambs increased growth hormone weight in lambs (Beermann et al., 1986; Kim et al., and T4 concentrations and decreased IGF-I concentra- 1987) and rats (Maltin et al., 1986), although some tions after 6 wk (Beermann et al., 1987). In subsequent changes in fiber type distribution have been observed. studies, no change in IGF-I, decreased T4, and markedly In general, the increase in hypertrophy of type II fibers decreased (50%) insulin concentrations were observed accounts for the increase in muscle mass without any after 3 wk of treatment (O’Connor et al., 1991a). Young quantitative change in muscle length. Similar changes et al. (1995) also reported no change in IGF-I following have been observed in pigs fed 20 ppm ractopamine chronic administration of clenbuterol to sheep. In cattle (Aalhus et al., 1992). No in vivo increase in myotube or fed cimaterol, an acute decrease in growth hormone muscle fiber number has been reported in βAR agonist- concentration was followed by a chronic increase in treated animals. Lack of response in very young nursing growth hormone and a decrease in IGF-I (Chikhou et lambs (Williams, 1989) and other species suggests that al., 1991). Dawson et al. (1993) observed no significant either muscle fibers lack sufficient βAR number or func- differences in growth hormone or IGF-I concentrations tion to respond early in life or that fractional growth in steers fed cimaterol. The normal episodic pattern of rate in very young muscle fibers is at the maximum. growth hormone secretion was absent in the steers fed Results of temporal studies show that 25% increases cimaterol, however. A similar result was observed in in RNA concentration and 85% increase in total mass lambs fed a high-energy diet with or without cimaterol of RNA occurred in parallel with 30% increases in the administration (Beermann, unpublished data). E20 Beermann Anderson et al. (1991) suggested that the reparti- study (Byrem et al., 1996). Close arterial infusion of tioning effects of phenethanolamines may be due in cimaterol for 21 d increased the rate of blood flow and part to opposing effects on insulin sensitivity in adipose extraction of essential amino acids from the circulation tissue vs skeletal muscle. Ractopamine reduced sensi- in the hind limb in a transient manner. Uptake of amino tivity to insulin of adipocytes from rats (Hausman et acids by the hind limb exhibited a large acute increase, al., 1989) and pigs (Liu and Mills, 1990). Budohoski et followed by a gradual continuous increase to a maxi- al. (1987) observed increased sensitivity to insulin in mum of 160% of controls on d 14, and then a decline soleus muscle receiving chronic treatment with βAR during the last 7 d of infusion. Fractional rates of pro- agonists. Beermann (1987) noted no change in sensitiv- tein accretion were 61 and 130% higher in cimaterol- ity to insulin in lamb hindquarters, using insulin chal- infused hind limbs on d 7 and 14, respectively. Calcula- lenges, despite a 50% reduction in circulating insulin tion of cumulative net protein accretion rates in the concentrations. More refined studies using the hyperin- treated and contralateral saline-infused hind limb pre- sulinemic and euglycemic clamp techniques in cattle dicted 10 to 15% differences in protein mass of the demonstrated a transient (Eisemann and Bristol, 1998) semitendinosus and semimembranosus muscles, re- and chronic (Sternbauer et al., 1998) decrease in insulin spectively. These differences were confirmed with prox- sensitivity with clenbuterol administration. Although imate analysis of the muscles taken on d 21 of the results available to date do not support significant experiment. Close arterial and systemic infusion of changes in insulin sensitivity as the mechanism by clenbuterol in sheep provided similar responses (Au- which skeletal muscle growth is stimulated by βAR rousseau et al., 1993) and confirm results obtained from agonists, more definitive studies are required to clarify cattle fed clenbuterol in which net flux across the portal- these relationships in muscle. drained viscera, the liver, and hindquarters was stud- The anabolic response of muscle in endocrine-altered ied (Eisemann and Huntington, 1993). animals to βAR agonist administration adds support The other approach taken by many to assess direct for mechanism(s) independent of hormone modulation. βAR-mediated effects of beta agonists is to demonstrate Muscle growth is significantly increased in animals dependence on βAR functionality in relation to a variety with genetic growth hormone deficiency (Bates and of response variables (Barnes, 1995). At present there Pell, 1991), in hypophysectomized rats (Thiel et al., are three subtypes of βAR, designated β1, β2, and β3 1987), and in diabetic (McElligot et al., 1987), adrenal- (Liggett and Raymond, 1993). Distribution and density ectomized (Buttery and Dawson, 1987), and hypothy- of βAR subtypes in mammalian tissues is known for roid rats (Beermann, unpublished data). Additive ef- several laboratory animal species (Minneman et al., fects of clenbuterol and growth hormone have been 1979), but as mentioned earlier, few studies provide demonstrated in veal calves (Maltin et al., 1990). Like- information for livestock species (Smith, 1989; Spurlock wise, cotreatment of pigs with salbutamol and growth et al., 1994; Bridge et al., 1998). hormone (Hansen et al., 1997) and ractopamine and Pretreatment and cotreatment of animals with gen- growth hormone (Jones et al., 1989) resulted in additive eral or selective βAR antagonists proved effective. responses, suggesting separate mechanisms of action Reeds et al. (1988) used propranolol, a nonselective β1 account for the independent effects. In summary, indi- and β2 antagonist, and altenolol (β2-specific) in combi- rect effects of βAR agonists, especially effects on endo- nation with clenbuterol feeding in rats. Dosages of the crine-mediated influences on muscle growth, appear to antagonists were 10 and 100 times the concentration be negligible in accounting for the hypertrophic re- of clenbuterol. Effects of clenbuterol on heart weight, sponses. Evidence in support of this conclusion is found fat deposition and energy expenditure were reduced in analysis of results from studies in which direct βAR- with the antagonists, but the muscle hypertrophy re- mediated effects were investigated. sponse was not. Similar results were obtained by Mac- Lennan and Edwards (1989) when similar doses of clen- Direct Effects of βAR Agonists buterol and propranolol were orally administered to rats. Muscle growth response was reduced in an addi- Close arterial infusion of cimaterol is a novel and tional treatment group that received intraperitoneal appropriate model for differentiating systemic and local injection of propranolol with subcutaneous administra- (direct) effects of cimaterol on skeletal muscle metabo- tion of clenbuterol. Blockage of the muscle growth re- lism and protein accretion in the hind limb of cattle sponse provided evidence for involvement of the βAR. (Byrem et al., 1998). Close arterial infusion is the direct, Unfortunately, these experiments do not reveal which continuous infusion of a compound into the arterial cell types the βAR are active in. Therefore, these results circulation of an organism to achieve a local elevated do not directly differentiate direct βAR-mediated and concentration of the compound relative to its concentra- indirect actions or effects. tion in the systemic circulation. Pharmacokinetic anal- Choo et al. (1992) conducted similar studies in which ysis of cimaterol administration in steers demonstrated the β2-specific antagonist ICI-118,551 blocked the ana- that a 10-fold elevation in concentration in the hind bolic responses of clenbuterol in rats. To date, similar limb is theoretically possible (Byrem et al., 1992), and studies have not been reported for livestock species, an appropriate dose was identified in a dose-response but other evidence for βAR dependence is available. Beta-agonists and skeletal muscle growth E21 These include quantitative measurement of cAMP re- administered phenethanolamines that enhance muscle sponse in skeletal muscle slices, cultured myoblasts, growth rate. Examples include total body and urinary and myotubes (Sillence and Matthews, 1994; Shappell excretion of 3-methyl hisitidine, indicative of a 25% et al., 2000) and documentation of desensitization or reduction in fractional degradation rates in rats fed down-regulation of βAR in muscle tissue following cimaterol for 7 d (Eadara et al., 1989); a 27% reduction chronic exposure to βAR agonists (Smith, 1989; Spur- in fractional degradation rate of skeletal muscle pro- lock et al., 1994). teins in steers fed L-644,969 (Wheeler and Koohmaraie, 1992), increased calpastatin mRNA in muscle of steers Effects of βAR Agonists on receiving close arterial infusion of cimaterol (Sun et al., Protein Synthesis and Degradation 1994), and elevated calpastatin activity in sheep and bovine muscle of animals fed the beta-agonist L-644,969 Early investigations failed to provide evidence for (Kretchmar et al., 1990; Koohmaraie et al., 1991; increased fractional rates of protein synthesis during Wheeler and Koohmaraie, 1992; Pringle et al., 1993; significant elevations in protein synthesis rates in rats and Killefer and Koohmaraie, 1994). Similar effects (Reeds et al., 1986) or sheep (Bohorov et al., 1987) fed were reported by Bardsley et al. (1992) and Parr et al. clenbuterol, leading to the suggestion that reduction (1992). Ractopamine-fed pigs fail to exhibit increased in protein degradation rates accounted for the muscle calpastatin activity or mRNA, however (Ji et al., 1991). growth observed. Similar conclusions were reported by Because accurate direct methods for measurement of MacRae et al. (1988), but the possibility exists that protein degradation rates are not available, definitive increases in fractional protein synthesis rates are tran- conclusions regarding effects of repartitioning pheneth- sient. This is supported by the steady increase from 1 anolamines cannot be made at this time. to 14 d, and then a dramatic decline during the subse- quent 7 d in net amino acid flux across the hind limb in Implications cattle treated by the close arterial infusion of cimaterol (Byrem et al., 1998). Using the Snell Dwarf mouse as A rapid increase in fractional rate of muscle protein a sensitive model, Bates and Pell (1991) demonstrated synthesis occurs with oral administration of β-agonists, that clenbuterol and growth hormone both increased and some, but not all (ractopamine), may reduce frac- whole-body and muscle fractional protein synthesis tional protein degradation rates. This appears to be a rates. They noted, however, that growth hormone treat- receptor-mediated response, rather than an endocrine- ment exhibited a larger response. Bergen et al. (1989) dependent one. A critical area for research is the need observed an increase in fractional protein synthesis to identify specific β-receptor subtypes in muscles of rate from 4.6 to 6.7%/d in muscle of pigs fed 20 ppm livestock species and determine their role in growth ractopamine. Similar results were observed by Culham enhancement. Subtype-specific agonists and antago- et al. (1990) using labeled lysine instead of tyrosine. nists must be used to investigate mechanism(s) of regu- Alternative measures that support an increase in pro- lation for these phenethanolamines. New technologies tein synthesis include measurement of mRNA abun- such as creation of β-receptor subtype gene knockout dance for myofibrillar proteins such as actin, myosin, animals, use of specific antibodies to β-receptor sub- and so on. Several investigators using this approach types, and use of transgenic approaches to alter their observed significant increases in response to feeding function, or their expression, will reveal the details of phenethanolamines. α-Actin mRNA abundance was in- the mechanisms by which these repartitioning pheneth- creased by feeding ractopamine to pigs (Bergen et al., anoloamines enhance muscle growth and reduce fat ac- 1989; Helferich et al., 1990; Grant et al., 1993). In- cretion. creased α-actin mRNA abundance was also observed in muscle of sheep fed the beta-agonist L-644,969 Literature Cited (Koohmaraie et al., 1991). Smith and co-workers ob- served elevated myosin mRNA abundance in cattle fed Aalhus, J. L., A. L. Schaefer, A. C. Murray, and S. D. M. Jones. clenbuterol and ractopamine (Smith et al., 1989; 1995). 1992. The effect of ractopamine on myofibre distribution and morphology and their relation to meat quality in swine. Meat These results are interpreted as evidence for increased Sci. 31:397–409. rate of transcription in muscle or as increased stability Anderson, D. B., E. L. Veenhuizen, A. L. Schroeder, D. J. Jones, and of RNA that would lead to greater rates of mRNA syn- D. L. Hancock. 1991. The use of phenethanolamines to reduce thesis in muscle fibers. Whether these events are linked fat and increase leanness in meat animals. In: Proc. Symp. on to protein kinase A activation via cAMP is not known. Fat and Cholesterol Reduced Foods—Advances in Applied Bio- technology Series. Portfolio Publishing Company, New Orleans, Protein degradation is usually determined by differ- LA. pp 43–73. ences between measured fractional protein accretion Aurousseau, B., A. Connell, D. K. Revell, H. J. Rocha, and G. E. and synthesis rates, rates of urinary 3-methyl histidine Lobley. 1993. Hind-limb protein metabolism in growing sheep: excretion, or by measurement of protease activities in Effects of acute and chronic infusion of clenbuterol by close arte- rial and systemic routes. Comp. Biochem. Physiol. C: Comp. muscle tissue (Goll et al., 1998). Common among results Pharm. Toxicol. 106:529–535. of many studies is the indication that muscle protein Bardsley, R. G., S. M. J. Allcock, J. M. Dawson, N. W. Dumelow, J. degradation may be reduced or not affected in animals A. Higgins, Y. V. Lasslett, A. K. Lockley, T. Parr, and P. J. E22 Beermann

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H. J. Mersmann1

USDA/ARS Children’s Nutrition Research Center, Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030-2600

ABSTRACT: β-Adrenergic receptor (βAR) agonists pharmacology of βAR agonists and antagonists in adi- reduce body fat in mammals and birds. Synthetic lipid pocytes is in many cases species-specific. Cloning of metabolism is decreased in βAR agonist-treated ani- individual βAR subtypes allows determination of the mals or in agonist-treated adipocytes in vitro. Degrada- pharmacology of subtypes from that species. For exam- tive lipid metabolism is increased by βAR agonists in ple, the pharmacology of the cloned porcine β1AR, adipocytes in vitro and in vivo. The βAR agonist effects β2AR, and β3AR indicates selected agonists or antago- are blocked by βAR antagonists. In mammalian tissues, nists can be used to assess the proportion of βAR sub- there are at least three distinct βAR subtypes; β-1 types. Nucleic acid sequences of the subtypes were used (β1AR), β-2 (β2AR), and β-3 (β3AR). Individual tissues to prepare probes to quantify the subtype mRNA. The have different proportions of subtypes. For example, pharmacological and mRNA data agree rather closely greater than 85% of the βAR in rat heart is β1AR, in and indicate porcine adipocytes contain over 70% β1AR. guinea pig lung is β2AR, and in rat adipose tissue is The effects produced by a βAR agonist (or antagonist) β3AR. Subtype distribution within a tissue varies with on adipose tissue in vivo depend not only on the species species (e.g., human heart has 65% β1AR and porcine and the adipocyte βAR subtype distribution, but also adipocytes have less than 10% β3AR). There is species on the pharmacokinetics and pharmacodynamics of the variation in the amino acid sequence of a βAR subtype. compound in that species, including blood flow to the Thus, it is expected that some βAR agonists would have tissue, and the multiple metabolic and endocrine effects different effects in the same tissue in different species of the compound in other tissues of the body. In short, because of different βAR subtype distribution and(or) it is expected that individual βAR agonists would have amino acid sequence. In support of these concepts, the somewhat different effects in different species.

Key Words: Adipocytes, Beta-Adrenergic Receptors, Growth, Metabolism

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E24–E29

Introduction β-adrenergic Receptors

The βAR are cell-surface receptors that interact with Understanding the mechanisms by which β-adrener- stimulatory G-proteins (Gs-proteins) to activate the en- gic receptor (βAR) agonists modulate adipocyte metab- zyme adenylate cyclase. Adenylate cyclase synthesizes olism and growth requires familiarity with the family cAMP, the intracellular messenger for responses to of membrane-bound βAR subtypes. This paper de- βAR. The physiological ligands for the βAR are the scribes the physiological functions and biochemical neurotransmitter norepinephrine and the adrenal med- characteristics of these receptors, including subtypes, ullary hormone epinephrine. Norepinephrine, in addi- tissue distribution, and species variation. This informa- tion to being a central nervous system and peripheral tion provides the basis for discussing animal responses sympathetic nervous system neurotransmitter, is pres- to oral administration of βAR agonists and effects on ent in the plasma and can act as a hormone. There are adipocyte metabolism and fat deposition. Secondary vs a large number of synthetic analogs of norepinephrine primary mechanisms of action are also discussed. and epinephrine, some of which are agonists (bind to the receptor to activate it and stimulate βAR-mediated activity), some of which are antagonists (bind to the receptor, but do not activate it, and thus compete with 1Correspondence: E-mail: [email protected]. agonists and inhibit βAR-mediated activity), and some Received June 11, 2001. of which are partial agonists (bind to the receptor and Accepted October 17, 2001. partially activate it). The βAR are present on almost

E24 βAR and adipocyte growth E25 every cell type and control an exceptionally large num- marization of studies in various species using a variety ber of physiological and metabolic functions. Examples of βAR agonists is noteworthy (Moloney et al., 1991), as are regulation of heart rate and contractile force, blood is a summarization of multiple studies on one agonist, pressure, bronchial muscle tension, uterine contrac- ractopamine, in pigs (Watkins et al., 1990). Attempts tion, and glycogen and lipid degradation. As functions to amalgamate data across species and agonists, albeit of norepinephrine, epinephrine, and synthetic analogs a gross oversimplification, indicate a hierarchy of re- were explored, incredible complexity arose that gave sponses across species with sheep ≥ cattle ≥ turkeys > rise to the concept of more than one type of receptor. pigs > chickens (Mersmann, 1998; Moody et al., 2000). First, α-adrenergic receptor (αAR) and βAR functions Perhaps one contributing factor to this hierarchy is were separated, then the βAR was separated into two that some species are closer to their maximal growth subtypes, the β1AR and β2AR. Later, the αAR was response because of intense selection for growth rate; separated into subclasses and, finally, evidence was consequently they respond to a lesser extent (e.g., chick- accrued for a β3AR. ens respond much less than sheep). The βAR is a continuous protein chain, > 400 amino acids in length. It contains seven relatively hydrophobic Factors Leading to Variable Responses segments that traverse the cell membrane (transmem- brane domains) and are connected by three extracellu- In addition to the variable response observed in dif- lar and three intracellular segments or loops. Also, ferent species, even when administered the same βAR there is a N-terminal extracellular tail and a C-terminal agonist, there is considerable variation caused by the intracellular tail. The ligand-binding site is composed use of different βAR agonists. This is to be expected of amino acids contributed by several of the transmem- because an individual agonist has not been tailored to brane domains. The binding of the G-proteins occurs all the species in which it has been studied. The rate primarily through intracellular loop 3. Phosphorylation of absorption, the metabolism of the agonist by various is one mechanism by which receptor activity is dimin- organs, and the excretion rate of the agonist or its deriv- ished, and there are phosphorylation sites on the C- ative(s) dictate the concentration achieved and main- terminal segment. Selected reviews about βAR struc- tained over time in the plasma, and ultimately at the ture and function include: Stadel and Lefkowitz (1991), target organ sites. The many aspects of these pharmaco- Caron and Lefkowitz (1991), and Strosberg (1992, 1993, dynamic properties of the agonist, coupled with the 1997). Reviews with emphasis on agricultural species blood flow to the target organs, dictate the pharmacoki- are Mersmann (1989b, 1998), Mills and Mersmann netic properties of the agonist. For a given βAR agonist, (1995), and Moody et al. (2000). many of these pharmacodynamic properties are ex- pected to vary among species. Animal Responses The pharmacodynamic properties of a particular βAR agonist administered to a particular species are ex- The use of βAR agonists to alter body composition of pected to be influenced by genetic-, sex-, and age-caused growing animals raised for meat production was re- variation in drug metabolism and delivery systems. ported in the 1960s (Cunningham, 1965). Beginning in Furthermore, the βAR population on target organs or the early 1980s, synthetic βAR agonists were shown to cell types may be expected to vary not only with species, affect body composition in several species, including but also with age and perhaps with breed or sex. Varia- cattle, chickens, pigs, sheep, and turkeys (Ricks et al., tion in the βAR number on a target cell would be ex- 1984). It was not until late 1999 that a βAR agonist, pected to cause variation in the response to a βAR ago- ractopamine, was approved in the United States for use nist. Examination of the response to a βAR agonist in one of these species, the pig. Oral administration of within a species must use a dose × response design any of a number of βAR agonists usually causes an because the response of one variable (end point) may increase in daily gain accompanied in many cases by a have a different relationship to dose than the response slight decrease in feed intake with a consequent im- of another end point. Thus, for ractopamine-fed pigs, provement in efficiency of gain. The striking observa- the half-maximal response for improvement in gain is tion is that βAR agonists preferentially increase skele- achieved at ∼1.5 mg/kg feed, whereas the half-maximal tal muscle protein mass, which is usually accompanied response for improvement in dressing percentage is by a decrease in adipose tissue mass, with little or no achieved at ∼10 mg/kg feed (Moody et al., 2000). Like- change in internal organ size. All of these responses wise, comparison of a given βAR agonist across species are quite variable and depend on the species, perhaps can only be validly accomplished when the studies uti- the breed, the animal age, the sex, the specific βAR lize a dose × response design because a dose that may agonist used, the dose, the diet, and many other factors be maximal in one species may be far less than maximal that vary in the numerous published studies. There are in another species. Factors influencing response to βAR multiple studies of the effects of βAR agonists on animal agonists have been discussed previously (Mersmann, growth; among the summaries of these results are Han- 1989b, 1995, 1998). rahan (1987), Bergen and Merkel (1991), Beerman Dietary components may influence absorption of the (1994), and NRC (1994). A rather complete tabular sum- drug, they may modify the transport of the drug, or they E26 Mersmann may increase or decrease the metabolism or excretion of each increase adipocyte degradative lipid metabolism the drug. With an anabolic agent such as a βAR agonist, and decrease synthetic lipid metabolism in vitro. One the protein content of the diet must supply sufficient enigma is that, overall, the βAR agonists that cause essential amino acids for optimal protein synthesis to changes in body composition, including decreases in allow expression of the anabolic properties of the βAR fat deposition, are not particularly potent or effective agonist on muscle synthesis (Mitchell et al., 1991; Duns- agonists to modulate adipocyte lipid metabolism in hea, 1993; Moody et al., 2000). vitro (Mills and Mersmann, 1995). These results may represent artifacts of the studies in vitro because the Fat Deposition βAR agonist clenbuterol, which does not increase por- cine adipocyte lipolysis in vitro, increases plasma fatty Mammals fed βAR agonists generally have decreased acid concentrations when infused in vivo (Mersmann, carcass fat, as indicated by backfat thickness or 1987; Hu et al., 1988). An increase in plasma nonesteri- trimmed fat from commercial cuts (Moloney et al., fied fatty acids usually results from an increase in adi- 1991). As genetic selection for lean carcasses has contin- pocyte lipolysis. ued, there is less subcutaneous fat, particularly in pigs. Thus, because the control pigs have less fat, a treatment βAR Subtypes to reduce fat deposition is more difficult to detect. Re- gardless, in most βAR agonist-treated animals, there There are three well-documented βAR subtypes: is a numerical decrease in carcass fat, even if the data β1AR, β2AR, and β3AR. Evidence for these subtypes do not always achieve statistical significance. When initially came from classic pharmacological studies of comparative slaughter techniques are employed to cal- organ, tissue, and cell systems using not only norepi- culate deposition rates, the rate of muscle or protein nephrine and epinephrine, but also numerous synthetic deposition is increased, whereas the rate of fat deposi- agonists and antagonists. These approaches were fol- tion sometimes is not significantly reduced (Mitchell et lowed by ligand binding to membranes from various al., 1991; Dunshea, 1993; Chwalibog et al., 1996). If tissues. It became obvious that certain tissues con- measured at a constant time (age), the βAR agonist- tained primarily one receptor subtype. For instance, treated animals weigh more and have more muscle rat heart contains mostly β1AR (> 90%), guinea pig mass; thus, even an unchanged rate of fat deposition bronchi contain mostly β2AR (> 85%), and rat adipo- translates to a lesser percentage of fat per animal. If cytes contain mostly β3AR (> 90%). These tissues, or measured at constant weight, the βAR agonist-treated membranes prepared from them, were prototypes used animals have more muscle mass; even with an un- to study an individual βAR subtype with minimal inter- changed fat deposition rate, there is less fat mass be- ference from other subtypes. The individual βAR sub- cause the βAR agonist-treated animals grow faster and types were eventually cloned and expressed in cells are measured at a younger age. The overall conclusion without endogenous βAR so that the properties of a is that βAR agonists decrease carcass fat in growing single βAR subtype could be studied without interfer- mammals and birds. ence from other subtypes. Using prototypical tissue or membrane preparations and, later, cloned subtypes ex- Adipose Tissue Lipid Metabolism pressed in cells, individual agonists and antagonists could be classified regarding their specificity for a sub- The βAR agonists markedly increase adipocyte de- type. A few agonists and antagonists were discovered gradative lipid metabolism. Activation of the βAR that had considerable specificity for a single subtype. causes an increase in cAMP that activates protein ki- Thus, CGP 20,712 is considered a β1AR antagonist, nase A, which in turn phosphorylates hormone-sensi- ICI 118,551 is considered a β2AR antagonist, and BRL tive lipase. Phosphorylated lipase is the activated form 37,344 is considered a β3AR agonist. These subtype- that initiates the degradative process, lipolysis. Fatty specific agonists and antagonists have become classic acids are produced and, to a large extent, exported from reagents to classify the βAR subtypes in other tissues the adipocyte to be used as oxidative fuels by other and species. However, such results must be considered tissues. Fatty acid synthesis and the esterification of tentative because the subtype specificity of these classic fatty acids into triacylglycerol, the primary energy stor- reagents is based solely on their interaction with proto- age molecule in the adipocyte, are both inhibited by typical receptors. Usually nothing is known about the βAR agonists. Thus, an increase in catabolic and a de- subtype specificity of the classic agonist or antagonist crease in anabolic lipid metabolic processes in the adi- in the species of interest. Overviews of the βAR sub- pocyte would both lead to decreased hypertrophy of the types with emphasis toward agricultural species are adipocyte with a consequent decrease in fat deposition. Mills and Mersmann (1995), Mersmann (1995, 1998), Much of these data have been summarized (Smith, and Moody et al. (2000). 1987; Mersmann, 1989b, 1995, 1998; Mills and Mers- The βAR subtypes have now been cloned from a num- mann, 1995). ber of species. These include the porcine β1AR (Cao et The synthetic βAR agonist isoproterenol and the al., 1998), β2AR (Liang et al., 1997), and β3AR (Smith physiological agonists norepinephrine and epinephrine and Mills, personal communication) and the bovine βAR and adipocyte growth E27 β1AR (Ha et al., 1999), β2AR (Einspanier et al., 1997), has 72% β1AR and pig lung has ∼ 67% β1AR, rather and β3AR (Pietri-Rouxel et al., 1995). The β1AR is the than the > 85% β2AR in the prototypical guinea pig largest of the three subtypes with approximately 460 lung (McNeel and Mersmann, 1999). Based on ligand- amino acids. The β2AR has approximately 420 amino binding studies, bovine muscle has almost exclusively acids and the β3AR has approximately 410 amino acids. β2AR (Sillence and Mathews, 1994), whereas bovine The homology for the three subtypes within a given (Sillence and Mathews, 1994; Van Liefde et al., 1994) species is usually between 45 and 60%, whereas the and ovine (Bowen et al., 1992) adipose tissue have pre- homology for a given subtype is rather high across spe- dominantly β2AR. These ligand-binding results must cies (usually > 70%). Although there is strong homology be considered tentative because a single classic β1AR for a given subtype across species, the variation in and β2AR antagonist was used to classify receptor sub- amino acid sequence allows for considerable variation types and there has been no verification that the antag- in ligand binding or functional properties of these ho- onists used have any specificity for the bovine or ovine mologous receptors. Consequently, it should not be as- β1AR or β2AR. Extensive pharmacological measure- sumed that a classic agonist or antagonist designated ments using porcine adipose tissue, isolated adipocytes, as specific for a βAR subtype (based on activities with and ligand binding to adipocyte membranes have prototypical receptors) will have the same subtype spec- clearly indicated that the βAR present do not respond ificity in another species. For example, cloned human, to classic βAR subtype-specific agonists or antagonists, mouse, and bovine β3AR expressed in Chinese hamster as do prototypical βAR subtypes (Mersmann et al., ovary (CHO) cells respond to propranolol (a classic βAR 1993; Mills and Mersmann, 1995). The cloned porcine antagonist) in different ways; propranolol is an antago- β1AR (Cao et al., 1998) and β2AR (Liang et al., 1997, nist for the mouse β3AR but a partial agonist for the 2000) have been expressed in CHO cells so that the human and bovine β3AR (Pietri-Rouxel et al., 1995). pharmacology of a single porcine βAR can be assessed. The uniqueness of the pharmacology of these porcine Tissue Variation in Distribution βAR subtypes has been verified; the classic β1AR antag- of βAR Subtypes onist CGP20712 is highly specific for the porcine β1AR, but the classic β2AR antagonist ICI 118,551 has no Although the prototypical tissues contain a predomi- specificity for the porcine β2AR, whereas a classic β3AR nant proportion of a single βAR subtype (e.g., rat heart agonist, BRL 37,344, is relatively specific for the porcine has > 90% β1AR), other tissues have a more equal distri- β2AR (Scott Mills, personal communication). These re- bution of the β1AR and β2AR subtypes. The β3AR has sults clearly indicate the species specificity of the phar- a limited tissue distribution, being present at high con- macological response of a βAR subtype and also force centration in adipose tissue of some species (e.g., rats the conclusion that use of classic βAR subtype-specific but not humans or pigs), and is expressed at substantial agonists or antagonists to classify βAR subtypes in levels in selected areas of the gut (Mersmann, 1998). other than the prototypical species produces, at best, Considering only the β1AR and β2AR, in humans there tentative conclusions. is approximately 35% β1AR in adipose tissue, 27% Using compounds specific for the cloned porcine β1AR β1AR in the lung, and 20% β1AR in the liver (Sano et and β2AR, it has been estimated that porcine adipo- al., 1993). cytes have ∼75% β1AR (Scott Mills, personal communi- The βAR subtypes may change with growth/age of cation). Using quantitative measurement of the mRNA the animal. For example, undifferentiated preadipo- for porcine βAR subtypes, it has been estimated that cytes from the rodent-derived clonal cell line 3T3- porcine adipocytes contain ∼73% β1AR, ∼20% β2AR, F442A have predominantly β1AR with essentially no and ∼7% β3AR (McNeel and Mersmann, 1999). Based β3AR, whereas the differentiated adipocytes have > on mRNA concentrations, the proportion of βAR sub- 90% β3AR (Feve et al., 1991). Thus, the age of an animal types in other pig tissues was estimated (McNeel and may dictate the response to an exogenous βAR agonist Mersmann, 1999), but these results have not been veri- because the βAR are less developed, or the proportion fied by ligand binding. There are no reports of βAR of the βAR subtypes may be altered during develop- subtypes in cattle or sheep tissues using appropriately ment. Of course, many other physiological responses of controlled mRNA measurements or ligand binding as- the animal may change with age to modify the pharma- sessments using agonists or antagonists verified as spe- codynamic and pharmacokinetic properties of a βAR cific for the βAR subtype of interest. agonist in vivo. Secondary Mechanisms βAR Subtypes in Agricultural Species In addition to direct effects on adipocytes, systemi- Compared with a prototypical tissue wherein a single cally delivered βAR agonists can have a number of sec- βAR subtype predominates (e.g., rat heart with ≥ 90% ondary effects that could alter adipocyte metabolism β1AR), the proportion of that receptor subtype in the and consequently growth. Blood flow to a variety of same tissue from another species may be lower. For tissues is increased in the presence of a βAR agonist; example, based on mRNA concentrations, the pig heart this has been demonstrated in cattle (Eisemann et al., E28 Mersmann

1988), sheep (Beermann et al., 1987), and pigs (Mers- region for the porcine β1-adrenergic receptor gene. J. Anim. Sci. mann, 1989a). The concentration of a number of hor- 76:1720–1721. Caron, M. C., and R. J. Lefkowitz. 1991. Structure-function relation- mones may be altered; some of these can directly or ships. In: J. P. Perkins (ed.) The Beta-Adrenergic Receptors. pp indirectly affect the adipocyte. For example, the lower 41–72. Humana Press, Clifton, NJ. plasma insulin in sheep chronically fed a βAR agonist Chwalibog, A., K. Jensen, and G. Thorbek. 1996. Quantitative protein might lead to decreased lipogenesis and increased lipol- and fat metabolism in bull calves treated with β-adrenergic ago- ysis (Beerman, 1987). Metabolic rate may be increased nist. Arch. Anim. Nutr. 49:159–167. by βAR agonists, although chronically treated calves Cunningham, H. M. 1965. Effect of epinephrine and nicotine on pro- tein and fat metabolism of pigs. In: K. L. Blaxter (ed.) Energy (Zimmerli and Blum, 1990), pigs (Yen et al., 1991), and Metabolism. pp 29–36. Academic Press, New York. sheep (Rikhardsson et al., 1991) do not have elevated Dunshea, F. R. 1993. Effect of metabolism modifiers on lipid metabo- metabolic rates. These and other secondary mecha- lism in the pig. J. Anim. Sci. 71:1966–1977. nisms have been discussed previously (Beerman, 1987; Einspanier, R., C. Gabler, and W. Kloas. 1997. Expression of the Mersmann, 1989b, 1995, 1998). beta2 adrenergic receptor in the cattle. GenBank Accession No. Z86037. Eisemann, J. H., G. B. Huntington, and C. L. Ferrell. 1988. Effects Conclusions of dietary clenbuterol on metabolism of the hindquarters in steers. J. Anim. Sci. 66:342–353. Although there is considerable evidence for effects of Feve, B., L. J. Emorine, F. Lasnier, N. Blin, B. Baude, C. Nahmias, several βAR agonists on adipocyte lipid metabolism, A. D. Strossberg, and J. Pairault. 1991. Atypical β-adrenergic receptor in 3T3-F442A adipocytes. J. 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S. E. Mills2

Department of Animal Science, Purdue University, West Lafayette, IN 47907

ABSTRACT: Receptor-mediated signals are tightly clinically in the treatment of human disease and may regulated by feedback inhibition and act to prevent sig- well limit the effectiveness of βAR ligands used to pro- nal overload and to reset the receptor to a changing mote livestock production. Pigs fed βAR ligands show environment. Short-term regulation (uncoupling) of a rapid response in growth and feed efficiency that tends beta-adrenergic receptors (βAR) involves receptor phos- to peak during the first 7 to 10 d but declines thereafter phorylation and uncoupling of the receptor from the G toward zero by approximately 6 wk. A similar pattern protein Gs. Chronic exposure to ligand leads to reduced was reported in rats fed clenbuterol and was accompa- receptor number (down-regulation), which results from nied by a 50% reduction in βAR in skeletal muscle. a combination of receptor internalization and degrada- Feeding clenbuterol every 2nd d prevented the decline tion, and decreased mRNA abundance. The extent of in the response to clenbuterol and gave a growth re- βAR regulation is subtype-specific with a rank order of sponse that was equivalent to daily dosing. These data β2AR > β1AR > β3AR. Differences between species are suggest that strategies to prevent or circumvent βAR expected also because amino acid sequences differ. Un- down-regulation may prolong the agonist response. In- coupling and down-regulation of βAR in pig tissues has termittent dosing of pigs may present logistical prob- been demonstrated in vivo and in vitro, although skele- lems. An alternative approach may be to incrementally tal muscle exhibits a blunted response compared with increase the dose of βAR ligand to compensate for the adipose tissue and changes in mRNA abundance have decline in response or to augment the ligand response not been observed. Desensitization presents a challenge by inhibiting the inhibitory G protein Gi.

Key Words: Beta-Adrenergic Receptor, Pigs, Regulation

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E30–E35

Introduction The short-lived nature of chemical messengers ex- tends to receptor molecules and signaling responses. Living organisms operate in a state of imbalance and Readjustments in these systems are multifaceted and the neural and endocrine systems have evolved to mod- include uncoupling of receptor responses from signaling ify the rates of physiological processes to maintain ho- events, degradation of receptors, and up- and down- meostasis. Whether we have binged on chocolate-chip regulation of signaling molecules that affect the pri- cookies, fasted for 10 d, or suddenly felt the need to run mary signaling pathway. a marathon, these two systems interact to fine-tune The need to regulate can be appreciated by consider- metabolic rate. One of the hallmarks of both regulatory ing several examples of disruption of regulation, includ- systems is the short-lived nature of the signals pro- ing goiter, which results from overstimulation by thy- duced. The half-life of neurotransmitters is measured roid-stimulating hormone when thyroid hormones are in seconds, whereas that of circulating hormones may deficient (Hetzel, 1983), tumor growth due to mutations be minutes to days. The rationale for the need for a in signaling molecules (Sebolt-Leopold et al., 1999), and system that is short-lived is to permit our systems to cholera, which results from overstimulation of G-pro- reset in order to meet the next challenge. tein signaling cascades (Fishman and Atikkan, 1980). Endocrine signals are tightly regulated so that animals can adapt to changing needs and to prevent the develop- ment of pathologies associated with unchecked sig- 1Journal paper no. 16655 of the Purdue Univ. Agric. Res. Prog. naling. This paper was presented at the 2001 Midwest meetings of the Am. Soc. Anim. Sci. [79 (Suppl. 2):53 (Abstr.)]. Beta-Adrenergic Receptors and Signaling 2Correspondence: Lilly Hall (phone: 765-494-4845; fax: 765-494- 9346; E-mail: [email protected]). Received April 5, 2001. The beta-adrenergic (βAR) belong to a large family Accepted October 30, 2001. of seven transmembrane-domain proteins that couple

E30 Down-regulation of beta-adrenergic receptors E31 and signal through guanine nucleotide binding proteins The βAR ligand-mediated desensitization has been (G-proteins). Three βAR subtypes have been cloned and demonstrated in several in vitro systems, including all signal in a similar manner. The binding of agonist ventricular strips (Temma et al., 1985), adipocytes promotes the interaction between the intracellular do- (Mills and Orcutt, 1989; Ding et al., 2000), and a variety mains of the βAR and the heterotrimeric G-protein Gs of cell lines (Su et al., 1980). The primary alteration is at (Strosberg et al., 1993). This interaction catalyzes the the level of the βAR itself or βAR-Gs coupling, because exchange of GTP for GDP in the Gα subunit and leads adenylyl cyclase activity assessed by direct activation to the dissociation of Gα from Gβγ. The activated Gα or via Gs is not impaired under conditions in which activates adenylyl cyclase, catalyzing the synthesis of signaling through the βAR is reduced (Mukherjee et cAMP from ATP. The cAMP in turn activates protein al., 1975; Hausdorff et al., 1990). Increased inhibitory kinase A (PKA), leading to subsequent phosphorylation activity of Gi was ruled out in studies in which Gi was events. Activation of adenylyl cyclase is terminated by inactivated with pertussis toxin, but recent studies the hydrolysis of GTP by an intrinsic GTPase and Gα have implicated some role of Gi in contributing to the recombines with Gβγ. Opposing the stimulatory effect reduced signaling through adenylyl cyclase (Tepe and of Gs proteins on adenylyl cyclase is the inhibitory ac- Liggett, 2000). tion of hormones that signal through Gi, including the α2-adrenergic and adenosine receptors. Mechanisms of Agonist-Induced Desensitization The ability of βAR ligands to increase cAMP is mea- sured in terms of potency (sensitivity) and efficacy Multiple mechanisms contribute to desensitization (maximum response) and can be modified by altering and can be divided into acute “uncoupling” responses any of the inputs affecting adenylyl cyclase activity. and chronic “down-regulation” responses (Figure 1). For instance, chronic treatment with a βAR antagonist Uncoupling of the βAR from Gs prevents signaling to increases the density of βAR in the plasma membrane adenylyl cyclase and represents the post-stimulation and results in an increase in sensitivity (Glaubiger and sequence that prepares the βAR for re-stimulation. Un- Lefkowitz, 1977), whereas chronic treatment with a coupling is initiated within seconds to minutes of ago- βAR agonist reduces the density of βAR and reduces nist exposure and the extent of uncoupling is closely sensitivity (Pecquery et al., 1984). The response to βAR related to the potency of the ligand to stimulate ade- can also be increased by ligands that signal through Gi nylyl cyclase (Su et al., 1979; Su et al., 1980). Desensiti- (Cumbay and Watts, 2001). In this case, the increased zation by partial agonists is directly related to their response is not a result of increased density of βAR, but ability to activate adenylyl cyclase, whereas antago- rather a component of G-protein signaling. In contrast, nists do not induce uncoupling (Su et al., 1980). Uncou- conditions in which Gi is elevated result in reduced pling is further characterized as being quickly and com- efficacy of βAR ligands (Tepe and Liggett, 2000). pletely reversible upon agonist removal, does not re- quire the synthesis of new protein, and does not involve Desensitization of βAR Signaling a change in receptor density (Su et al., 1980; Hausdorff et al., 1990). Desensitization is defined as the attenuation of re- The primary mechanism for uncoupling involves sponse despite continued presence of the stimulus and phosphorylation of the βAR, which interferes with cou- has been demonstrated in a variety of systems. One pling to Gs. In addition, the uncoupled βAR has a re- early report from the Lefkowitz group demonstrated duced affinity for agonists that decreases the signaling that frogs given four doses of isoproterenol over 24 h had response at submaximal ligand concentrations. Two ki- greater than 50% reduced stimulation of erythrocyte nases are involved in βAR phosphorylation. The first adenylyl cyclase (Mukherjee et al., 1975). Similar is a true “feedback” regulation by PKA, which is acti- changes were induced in adipocytes from hamsters vated by cAMP from adenylyl cyclase (Hausdorff et al., given once-daily subcutaneous injections of epinephrine 1990). Phosphorylation by PKA alters the conformation for 6 d (Pecquery et al., 1984). The primary alteration of βAR and its affinity for Gs. Because cAMP is a second was a reduced maximal response to isoproterenol for messenger for a number of hormones, regulation by the stimulation of glycerol release, cAMP accumulation, PKA can phosphorylate and uncouple the receptor in- and adenylyl cyclase activity, with no appreciable volved in initiating the signal (homologous regulation) change in sensitivity to isoproterenol. Reduced respon- or other G protein-coupled receptors (heterologous reg- siveness of adipocytes has been observed also in rats ulation) (Richelsen and Pedersen, 1985; Hausdorff et with a norepinephrine-secreting tumor (Prokocimer et al., 1990). A second family of kinases, the G-protein al., 1988). However, in this instance reduced sensitivity coupled receptor kinases (GRK), includes soluble en- to isoproterenol was more pronounced than was re- zymes that mediate only homologous desensitization. duced maximal response. The effects of chronic eleva- The specific enzyme that phosphorylates Ser and Thr tion of catecholamines shown for adipocytes and eryth- residues in the carboxy tail of the βAR is βARK1 (β- rocytes appear to be a general phenomenon and likely adrenergic receptor kinase 1). The βARK1 enzyme extend to most, if not all, tissues that express βAR. phosphorylates only agonist-occupied βAR, or that con- E32 Mills kinase to the βAR. The agonist-occupied receptor has a high affinity for βARK1, resulting in phosphorylation of multiple residues. Phosphorylation by βARK1 in- creases the affinity of the βAR for one of the arrestin family of proteins (β-arrestin). The binding of β-arrestin causes steric hindrance of βAR association with Gsα and interrupts signaling. Receptors are subsequently reactivated by a process that involves sequestration and internalization into cytoplasmic vesicles, dephos- phorylation, and recycling to the plasma membrane (Jockers et al., 1996; Kallal et al., 1998). Desensitization may also involve activation of Gi and inhibition of ade- nylyl cyclase (Tepe and Liggett, 2000). It has been shown that PKA phosphorylation of βAR increases the affinity for Gi, thus effectively switching signaling path- ways from Gs to Gi. Down-regulation is defined as a decline in the total number of βAR and contributes to desensitization from chronic exposure to agonists. Down-regulation develops more slowly than uncoupling, taking hours to days in cell culture systems (Su et al., 1980; Hausdorff et al., 1990) or in vivo (Mukherjee et al., 1975; Pecquery et al., 1984). The rate of decline in βAR binding sites is related to potency and efficacy of the drug (Su et al., 1980; Hausdorff et al., 1990). High concentrations of full agonists cause a more rapid decline in βAR binding, but even low concentrations of full agonists and partial agonists lead to loss of βAR number that is equivalent to that induced by a full agonist. Antagonists do not cause down-regulation, and in fact they have been shown to induce a compensatory increase in βAR num- ber and responsiveness (Glaubiger and Lefkowitz, 1977). Recovery from down-regulation is also slower than that from uncoupling, taking days in cell culture (Su Figure 1. Cellular signals involved in β-adrenergic re- et al., 1980). The total βAR protein content of a cell is ceptor uncoupling and down-regulation. Initial steps in the sum of the rates of synthesis and degradation, and desensitization are similar for uncoupling and down-reg- it seems that both processes may contribute to the de- ulation. Agonist binding leads to phosphorylation of the cline. Degradation of βAR involves endocytosis and hy- βAR by the kinases PKA and BARK I (A). The βAR subse- drolysis in lysosomes. The process of endocytosis re- β β quently binds -arrestin ( AR-Arr), which effectively un- quires βARK1 and β-arrestin, the latter of which binds couples the receptor from further G-protein activation to caltherin in coated pit regions of the plasma mem- (Gsα). The βAR is reactivated and recycled to the plasma brane and initiates endocytosis (Lefkowitz, 1998). If membrane following dephosphorylation in intracellular βARK/β-arrestin are involved in both uncoupling and vesicles. (B) Down-regulation differs from uncoupling in down-regulation, it is not clear whether these processes that internalized βAR are degraded in lysosomes and are always linked or a switch is made to induce down- mRNA abundance for βAR may be decreased. Both pro- regulation. It now appears that endocytosis is far more cesses contribute to a reduction in total βAR in the complex than simply βAR degradation and includes plasma membrane. βAR reactivation and the activation of alternative sig- naling cascades (Lefkowitz, 1998). formation of the βAR induced by agonist binding (Lig- Down-regulation of βAR occurs in cell lines express- gett and Lefkowitz, 1994). ing βAR genes that lack promoter elements, indicating The sequence of events that results in βAR phosphor- that changes in expression are not required for down- ylation has been determined (Liggett and Lefkowitz, regulation. Nonetheless, chronic exposure to βAR li- 1994). Upon ligand binding and Gs activation, Gsα asso- gands leads to a decrease in mRNA abundance for the ciates with adenylyl cyclase, leaving Gs(βγ) tethered to β1AR and β2AR (reviewed by Liggett and Lefkowitz, the membrane in close proximity to the βAR. The 1994) and likely contributes to the full decline in βARK1 enzyme binds Gs(βγ), effectively targeting the βAR protein. Down-regulation of beta-adrenergic receptors E33 Subtype Differences a twofold decrease in sensitivity to ractopamine (Mills et al., 1990). In vitro incubation of pig adipocytes with Phosphorylation is a key element of desensitization, isoproterenol reduced total βAR binding sites 40% but so it is logical that subtype difference in phosphoryla- did not affect mRNA abundance for either the β1AR or tion may determine rates and extent of desensitization. β2AR (Ding et al., 2000). For the human βAR, the β2AR contains two potential PKA sites and 11 βARK sites, whereas the β1AR has Implications of βAR Desensitization only one PKA site and 10 βARK sites. The β3AR is the most divergent, having no PKA sites and only three There is little question that desensitization can affect βARK sites (Hausdorff et al., 1990). Small differences physiological processes and therapeutic approaches to have been noted between the β1AR and β2AR (Zhou the treatment of disease. For instance, a decrease in et al., 1995), but the β3AR is resistant to short-term βAR density and responsiveness to catecholamines in uncoupling responses (Granneman, 1992; Liggett and heart muscle may precipitate heart failure (Choi and Lefkowitz, 1994). Chronic down-regulation of the β3AR Rockman, 1999). Furthermore, βAR agonists are often may not involve the mechanisms outlined above be- administered to counteract desensitization in heart cause it appears to be absent in some cell types (Nantel muscle, but the treatment may instead hasten desensi- et al., 1993; Liggett and Lefkowitz, 1994) but has been tization and the loss of effectiveness of administered shown to develop slowly in others (Nantel et al., 1993). drugs (Colucci et al., 1981). The pig βAR have been cloned and the putative phos- Animals treated with βAR agonists exhibit growth phorylation sites are similar to those found in human patterns indicative of desensitization. Weight gain in βAR (Liang et al., 1997; Cao et al., 1998; Smith et al., rats fed clenbuterol was increased 20% over the first 7 2000). The pβ1AR has one potential PKA site and 10 d, but the response diminished to zero by d 14. A similar Ser + Thr residues in the carboxy tail for βARK phos- pattern for weight gain was shown for pigs fed racto- phorylation. The pβ2AR may have only one PKA site pamine (Williams et al., 1994) or L-644,969 (Convey et and 14 Ser + Thr residues. We would predict, therefore, al., 1987). Maximal responses were observed in the first that desensitization would be similar in the pig as de- week to 10 d followed by a decline in the response to scribed for other species. The pβ3AR is similarly defi- each drug to near zero by wk 7. Further, we favor the cient in potential phosphorylation sites and its regula- idea that down-regulation of βAR in adipose tissue pre- tion would be expected to differ from the β1AR and vents full expression of the βAR response and may lead β2AR. to little or no change in the rate of fat accretion (Duns- hea et al., 1993; Liu et al., 1994). If desensitization Desensitization of Pig Tissues limits the effectiveness of βAR ligands, it is doubtful that effects are fully lost because removal of the ligands Clenbuterol and cimaterol are two βAR agonists that from the diet invariably results in a reversal of the induce muscle hypertrophy in rodents and are accompa- favorable change in body composition (Jones et al., nied by a down-regulation of βAR in muscle tissue 1985; Mills and Orcutt, 1989). (Rothwell et al., 1987; Kim et al., 1992). Cimaterol simi- larly induced βAR down-regulation in pig skeletal mus- Circumventing Desensitization cle (Smith, 1989). Chronic feeding of ractopamine to swine reduced βAR in adipose tissue 25% after1dand The question of greatest interest is, does desensitiza- up to 50% by 7 d (Spurlock et al., 1994). Curiously, tion limit the effectiveness of βAR agonists to modify however, significant βAR down-regulation was not de- growth and composition, and how can desensitization tected in skeletal muscle and only trends for a decrease be prevented or overcome? One approach tested in rats were observed. Smith (1989) also did not detect down- was to feed clenbuterol every 2nd d, thereby allowing regulation of βAR in skeletal muscle of pigs fed racto- the βAR system an opportunity to reset (McElligot et pamine. Why do the adipose and muscle tissues appear al., 1989). Intermittent feeding seemed to prevent de- to differ? One possibility is that βAR subtypes are dis- sensitization because growth rate was stimulated by tributed and regulated differently or that ractopamine clenbuterol to a similar extent at each supplement in- preferentially targets one subtype. Indeed, the β1AR terval over 14 d, whereas rats fed clenbuterol continu- accounts for 75% of the βAR in adipose tissue and only ously showed no response by d 14. Overall, growth rate 50% in skeletal muscle (Liang and Mills, 1999; McNeel and skeletal muscle gain were the same in the two and Mersmann, 1999). It is unknown whether these groups, but the intermittently-fed rats received 50% subtypes respond differently to elevated ligands, but less clenbuterol. It is not known whether intermittent ractopamine does not show specificity for one subtype feeding is advantageous over longer time periods. A over the other. One other possibility is that ractopamine second approach may be to incrementally increase the might stimulate the synthesis of new βAR protein in dose of drug to compensate for the decline in response. skeletal muscle and effectively mask down-regulation. Drs. Schinckel and Richert at Purdue are taking this In adipose tissue, the decrease in βAR is consistent approach to determine whether a greater response can with the observed 40% decrease in rates of lipolysis and be achieved with ractopamine. E34 Mills Implications beta 2-adrenergic receptor desensitization and sequestration. Delineation using chimeric beta 3/beta 2-adrenergic receptors. Compensatory desensitization is a well-characterized J. Biol. Chem. 271:9355–9362. Jones, R. W., R. A. Easter, F. K. McKeith, R. H. Dalrymple, H. M. response to chronically elevated agonist concentration Maddock, and P. J. Bechtel. 1985. Effect of the beta- adrenergic that results in a net loss of beta-adrenergic receptors agonist cimaterol (CL 263,780) on the growth and carcass charac- (βAR) from the plasma membrane and reduced tissue teristics of finishing swine. J. Anim. Sci. 61:905–913. response. The ability of βAR agonists to affect pig Kallal, L., A. W. Gagnon, R. B. Penn, and J. L. Benovic. 1998. Visual- growth and alter body composition may well be limited ization of agonist-induced sequestration and down-regulation of by βAR desensitization. Experimental approaches a green fluorescent protein-tagged beta2-adrenergic receptor. J. Biol. Chem. 273:322–328. should be investigated to determine the extent to which Kim, Y. S., R. D. Sainz, R. J. Summers, and P. Molenaar. 1992. desensitization limits drug response, and to develop Cimaterol reduces beta-adrenergic receptor density in rat skele- practical approaches to optimize the effectiveness of tal muscles. J. Anim. Sci. 70:115–122. exogenously administered compounds. Three βAR sub- Lefkowitz, R. J. 1998. G protein-coupled receptors. III. New roles for types have been cloned from mammalian species and receptor kinases and beta-arrestins in receptor signaling and desensitization. J. Biol. Chem. 273:18677–18680. most tissues express more than one subtype. It is not Liang, W., C. A. Bidwell, S. K. Williams, and S. E. Mills. 1997. Rapid known which βAR subtypes are linked to growth re- communication: Molecular cloning of the porcine β2-adrenergic sponses in the pig or how βAR agonists affect the bal- receptor gene. J. Anim. Sci. 75:2824. ance of synthesis and degradation of each subtype in Liang, W., and S. E. Mills. 1999. Distribution of beta-adrenergic different tissues. Answers to these questions may lead receptor subtypes in pig tissues. FASEB J. 13:A258. to targeting strategies that improve the efficacy of Liggett, S. B., and R. J. Lefkowitz. 1994. 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W. D. Purcell1

Agricultural and Applied Economics Department, Virginia Tech, Blacksburg 24061

ABSTRACT: Price systems in the beef industry have attribute identification, the price-based system will con- failed to communicate needed changes to producers. tinue to disappear. Producers will need to look at alter- The lack of alignment between what is produced and natives and find the approach that allows them to par- what consumers want and are willing to pay for pushed ticipate in a beef system that profits from better serving beef demand down nearly 50% from 1980 through 1998. a discriminating and changing consumer. Packers have Ineffective grades allowed the price-driven system to revised their business models as contractual arrange- fail, and current public policy blocks needed changes ments and alliances have allowed them to coordinate in quality grades for fresh beef. Product attributes of what they buy with needs for new consumer-friendly products and to accomplish at least a modicum of qual- importance to consumers, such as tenderness, have not ity control. Once low-cost commodity operations, the been identified and brought into the pricing process. In large beef packers are now more nearly quality-ori- the presence of a failed pricing system, producers have ented. The huge investments in new products and mar- looked to pricing grids, contracts, and vertical alliances kets they have made are the catalysts that have turned to be paid for value and for investments in genetics and the beef demand picture to the positive. Those invest- technology. The future will see continued competition ments are important to the industry and will set the between price-based systems and non-price means of future of the beef industry for the year 2010 and beyond. coordination and quality control. If grades are not mod- The future of beef can be very positive if needed changes ernized and new technology brought in to allow more are made to ensure the consumer is better served.

Key Words: Communication, Contracts, Coordination, Prices

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E87–E93

Introduction are supposed to communicate to producers and prompt needed changes in production. In practice, the price Demand for beef decreased dramatically from the late system failed as a communication mechanism. Cattle 1970s through 1998. The primary reason for the de- were being sold on broad averages, with little or no price creases was a growing divergence between the fresh differences for value. Important product attributes such beef offering and what the consumer wanted as consum- as tenderness were not identified by the quality grades, ers’ lifestyles changed. Beef lost over 30% of its market and price premiums or discounts cannot be generated share and cattle inventories decreased from over 132 if tenderness is not identified and brought into the price million head to 91 million head. Many producers were discovery process. In the presence of a failed pricing forced out of business and others consolidated to reduce system, producers will increasingly look to non-price costs. The prices consumers were willing to pay were systems such as vertical alliances for more accurate not sufficient to support a beef industry that had grown identification of cattle value and for potential returns to 43 kg in per capita consumption, retail weights, in to investments in new genetics and new management 1976. By 1993, the inventories were much smaller and practices. Cattle producers of all sizes will need to un- per capita consumption was down to 30 kg (Livestock derstand the new approaches in order to evaluate Marketing Information Center, 2000c). whether and how those approaches fit their particular In theory, the price system should keep such a diver- needs and operations. gence from happening. Price premiums and discounts Background

1Correspondence: Mail Code 0401 (phone: 540-231-7725; fax: 540- 231-7622; E-mail: [email protected]). The value of calves from beef cow herds is derived Received July 30, 2001. from beef consumers’ dollars. It is a derived value that, Accepted October 30, 2001. in the long run, will compensate cattle producers as a

E87 E88 Purcell consumer-level prices. The price consumers pay to clear the market is pushed down from supply-side pressure when seasonal and cyclical forces increase slaughter levels. The prices consumers will pay for the quantity being offered are also pushed down when they do not like the offering and demand decreases. The supply- side pressures on price are related to time, and although they can last for several years as the cattle cycle moves through the liquidation phase, they do eventually come to an end. When decreasing demand is the problem, the price pressure will last as long as the consumer is dissatisfied with the product offering. Demand will Figure 1. Price spreads for beef, 1970 through 2000. decrease until the problem is fixed and until the sector Source: Livestock Marketing Information Center (2000a; moves to a “consumer-driven” status. http://lmic1.co.nrcs.usda.gov/, Members Only section, Sustained problems on the demand side mean the spreadsheet BFSPREAD.xls). communication system inherent to any marketplace is not working. As lifestyles and eating behavior change, what the consumer wants and is willing to purchase will residual claimant. The beef system can be described as also change. There must be a communication channel a supply chain that adds value as the product moves to send a message to producers, a message that both up toward the consumer. Middlemen try to extract a identifies the needed changes and provides an economic per-animal margin, and retail price minus the com- incentive for producers to make new investments and bined margins determines the calf price to producers. adjust their production programs. If there are no effec- The individual producer is a residual claimant and has tive communications and no recognizable set of eco- no power to set selling prices. nomic incentives, what producers offer can increasingly In the current marketplace, one firm may perform diverge from what consumers want. A market failure two or more of the economic functions along the supply has then occurred, and this “failure” is the dominant chain, but the net result is the same. Consumers place feature of the beef business in the United States during a value on the fresh beef offering and decide whether the decades of the 1980s and 1990s. to buy. If beef is offering the quality, consistency, and convenience the consumer is looking for, prices get bid Pricing Systems up. If the fresh offering falls short in ways of importance to consumers, the packages stay in the counter and Price-driven marketplaces with separate profit cen- prices are pushed lower. The ability to always extract ters between the producer and consumer dominate the a margin sufficient to cover all costs weakens in moving historical literature on the markets in any food sector. from the retailer down through the cattle feeder, but all In theory, the consumer will send a message, a price the “middlemen” want to extract targeted per-animal premium or a price discount, down through the various operating margins. profit centers to the producer. Price is bid up at retail on If all the middlemen margins were constant over any product offering that finds favor with the consumer. time, producer-level prices would move directly with The display counter empties, and the retailer bids up changes at the consumer level. But the margins be- the price to get more of that particular product offering. tween the producer and the consumer are not constant. Increasing prices and profits pull new investments into They tend to move up with overall price inflation as the sector, and growth in terms of increasing per capita labor, packaging, fuel, refrigeration, and transportation supply, and, therefore, in increasing per capita con- costs increase. Figure 1 shows wholesale, retail, and sumption, is seen. The pattern in beef in the 1960s and total price spreads for beef across recent decades. The into the early 1970s was one of growth, and increasing spreads are in dollars per hundredweight, retail demand was the catalyst. weights. When retailers have to lower price to get the con- The price spreads are reported by the USDA and sumer to take the product offering, just the opposite the “farm-wholesale” margin is essentially the packer/ occurs. Lower prices are pushed down toward the pro- processor margin and the “wholesale-retail” margin or ducer, and downsizing, disinvestment, and a loss of spread is the margin extracted by the retailer. It is market share are the inevitable results. Dissatisfaction clear that the price spreads have increased over time. with the product offering brings decreases in demand, Either costs, profits to middlemen, or some combination and producer-level price decreases can be significant of the two are increasing. The expanding spreads are and sustained. one source of downward pressure on producer prices Before turning to what is needed to prompt correct (Livestock Marketing Information Center, 2000c). valuations and prices to beef cattle producers, it is use- The other primary source of downward pressure on ful to look at what happens when there is no effective producers’ calf and cow prices comes from decreases in communication in price-driven systems. It is problems Communicating value to cattle producers E89

Figure 3. Per capita consumption and deflated Prices (CPI, 1982 through 84 = 100) for beef, 1960 through 2000. Figure 2. Price and quantity changes along a constant Source: Livestock Marketing Information Center (2000b; demand surface. Source: Original development. Price is http://lmic1.co.nrcs.usda.gov/, under Members Only price per unit such as cents per pound. Quantity is often section, spreadsheets RETMT.xls and SUMQ.xls). per capita consumption in pounds. demand line running through 1980, 1985, or even 1990. Any move down and to the left is a decrease in demand, in this pricing dimension that cause market failures in and the new demand line is below the prior line. If economic systems. that pattern persists over time, low prices will force producers out of business, reduce per capita supply and A Look at Demand per capita offerings, and the result is a loss of market share. That loss is apparent in the decrease in per capita Demand is the schedule of quantities that consumers consumption from near 43 kg in 1976 to 30 kg in 1993. will take at alternative prices. If this schedule is plotted Table 1 presents a demand index developed for the with price on the vertical axis, the curved line that Beef Promotion and Research Board as a part of an results slopes down and to the right. Any price/quantity industry demand study launched in 1997. The index combination along the line represents the same level was calculated using 1980 as a base period (index value or strength of demand. If demand is not changing due of 100) and it declined to 51.45 in 1998. To facilitate to changes in preferences, in incomes, or in prices of monitoring any progress from the 1997 study date, a substitutes, then any increase in quantity to Q2 will transformed index with 1997 = 100 is also shown in bring a market-clearing price, P2, that is below the ear- the table. In 2000, the index with 1997 = 100 suggests lier P1 price. If quantity decreases due to seasonal or demand was 6.7% above demand in 1997. cyclical influences, the smaller quantity, or Q3, will The dramatic and sustained decline in demand, run- clear the market at a higher price than P1, price P3. ning unabated from 1979 or 1980 through 1998, is evi- Quantity and price change over time with no change dence of a massive market failure. There was no sig- in demand. In particular, quantity as measured by per nificant response to consumers’ changing wants and no capita consumption can change, and will change, with significant and badly needed modification of the fresh changes in per capita supply, but such changes in no beef product offering for nearly 20 yr. way indicate whether the level or strength of demand has changed. A Failed Pricing System Figure 2 illustrates these possibilities. If Q1,Q2, and Q3 are measuring per capita consumption, it is apparent The beef pricing system of the 1980s and 1990s ap- that per capita consumption, which is a measure of per pears to have failed in its role as a communication capita supply, can change while demand is constant. A system. Many fed cattle sell each week in a few hours new set of price-quantity coordinates is representing a at one average price with no significant price discrimi- different level of demand only if they are not on the nation in spite of a huge variation in final quality and curve labeled “Demand” in Figure 2. value of fed cattle in any pen and in every feedyard. Figure 3 presents a scatter plot with inflation-ad- Producers who have invested in superior genetics, pro- justed prices of Choice beef at retail on the vertical axis. duction technology, and management expertise get lit- Per capita consumption as a measure of quantity is on tle or no reward if they sell at the average price. There the horizontal axis. The average prices paid for the is a subsidy flowing from producers of the high-value yearly quantity offered are identified by years in the cattle in the overall quality and value distribution to plot. It is apparent that any negatively sloping line producers of the low-value cattle. Schroeder and Graff running through 1998, for example, is far below the (2000) estimate the subsidy at $30 per animal. E90 Purcell Table 1. Beef demand index, 1980 through 2000a

Constant Per-capita Deflated demand consumption, price, price, Index Index Year lb cents/lb cents/lb 1980 = 100 1997 = 100

1980 76.6 283.5 — 100.00 193.5 1981 78.3 258.2 274.13 94.19 182.3 1982 77.1 247.0 280.71 87.99 170.3 1983 78.6 235.0 272.40 86.27 167.0 1984 78.5 227.3 271.82 83.26 161.1 1985 79.3 212.7 268.59 79.19 153.3 1986 78.9 206.9 270.78 76.41 147.9 1987 73.9 209.8 298.35 70.32 136.1 1988 72.7 211.6 304.99 69.38 134.3 1989 69.3 214.2 323.76 66.16 128.0 1990 67.8 214.5 332.04 64.60 125.0 1991 66.8 212.0 337.63 62.79 121.5 1992 66.5 203.3 339.29 59.92 116.0 1993 65.1 203.1 346.94 58.54 113.3 1994 67.0 190.9 336.57 56.72 109.8 1995 67.5 186.6 333.81 55.90 108.2 1996 68.2 178.6 329.89 54.14 104.8 1997 66.9 174.2 337.14 51.67 100.0 1998 68.1 170.0 330.42 51.45 99.6 1999 69.1 173.4 325.02 53.35 103.3 2000 69.4 178.1 323.17 55.11 106.7 aSource: Original calculation. Per capita consumption data and nominal prices can be found at the Livestock Marketing Information Center (2000; http://lmic1.co.nrcs.usda.gov/, Members Only section, spreadsheet SUMQ.xls and RETMT.xls). The Consumer Price Index used to deflate the prices is available at the Bureau of Labor Statistics (http://stats.bls.gov/cpihome.htm).

The reasons for the long-running decrease in beef for lack of marbling. To the extent that marbling is demand are widely known, and they are associated with correlated with tenderness, this reduction in marbling the effectiveness of the price system (Schroeder et al., requirements for the Choice grade may have contrib- 2000). Moving through the 1980s, consumers were in- uted to the tenderness problem of the 1980s and 1990s. creasingly concerned about fat and cholesterol levels The economic purpose of grades in any food product and they were willing to pay for convenience in the is to identify and categorize the product attributes of form of a reduction in meal preparation time (Purcell, importance to the final consumer. In beef, if any attri- 1993). Perhaps most important, consumers of fresh beef bute such as tenderness varies significantly within a or beef entrees in their favorite restaurant wanted a grade such as Choice, the necessary conditions for effec- consistent, high-quality eating experience. Instead of tive price signals are not met. There is no price signal consistency, consumers were treated to a fresh beef mechanism for consumers to use in sending a message offering with Choice cuts that were often tough. A “qual- to either encourage production of tender beef or discour- ity concerns” strategy workshop that examined the age production of tough beef. There is no categorization findings of the national beef quality audit conducted in within Choice beef to which a price signal can be 1991 ranked excessive external fat, excessive seam fat, attached. A market failure results and consumers sim- low overall palatability, and inadequate tenderness ply walk past the beef counter in the grocery store and numbers 1 through 4 in the top 12 list of concerns about order something else at the restaurant, even at their beef quality (NCA, 1992). favorite steakhouse. The study by Lusk et al. (1999) The failure of the pricing system to provide coordina- indicated that consumers will pay significant premiums tion between what is produced and what consumers for guaranteed tenderness in beef, but tenderness is want has roots in both the public and private sectors. not identified as part of the quality grade specifications. Public quality grades for beef are administered by the Across the 1980s and into the 1990s, the beef quality Agricultural Marketing Service (AMS) of the USDA on grades have been inadequate and outdated. Without a user fee basis. The policy of AMS is to change the reference to the widespread concern about consistency grades for any food or fiber product only when the indus- across USDA graders, the grade specifications were try demands change. The last significant change was simply not sufficient. The volatile industry reaction to in the late 1970s when the industry supported a move the late 1990s move to get B-maturity cattle out of the to “flatten” the marbling requirements against age of Choice and Select grades made the USDA and AMS the animal. The argument was that declining age of leaders even more cautious about changing grades, and the steer and heifer at slaughter would compensate the beef quality grade program continues largely un- Communicating value to cattle producers E91 changed since the 1970s. Helming (1996) stated that The Move to Non-Price Systems the USDA grading system stifled development and use of branded beef products and blocked value-based beef The recent move to non-price means of coordination production and pricing from becoming a reality. Helm- and quality control was very predictable. The profit ing (1996) wanted to see the public grades replaced with opportunities coming from better serving consumers, private grades that included specifications on product long ignored, were becoming so apparent that they de- attributes such as tenderness that influence the quality manded attention. In-house analysts at the large pack- of the consumer’s eating experience. However, this call ers were looking at the massive decline in demand and for a dramatic change was buffered by observations on starting to analyze what could happen if that trend the usefulness of the quality grades in selling to other could be reversed. In pork, where grades and the price countries, and exports were starting to grow rapidly in system had suffered a similar market failure, the move the middle of the decade. There was no widespread away from price-driven systems occurred at an incredi- support for elimination of public grades and no proposal ble pace. The largest firm, Smithfield Foods, Inc., ac- knowledges publicly that some 70% of their kill is from for new approaches emerging from private firms along their own genetics in company-owned and contract pro- the supply chain. duction programs. As the company achieved a measure The beef industry of late 2001 will likely not demand of coordination and quality control, they were able to a change in grades. Those cattle owners feeding cattle meet a company goal to move into export markets such with overall quality below the mean level of the quality as Japan, and company profits surged. The message distribution are receiving a significant subsidy. This was not lost on other meat packers and processors. part of the feeding industry will not want a change In beef, smaller firms in the further processing busi- such as tenderness scores within the Choice and Select ness moved first into pre-cooked product and attacked quality grades. They fought the proposed change in the the problems of quality inconsistency and too much B-maturity issue. time in meal preparation. Contracts, price grids, verti- The public sector that includes federal agencies and cal alliances, and other forms of non-price coordination the land-grant university system, perhaps predictably were developed to help processors buy what they needed because of long-standing USDA policy, has been slow for a new product line. Non-price approaches to vertical to make progressive change in the beef grades. As a coordination and quality control are now a reality. As result, the price-driven system could not prompt the the price-driven system is increasingly being aban- vertical coordination and related quality control so nec- doned, processors are identifying and buying the genet- essary to improvement in the offerings to the consumer ics and the quality control they need to serve the con- market, especially in fresh beef. Market share for beef sumer with a new and branded product offering. In declined sharply across the 1980s and 1990s. Cattle 2001, the large packers have largely abandoned a busi- producers were forced out of business as inflation-ad- ness model built around being a low-cost commodity justed retail prices plunged, even in the presence of operator and are using price grids, contractual procure- declining per capita offerings, and prices were forced ment, vertical alliances and quality control and are down at the cow-calf level. offering branded and cooked product lines. The private sector reluctance to make changes was It is not impossible for the price-driven system to predictable and grows out of the way the sector has been come back and compete with these non-price systems, structured under the traditional price-driven system. but it is not likely to happen. At a minimum, it would There are a number of profit centers between the origi- appear that the quality grades would need to be mod- nal cattle producer and the consumer. Inter-level rela- ernized or widely used private sector replacements tionships have been adversarial for decades. Producers would need to be developed, and neither of these devel- and producer groups complained when told by industry opments appears to be very likely. Instead, vertical alli- ances are sorting carcasses using in-house measures of experts that they should make sure the new product tenderness to identify carcasses acceptable for branded development work was done and pointed fingers at the and quality-assured product lines. Because these pro- packers. But the large packers were following a busi- cesses and procedures are developed with private mon- ness model of being the low-cost provider of commodity ies, there will be less interest in participating in open- beef, and they were efficient in that role. The packers market, price-driven systems in which prices are aver- had no strong motivation to spend billions of dollars on ages and do not reflect the premiums that can be gener- new product lines when the raw material supply (fed ated from branded beef products. cattle) was heterogeneous, unpredictable, and variable in quality and value. Without quality control to allow What Producers Will Need to Do them to brand and stand behind their products, the large packers could see no way to build a fence around Producers looking to be paid for value will have, the benefits of investments in new products and in new therefore, two broad approaches they can follow. They markets. The industry drifted and needed corrections can campaign hard to fix the price system to give it a were not made. chance to compete with the non-price approaches, or E92 Purcell they can go to the contracts, grids, and vertical al- Implications liances. The price system will not be easy to fix. It will need The move to non-price systems for beef products will a change in grades toward a more refined identification not block all opportunities to make needed changes to of product attributes that influence value to the con- give the price-driven system a chance. Modernization sumer, and a corollary move to objective and high-tech of beef grades, including taking into account varying measures will be essential. Any progressive proposal levels of tenderness, would provide needed quality con- for change splits the cattle-feeding sector and seed stock trol. The price-driven model can be effective if it has producers. Firm-level profits and opportunities to in- the grades, technology, and policies to allow it to work. crease those profits are still usually seen as more im- In the meantime, it will be the non-price alternatives portant than a progressive change for the industry as that offer some chance of “pricing to value,” and it will a whole. That was clear in the sometimes bitter dialogue be the price grids, contract arrangements, and vertical surrounding the move away from B-maturity cattle. alliances that will likely continue to grow in impor- With leading meat scientists saying tenderness is a key tance. Producers wishing to be compensated for high- factor in consumer acceptability and describing USDA value cattle will need to take a look. If better coordina- tion and quality control is the result of these new sys- quality grading as “mass inspection” and lacking in tems, both the individual cattle producer and the beef precision, those feeding the B-maturity cattle still put industry as a whole will face a more promising future. their bottom line first and some tried to block the change (Tatum et al., 1999). In the near term, the chances of Literature Cited an industry consensus to incorporate tenderness scores or other more refined measures in the quality grades are Helming, B. 1996. Is it time to eliminate the USDA beef grading small. The price-based system will, in all probability, system? In: Proc. Natl. Cattlemen’s Beef Assoc. Live Cattle Mar- continue to languish and falter. keting Council, Nashville, TN. In the presence of a failed price system, producers Hudson, T., and W. Purcell. 2001. Risk sharing and compensation guides for managers and members of vertical beef alliances, need to look at the non-price coordination opportunities Research Inst. on Livestock Pricing Research Bull. 1-2001, Virg- to learn what will be valued in a consumer-driven mar- nia Tech., Blacksburg. ketplace, and try to offer what consumers want. De- Livestock Marketing Information Center. 2000a. Available at: http:// pending on geographical location and the constraints lmic1.co.nrcs.usda.gov/, Members Only section, spreadsheet BFSPREAD.xls. Accessed October 26, 2001. that geography place on genetic mix, the need is to find Livestock Marketing Information Center. 2000b. Available at: http:// an opportunity that will reward producer-level invest- lmic1.co.nrcs.usda.gov/, Members Only section, spreadsheet ments in technology and management expertise. Indi- RETMT.xls. Accessed October 26, 2001. vidual animal identification and feedback is essential Livestock Marketing Information Center. 2000c. Available at: http:// lmic1.co.nrcs.usda.gov/, Members Only section, spreadsheet and is increasingly available. No one grid or contract SUMQ.xls. Accessed October 26, 2001. arrangement or alliance will fit every producer’s re- Lusk, J., J. Fox, T. Schroeder, J. Mintert, and M. Koohmaraie. 1999. source base and opportunities, so there is a need to look Will consumers pay for guaranteed tender steak?, Research Inst. and compare. Fortunately, the alliances recognize the on Livestock Pricing Research Bull. 3-99, Virginia Tech, Blacksburg. reality of geographical and other influences on genetic NCA. 1992. Executive summary. National Beef Quality Audit. Engle- mixes, and the alliance within a producer’s region will wood, CO. probably fit some or most producer needs. The best Purcell, W. D. 1993. Price and non-price factors affecting demand for situation, arguably, is one in which producers get a meats: Emphasis on beef, Research Inst. on Livestock Pricing Research Bull. 4-93, Virginia Tech, Blacksburg. significant share of the “premium pool” associated with Schroeder, T. C., and J. L. Graff. 2000. Estimated value of increased high-quality cattle moving into premium-priced, pricing accuracy for fed cattle. Rev. Agric. Econ. 22:89–102. branded product lines, and some vertical alliances are Schroeder, T. C., T. L. Marsh, and J. Mintert. 2000. Beef demand offering producers who maintain ownership of their cat- determinants, report prepared for the Joint Evaluation Advisory Committee, Department of Agricultural Economics, Kansas tle a share of those premiums. It is this set of premiums State University. Available at: www.agecon.ksu.edu/livestock/. for high-quality and branded product lines that can Accessed October 29, 2001. expand the number of consumer dollars shared across Tatum, J. D., K. E. Belk, M. H. George, and G. C. Smith. 1999. all participants, and competing for more of those con- Identification of quality management practices to reduce the incidence of retail beef tenderness problems: Development and sumer food dollars is very important (Hudson and Pur- evaluation of a prototype quality system to produce tender beef. cell, 2001). J. Anim. Sci. 77:2112–2118. Using genetic tools to meet market targets without sacrificing maternal performance

M. W. Tess1

Animal and Range Sciences Department, Montana State University, Bozeman 59717

ABSTRACT: Changes in beef production/marketing performance due to selection for increased marbling or systems are creating incentives for beef producers to tenderness. However, selection for increased lean yield improve carcass quality. Current grid marketing carries risks of older age at puberty, increased mature schemes are characterized by premiums for carcasses size, decreased fertility, and possibly increased mater- that excel for marbling and(or) retail product and by nal calving difficulty. Systems research indicates that large discounts for noncompliance factors. Economic reduced fertility would have large negative effects on incentives for producers to select for tenderness may profitability. To avoid detrimental effects on maternal be on the horizon. Effective tools are available for pro- traits, genetic management should consider EPD for multiple traits as part of a systems approach. Managers ducers to genetically tailor cattle to specific markets. will deal with increasing amounts of information and New DNA markers for carcass trait QTL will help cattle detailed monitoring of production costs will become in- producers identify genetically superior animals at creasingly important to success. Structured crossbreed- younger ages. Markers will be used most efficiently ing, exploiting heterosis and breed strengths, should when marker genotype information is incorporated di- be used to increase profitability of production systems. rectly into the computation of EPD, and after potential Management strategies to reduce phenotypic variation epistatic interactions and genotype × environment in- in market groups and to match genetic potential with teractions are characterized for each QTL. Limited re- nutritional management are expected to increase com- search has not identified risks associated with maternal petitiveness in grid marketing schemes.

Key Words: Beef Cattle, Carcasses, Genetics, Maternal Transmission, Systems

2002 American Society of Animal Science. All rights reserved. J. Anim. Sci. 80(E. Suppl. 1):E94–E103

Introduction My objectives are 1) to identify the economic aspects of carcass quality and the traits that will likely receive U. S. beef production and beef marketing are chang- increased attention in future breeding programs, 2) to ing. Beef production systems are becoming more prod- anticipate how important maternal traits might change uct-oriented rather than commodity-oriented. Value- in response to selection for different aspects of carcass based marketing is being stimulated by technologies quality, and 3) to discuss genetic tools and their applica- such as electronic identification and Internet com- tion in meeting these new carcass targets without sacri- merce. Production segments are becoming more coordi- ficing maternal performance. nated, sometimes by formal business alliances, some- times by improved information transfer and measure- Review and Discussion ment of value. Industry changes are providing new incentives for Economic Incentives for Genetic Change genetic changes in carcass traits. Developing technolog- ies are providing new genetic tools for making these Grids. Current marketing systems for fed cattle re- changes. Cow-calf producers are challenged to geneti- flect value differences associated with several traits, cally tailor cattle to meet new and dynamic markets, most of which are measured after slaughter (USDA, yet continue to manage cattle and natural resources in 2001). Primary traits include USDA quality grade and a sustainable manner. USDA yield grade. Quality grade, largely determined by marbling, is intended to be a measure of palatability (USDA, 1997). Yield grade, determined by carcass 1Correspondence: E-mail: [email protected]. weight, longissimus muscle area, fat thickness, and kid- Received July 30, 2001. ney, pelvic and heart fat, predicts retail lean yield. Some Accepted October 4, 2001. systems recognize additional indicators of quality grade

E94 Genetic tools for carcass traits E95 and yield, such as breed identification and muscling. the importance of tenderness to beef demand, it seems Quality grade and yield grade form the basis of most likely that affordable technology will be soon developed grid pricing systems (two-dimensional set of premiums to address this issue, and economic incentives for pro- and discounts) for individual carcasses and motivate ducers to select for tenderness may be on the horizon. price differentials paid for groups of cattle sold before slaughter. Magnitudes of these premiums and dis- Genetic Relationships Between counts vary seasonally in response to supply and de- Carcass and Maternal Traits mand factors (LMIC, 2001). Grid prices may also differ among alliances and(or) packers. The components of quality grade and yield grade are Grid marketing schemes normally take into account moderately to highly heritable (Marshall, 1994; several compliance factors, measures that largely iden- Shanks, 1999; Shanks et al., 2001); hence, selection is tify carcasses that cannot be marketed through main- expected to be effective in changing these traits. Histor- stream channels. Compliance factors include extremes ically, carcass traits have not received much attention in carcass weight (light or heavy), undesirable levels of in the breeding plans of U.S. beef producers. As beef maturity, carcasses from bulls or dairy breeds, and dark producers now focus on carcass traits, answers to im- cutters (USDA, 2001). Essentially, there are no premi- portant questions are needed: 1) Will selection for im- ums for compliance factors, only discounts. Generally, proved carcass characteristics lead to undesirable cor- discounts for the most undesirable quality and yield related responses in important maternal traits and 2) grades (standard quality grade and yield grades ≥ 4), Will breed substitutions to improve carcass quality neg- and for noncompliant carcasses are much larger than atively affect maternal traits? Regrettably, few experi- premiums paid for desirable quality grades and yield ments focusing on selection for carcass quality have grades. Hence, these carcasses are referred to as “out- been conducted. Therefore, few correlated selection re- cattle” (i.e., cattle that do not fit the grid). Discounts sponses have been documented, and we must rely on for out-cattle are so severe that it can easily take two less direct types of information. I will focus on three or more premium carcasses to balance one noncompli- traits: marbling, retail product (or yield grade and its ant carcass. components), and tenderness. Grid marketing schemes provide incentives for beef Marbling. At the University of Nebraska, Angus sires producers to genetically improve quality grade and with high and low EPD for marbling were randomly yield grade. In addition, there are strong incentives for mated to MARC II composite cows. Progeny were evalu- producers to avoid marketing out-cattle. Because out- ated for growth and carcass traits (Vieselmeyer et al., cattle are associated with extreme values for quality 1996) and palatability traits (Gwartney et al., 1996). grade, yield grade, and carcass weight, the incentive to Because heifers were slaughtered, this study provided avoid out-cattle is largely an incentive to market cattle no information on correlated responses in maternal with less phenotypic variability. traits. Niche Markets. Utilizing coordinated networks or in- Frazier et al. (1999) used field data to evaluate associ- tegrated systems some beef producers are targeting ations of Angus sire marbling score EPD with reproduc- specialized or niche markets. Generally, these systems tive performance of females. Marbling score EPD was market beef products at the retail level with brand significantly associated with age at first calving; how- name identification. Examples include Laura’s Lean ever, predictors were not consistent across states, even Beef (Laura’s Lean Beef, 2001), Montana Range (Leach- switching signs. Classifying sires into marbling score man, 2001), and Coleman Natural Products (Coleman, EPD categories produced inconsistent relationships 2001). Product definition varies among systems but with age at first calving. The authors concluded that usually focuses on a specific aspect of eating quality marbling score EPD was not a good predictor of age (e.g., tenderness or leanness) or food safety (e.g., free of at first calving or calving interval and suggested that antibiotics). Depending on the market niche, producers selection for marbling should not affect these mater- participating in these systems experience incentives for nal traits. genetic change that may be different from those for Splan et al. (1998) estimated genetic correlations be- producers marketing through mainstream channels. tween male carcass traits and female reproductive Tenderness is considered a very important heritable traits. Genetic correlations between marbling score and palatability trait (Dikeman et al., 2001). Tenderness age at puberty, calving rate, and calving difficulty were must be measured on cooked meat, and available tech- close to zero (Table 1), suggesting that little change nology to measure tenderness in commercial packing would occur in these maternal traits in response to plants is considered too expensive (NCBA, 1998a). selection for increased marbling. Therefore, there is currently no industry mechanism to As the most popular U.S. beef breed (Taylor and provide information feedback to producers on a large Field, 1999), Angus has served as a benchmark for scale. Except for production systems marketing many breed evaluation studies. Angus cattle have branded retail products (e.g., Montana Range; Leach- higher marbling scores than most other beef breeds man, 2001), there is currently little economic incentive used in the United States (Cundiff et al., 1986). Breed for producers to select for tenderness. However, due to comparisons have not identified problems with age at E96 Tess Table 1. Genetic correlations between maternal performance traits and carcass traitsa

Age at Conceptions/ Calving Calving Mature Trait puberty service rate difficulty wt

Retail product wt 0.30 0.28 −0.02 0.25 Retail product % −0.01 −0.13 0.18 Carcass wt 0.17 0.61 0.05 −0.31 0.21 0.06 −0.17 Fat thickness −0.29 0.21 0.19 −0.36 −0.09 −0.01 −0.14 Longissimus muscle area 0.04 0.15 −0.04 Marbling score −0.04 −0.05 −0.09 Warner-Bratzler shear force 0.01 0.11 0.19 Taste panel tenderness −0.32 0.07 −0.42 aCorrelations involving retail product weight, conceptions/service, and mature weight are from MacNeil et al. (1984). Correlations involving retail product %, longissimus muscle area, marbling score, Warner- Bratzler shear force, and taste panel tenderness are from Splan et al. (1998). When two values are listed, the upper value is from MacNeil et al. (1984) and the lower from Splan et al. (1998). All carcass traits are adjusted to constant age. puberty, calving difficulty, pregnancy rate, calf crop heifers sired by these breeds tend to be about 1 mo weaned, or maintenance requirements for beef breeds older at puberty and have lower heifer pregnancy rates that rank high in marbling (Ferrell and Jenkins, 1985; than breeds with less rapid growth and lean yield (Cun- Cundiff et al., 1986; Martin et al., 1992). diff et al., 1986; Martin et al., 1992); however, most Retail Product. MacNeil et al. (1984) reported genetic heifers from these breeds should still reach puberty correlations between retail product weight and age at before their first breeding season under good nutri- puberty, conceptions/service, maternal calving diffi- tional management. Means for calving difficulty and culty, and mature weight (Table 1). Their results sug- calf crop weaned were similar for breeds with moderate gest that selection for increased retail product weight vs more rapid growth and lean yield (Cundiff et al., would lead to older ages at puberty, improved fertility, 1986). Bennett and Williams (1994) hypothesized that and increased mature size with no change in maternal cows from larger, leaner breeds would require better calving difficulty. nutritional management than more moderate types. Retail product percentage and retail product weight Nugent et al. (1993) reported that these biological types are not the same traits but are genetically correlated of cows had shorter postpartum intervals in response (0.46, Koch et al., 1982). Splan et al. (1998) computed to increased feed. Compared to progeny of sires from genetic correlations between female traits and retail moderate breeds, progeny of sires from breeds charac- product percentage. Correlations reported by Splan et terized by rapid growth and high lean yield and dams of al. (1998) suggest that selection for retail product per- moderate types are expected to experience more calving centage will lead to no change in age at puberty, a small difficulty and calf mortality, leading to reduced rebreed- decline in calving rate, and a slight increase in maternal ing rates in dams (Cundiff et al., 1986). Feed requirements for maintenance account for over calving difficulty. 70% of annual requirements for beef cattle. Many stud- As components of retail product (or yield grade), fat ies have attempted to estimate maintenance require- thickness and longissimus muscle area are also of inter- ments for different breeds and biological types (Ferrell est. Because breed associations report EPD for these and Jenkins, 1985, 1998a,b). In some studies larger, traits, some producers may select for decreased fat leaner breeds had higher maintenance requirements thickness or increased longissimus muscle area. Ge- (Mcal ME/kg0.75) than more moderate types, but not netic correlations reported by MacNeil et al. (1984) and in others. Ferrell and Jenkins (1985) concluded that Splan et al. (1998) indicate that selection for decreased maintenance requirements are positively associated fat thickness would negatively affect age at puberty, with genetic potential for measures of production such fertility, calving rate, and maternal calving difficulty. as growth rate and milk yield. However, the relation- Selection for increased longissimus muscle area would ship between maintenance and milk potential seems to have no effect on puberty or calving difficulty but would be stronger than that between maintenance and growth improve calving rate slightly (Table 1). Undesirable as- potential, and U.S. feeding standards (NRC, 1996) do sociations between maternal traits and retail product not adjust maintenance requirements upward for appear to be mediated through fat thickness. larger, leaner breeds. The lack of independence between Breed comparisons involving breeds that excel for estimates of maintenance and the efficiency of energy retail product are confounded by the fact that some of use for growth, and the change in efficiency of energy these breeds also have high milk production. Re- use for growth with changing energy intake makes esti- stricting comparisons to breeds that have moderate mation and interpretation of these parameters difficult milk but increased growth and lean yield suggests that (Ferrell and Jenkins, 1998a,b). Genetic tools for carcass traits E97 Although easily classified as high retail product ing difficulty. Breed comparisons suggest that com- breeds (Wheeler et al., 1996), Belgian Blue and Pied- pared to moderate breeds, breeds that excel for growth montese are distinct from other breeds due to their and lean yield are older at puberty, except for the dou- frequent expression of muscle hypertrophy (double- ble-muscled Belgian Blue and Piedmontese, which are muscling), a condition caused by the effects of alleles younger. Breed comparison studies have not identified at a single locus (inactive myostatin, Casas et al., 1998, important differences in maternal calving difficulty or 1999). Compared to homozygous normal animals, indi- calf crop weaned due to increased growth or leanness. viduals carrying a single copy weigh more at birth but It is important to know just how detrimental changes experience similar calving difficulty, have larger longis- in these maternal traits might be to ranch profitability. simus muscle area, less fat thickness, and greater retail Using systems analysis techniques (Tess and Kolstad, product yield, yet do not express double-muscling (Ca- 2000a,b), Tess (1999) evaluated the effects of changes sas et al., 1998, 1999). Heifers sired by Piedmontese in several maternal traits on cow-calf enterprise profit- bulls average 15 to 20 d younger at puberty than Angus ability in a range environment. This study demon- or Hereford crossbreds (Thallman et al., 1999; Lammog- strated that system performance was quite sensitive to lia et al., 2000). Pregnancy rates, calving and weaning performance levels for postpartum interval to estrus percentages, and postpartum intervals to estrus for Pie- and mature cow conception rate (i.e., fertility). Postpar- dmontese- and Belgian Blue-sired heifers appear to be tum intervals greater than about 70 d led to precipitous similar to those for Hereford and Angus crossbreds drops in gross margin. Gross margin declined in a (Freetly and Cundiff, 1998; Thallman et al., 1999). nearly linear fashion with mature cow conception rate. Tenderness. Splan et al. (1998) reported genetic corre- Together, postpartum interval and conception rate lations between two measures of tenderness (Warner- largely determine calving rate. Profitability was rela- Bratzler shear force and taste panel tenderness) and tively insensitive to calving difficulty rate. Increased maternal traits (Table 1). Inconsistencies among corre- age at puberty and decreased heifer conception rate led lations for shear force and taste panel tenderness make to higher replacement rates, yet their effects on gross conclusions difficult; however, their results suggest that margin depended on marketing strategies for cull heif- selection for increased tenderness should lead to less ers. If nonpregnant heifers were sold following the graz- maternal calving difficulty and possibly younger age ing season, gross margin was relatively unaffected, es- at puberty. sentially shifting income from calves to yearlings. If Differences in Warner-Bratzler shear force and(or) nonpregnant heifers were given an additional year to taste panel tenderness have been demonstrated among breed, gross margin declined after age at puberty ex- several breeds (Marshall, 1994; Wheeler et al., 1996). ceeded approximately 420 d. Similarly, if nonpregnant Generally, tenderness measures for F1 crosses have heifers were retained profitability declined in a nearly been within acceptable ranges. Bos indicus tend to be linear manner as heifer conception rate declined. less tender than Bos taurus breeds. Two breeds that Most female reproductive traits have a minimum appear to excel for tenderness are Pinzgauer and Pied- level of performance below which herd size cannot be montese (Marshall, 1994; Wheeler et al., 1996). The maintained without purchasing replacements. The re- tenderness advantage for Piedmontese does not appear sults of Tess (1999) suggest that the relationship be- to be associated with the myostatin locus (Casas et al., tween profitability and maternal performance is mir- 1998). Reproductive comparisons involving Piedmont- rored by the effects of maternal traits on calf weight ese were reviewed above. Pinzgauer heifers tend to be weaned per cow exposed, or by weight sold per cow about 2 wk younger at puberty and have higher preg- exposed. Any genetic or management change that de- nancy rates than Hereford and Angus crosses, whereas creases these measures is likely to be detrimental to calving difficulty and calf crop percentages are similar ranch profit. Estimates of economic weights for fertility for these breeds (Cundiff et al., 1986; Martin et al., have consistently been positive (Kolstad, 1993; MacNeil 1992). et al., 1994; Koots and Gibson, 1998a,b). Potentially Sensitive Traits in Range Cow-Calf Pro- Several researchers have studied the effects of ma- duction Systems. Based on this brief review, there is ture size on beef enterprise profitability. Using systems little evidence to suggest that selection for increased analysis techniques, Kolstad (1993) found that in- marbling or increased tenderness or that substituting creases in mature weight for maternal strains, indepen- with breeds that excel in these traits will lead to de- dent of other growth traits, had negative effects on clines in maternal performance. It should be empha- profit. Similarly, MacNeil et al. (1994) reported that sized, however, that direct measures of correlated re- the relative economic value of mature weight was nega- sponses to selection have not been reported. On the tive for maternal strains and general-purpose strains other hand, selection for increased retail product is ex- in all production/marketing scenarios studied. Koots pected to lead to older ages at puberty for heifers, heav- and Gibson (1998a) reported that mature weight had ier mature weight, and possibly increased maternal a positive economic value in general-purpose lines; how- calving difficulty. Selection for decreased carcass fat ever, in their model mature weight accounted for other thickness is expected to lead to older ages at puberty, growth traits. Further, the economic value of mature decreased fertility and calving rate, and increased calv- weight was sensitive to the marketing assumptions and E98 Tess base trait means (Koots and Gibson, 1998b). Heavier carcass quality. Real-time ultrasound has proved to be mature weight represents added overhead in terms of an effective technology to meet this goal (Wilson, 1992). feed for maintenance (per cow, Cundiff et al., 1986). An ultrasound measurement made on potential For a given feed resource (e.g., rangeland), increasing breeding stock raised under ranch conditions (e.g., bulls feed requirements per cow will require reductions in or heifers at 1 yr) is not the same trait as the direct herd size (Tess and Kolstad, 2000b). Cows that excel measurement made on carcasses from animals grown in rapid early growth but mature at lighter weights in a feedlot, but a genetically correlated trait that ex- should be desirable in most systems. plains (or accounts for) some but not all the variation in the carcass trait. Estimates of genetic correlations The Genetic Toolbox between carcass measures and ultrasound measures are needed before EPD for carcass traits can be com- There are a variety of genetic tools currently avail- puted using ultrasound records (e.g., Moser et al., 1998). able to beef producers, and new tools are sure to be Hence, breed associations have taken one of four ap- developed. Just like a mechanic uses different tools for proaches to computing EPD for carcass traits: 1) using different jobs, beef producers should use genetic tools carcass data to compute carcass EPD, 2) using ultra- based on their effectiveness, ease of application, and sound data to compute ultrasound EPD, 3) computing cost. separate EPD for carcass and ultrasound, or 4) using Breed Resources. Breed differences in carcass traits carcass and ultrasound records to compute a single car- have been reported in several studies (Marshall, 1994; cass EPD (i.e., via the genetic correlations). Wheeler et al., 1996). For some producers, exploiting Due to the fact that ultrasound measurements are breed differences will be the fastest way to make genetic made on potential parents, one might predict that fu- adjustments in carcass quality. If females will be used ture carcass trait EPD will be essentially based on ul- as replacements, choices among breeds should be based trasound measurements. However, developing technol- on maternal as well as growth and carcass traits (Cun- ogy related to source and process verification (i.e., elec- diff et al., 1986). For some niche markets, production tronic animal identification, animal tracking networks, of animals that carry a single copy of the inactive myos- and Internet data transfer) might facilitate a different tatin allele may form an important component of the course. If this electronic technology is widely adopted, breeding strategy (e.g., Montana Range, Leachman, and if the expense is low, direct carcass measures might 2001). eventually be the dominant source of genetic informa- Expected Progeny Differences. To practice effective tion on carcass traits. selection within breeds, estimates of breeding value Genetic Markers. Molecular genetic markers (i.e., or EPD are needed. Compared to most growth traits, DNA markers) have been reported for QTL affecting traditional performance and progeny testing for carcass several carcass components, including rib bone and merit is more difficult. Carcass traits cannot be mea- dressing percentage (Stone et al., 1999) and marbling sured directly on potential parents. Carcass measure- score (Casas et al., 2001; GeneStar, 2001). Research ments on cattle (i.e., progeny or other relatives of poten- also suggests the possibility of QTL for carcass weight, tial parents) marketed and harvested through main- fat depth, longissimus muscle area, and retail product stream commercial channels are expensive to collect. yield (Stone et al., 1999; Casas et al., 2000, 2001). The Ownership of calves can change several times prior to muscle hypertrophy locus (inactive myostatin allele) slaughter and cattle are typically moved to different found in the Belgian Blue and Piedmontese breeds has locations during their lifetime; hence, maintaining ani- been mapped (e.g., Fahrenkrug et al., 1999) and shown mal identification and information feedback to the cow- to affect longissimus muscle area, retail product yield, calf segment (i.e., where breeding decisions are made) marbling, yield grade, fat thickness, and kidney, pelvic, are difficult tasks. Nevertheless, carcass traits are mod- and heart fat. Undoubtedly, additional markers will erate to highly heritable (Marshall, 1994; Shanks, 1999; be discovered and validated for these and other traits Shanks et al., 2001) and several breed associations cur- (NCBA, 1998b). Markers that prove to be closely linked rently compute and publish EPD for carcass traits to important QTL (i.e., QTL having a large economic based on direct measures of carcass quality. effect) or markers that actually map to the QTL locus Current technology permits computation of adjust- will likely be rapidly commercialized (e.g., GeneStar, ment factors for comparing EPD across breeds (across- 2001). breed EPD; e.g., Barkhouse et al., 1998) or by computing Due to the part/whole relationship between QTL and EPD for animals from different breeds using one the target trait, DNA markers explain a portion, but multibreed dataset (multiple-breed EPD; Pollak and not all, of the genetic variation in the target trait. In Quass, 1998). Across-breed EPD adjustments are cur- other words (and similar to ultrasound), DNA markers rently not available for carcass traits. account for part, but not all, of an animal’s carcass EPD. Ultrasound. Problems associated with direct mea- The utility of DNA markers is due to the fact that the sures of carcass quality have motivated searches for genotype for the marker, and hence a portion of the traits that could be measured on live animals, especially EPD, can be determined early in life on potential par- potential parents, that could provide information on ents. The most valuable markers will be those that Genetic tools for carcass traits E99 explain a significant portion of one or more EPD and can obvious problem with heterozygous bulls is that only be economically assayed. Physiological measurements half their progeny will carry the desirable allele. More such as blood hormone concentrations could potentially important, the danger is that producers will value these have similar utility in providing early indicators of ge- bulls highly when it is possible, if not likely, that other netic merit. For example, Davis and Simmen (2000) bulls within the same breed are genetically superior found that bulls with lower serum IGF-I concentrations because they are homozygous for the desirable allele had higher marbling scores, quality grades, fat thick- and(or) because their net genetic makeup (EPD) is supe- ness, and yield grades. rior. Because QTL explain only part of the genetic varia- In the future, it is likely that marker genotype infor- tion for the target trait, it is possible for a bull to carry mation will be incorporated into national genetic evalu- a desirable QTL allele yet not rank near the top of the ation programs. In other words, information from DNA breed for the trait. markers will supplement performance information in Research relating to DNA markers has made great computing EPD. As described for ultrasound, genetic progress in a short time. Much effort has been expended correlations between marker genotype and the trait to identify potentially useful markers, yet little is of interest will be needed in order to incorporate this known regarding interactions involving QTL. In other information into the computation of EPD. If ultrasound words, for many QTL it is not known how or whether and carcass measures are both used to compute EPD, the QTL will interact with other QTL or the background genetic correlations between the QTL and both mea- genotype (epistasis), or whether the QTL performs the sures will be needed. same in all production environments (genotype × envi- Risks and Questions. Marker-assisted selection will ronment interaction). For example, Casas et al. (2000, no doubt help beef producers be more competitive in 2001) detected evidence of interactions between the my- the future. However, like most technology, it is accom- ostatin gene and QTL affecting Warner-Bratzler shear panied by risk. force and fat thickness. Future research will likely ad- In the future, one can envision several DNA markers dress these questions. However, until these questions and possibly some physiological markers documented are answered for each QTL, there will be uncertainty for each carcass trait, with private companies conduct- in predicting their effects on performance. ing assays for the markers. For a given trait, each Application of DNA technology in the beef industry marker will explain a portion, but not all of the EPD faces obstacles and questions beyond the scope of this for an animal. To my knowledge no current genetic paper. Statistical procedures to incorporate QTL infor- evaluation system explicitly includes marker genotype mation into the computation of EPD are currently being information in the analyses, but it is likely that such developed. The computational complexity of these pro- technology will be soon developed. Nevertheless, cur- cedures may restrict the number of QTL that can be rent carcass EPD (computed from direct carcass mea- incorporated simultaneously. It may be necessary to surements and[or] ultrasound) already account for validate that QTL (and their markers) are segregating some of the effects of any QTL that exist for the trait. in each breed. Private ownership of DNA technology In other words, QTL affect performance whether we and competition for business will limit information know they are there or not, and performance differences sharing. Cost/benefits of the technology to the beef in- are the basis of EPD. dustry may be difficult to calculate, and marketing of Consider a beef producer who is considering using perceived benefits may drive application. semen from a bull based on genetic merit for some carcass trait, and assume that marbling is the trait of A Systems Approach to Genetics and Management interest. It is easy to envision a situation in which the bull under consideration has a marbling EPD, plus one Similar to other industries, the beef industry is be- or more known marker genotypes for marbling QTL. coming more complex. Managers must deal with in- Regardless of whether the EPD was calculated using creasing amounts of information to be competitive. Co- the marker genotype information, the genetic informa- ordinated and integrated production systems may need tion provided by the EPD and that provided by the to supply beef products on a weekly basis to meet mar- marker genotypes overlap (i.e., they are not indepen- ket demand. Calving in different seasons will make dent). In such cases the marker genotype information matching forage resources and nutrient demand more is likely to be overvalued by the beef producer. difficult (Reisenauer et al., 2001). Detailed monitoring As mentioned above, some DNA markers detect al- of production costs will become increasingly important. leles at loci that are closely linked to QTL, whereas Marketing strategies and genetic tools are just part of others detect actual QTL alleles. Using current technol- beef cattle management systems. Management from a ogy for linked markers, usually only heterozygous ani- total systems approach will be increasingly important mals can be identified (i.e., animals that carry one, to success. but not two, copies of the desirable allele). Hence, it is As reviewed above, evidence suggests that producers possible for an animal to carry both desirable QTL al- breeding for increased lean growth risk loss of perfor- leles but they could be undetectable. In the industry, mance for some maternal traits, including age at pu- heterozygous bulls are generating much attention. An berty, fertility, calving ease, and mature size. Yet, the E100 Tess Table 2. Reduction in phenotypic standard deviation (%) due to the production of various types of related calvesa

Additive genetic variance: 20.00 40.00 40.00 Mating scheme Dominance variance: 20.00 20.00 40.00

Multiple unrelated sires 0.00 0.00 0.00 Sires = ¹⁄₂ sibs 0.63 1.26 1.26 Sires = ³⁄₄ sibs 0.78 1.57 1.57 Sires = full sibs 1.26 2.53 2.56 Sires = inbred full sibs (F = 0.25) 1.51 3.05 3.05 Single sire 2.53 5.13 5.13 Single inbred sire (F = 0.25) 3.18 6.46 6.46 Full-sib embryo transfer calves 7.80 13.40 16.33 Calves = clones 22.54 36.75 55.28 aPhenotypic variance = 100.00. No maternal effects. research available regarding the relationships between counteracting the potential increase in age at puberty maternal traits and marbling or tenderness is not ex- expected from using breeds that excel for lean growth. tensive. Hence, uncertainty exists regarding how ge- Crossbred heifers reach puberty 1 to 3 wk earlier than netic emphasis on carcass performance will affect ma- purebreds (Gregory et al., 1991; Martin et al., 1992). ternal performance. How do genetic tools fit into the Heterosis effects are desirable for most carcass traits management system to meet future markets? except fat thickness and retail product percentage Multiple Trait Genetic Management. First, it is im- (Cundiff et al., 1986; Marshall, 1994). portant to emphasize that effective genetic manage- Practical, structured crossbreeding systems and well- ment will be multiple trait management. In coordinated designed composite breeds permit breed differences and or integrated production systems profitability is deter- heterosis to be combined in a complementary manner mined by production efficiency in each production seg- and manage trade-offs due to genetic antagonisms ment (cow-calf, background or stocker, and feedlot), (Gregory and Cundiff, 1980; Kress and MacNeil, 1999). plus end-product value. Carcass traits should never be Using systems analysis techniques, Lamb and Tess the sole focus of genetic decisions. Markers (and possi- (1989) reported that two- and three-breed rotational bly other indicator traits) will provide valuable early crossbreeding systems produced about 11% more in- indicators of genetic merit. The industry will use mark- come than purebred systems in small herds. Davis et ers most efficiently when information from marker ge- al. (1994) determined that maternal heterosis improved -notypes is incorporated directly into the computation profit $70ؒcow−1ؒyr−1 for range-based cow-calf enter of EPD. prises. Lamb et al. (1992a) found that rotational cross- Fortunately, EPD are available for many important breeding systems improved economic efficiency by an beef cattle traits. Widespread whole-herd reporting average of 16 to 17% over purebred systems. Tess and could facilitate genetic evaluation of additional mater- Kolstad (2000b) reported that a rotation of three com- nal traits (Tess, 1999). Monitoring an array of traits posite breeds improved annual net income per cow by in making selection decisions can prevent undesirable 12% and ranch gross margin by 6% when compared to correlated responses. Most breed associations compute the average of the same composite breeds managed as EPD for calving ease or birth weight, which can be used purebreds. Adding a composite terminal sire breed im- to avoid increases in calving difficulty. Some breeds proved net income per cow and gross margin by 18 and compute EPD for mature weight, and most compute 19%, respectively. Lamb et al. (1992b,c; 1993) found EPD for weaning weight, yearling weight, and carcass that rotational crossbreeding systems were economi- weight. These growth traits are highly correlated (Bul- cally more efficient than purebred systems in the feedlot lock et al., 1993; Northcutt and Wilson, 1993; Kaps et segment and in integrated systems, regardless of the al., 1999); hence, monitoring EPD for these traits will marketing end point. help producers avoid unwanted increases in mature Use of terminal sires probably gives producers the size. Grid discounts for extremely heavy carcasses pro- greatest flexibility in genetically tailoring cattle to spe- vide further incentive to constrain mature size. Devel- cific market end points (Bennett and Williams, 1994). opment of EPD for scrotal size and stayability will be Terminal sire systems will be the most practical way useful in maintaining or improving age at puberty and of exploiting the advantages of animals that are hetero- fertility (Tess, 1999). zygous for single copies of QTL such as the inactive Heterosis. Second, the economic value of heterosis myostatin allele. Many producers, especially those should be exploited, especially in reproductive females. managing small herds, find terminal sire systems diffi- Favorable heterosis effects for reproductive traits are cult to implement due to problems associated with bull/ well documented (Long, 1980; Cundiff et al., 1986; Greg- cow ratios, generation of replacement heifers, and vari- ory et al., 1992). Crossbreeding is an effective way of ation in performance between progeny of terminal sires Genetic tools for carcass traits E101 Table 3. Effects of breed variation and sorting on the range in sale weights (kg)a

Scenario No sort Sort off 5% Sort off 10%

Uniform crossbreeding system 121 94 83 Inconsistent crossbreeding, maximum performance difference among breeds 137 107 93 aSteer calves sold at weaning. 60-d calving season. Calves removed are those with extreme heavy or light weights. and maternal-line sires. It seems likely that economic els (i.e., the same biological type) will produce variation incentives created by some markets may motivate more similar to purebred herds. Similarly, well-planned ter- producers to purchase replacement heifers. minal-sire crossbreeding systems and straight-breed- Reducing Variation. The large discounts associated ing systems based on composite breeds will also exploit with nonconforming carcasses in most grid-marketing heterosis without increasing calf variation. Across- systems can be avoided by marketing phenotypically breed EPD and multiple-breed EPD can be used to uniform groups of cattle. Hence, managers have incen- choose bulls with similar EPD from breeds that might tive to limit genetic variation as well as variation have quite different performance means, although caused by management and environmental factors. these tools are not yet available for carcass traits. Cross- Related animals are genetically more similar than breeding systems that indiscriminately use sires from unrelated animals. Table 2 shows the expected reduc- different biological types and switch sire breeds fre- tion in phenotypic variation (i.e., the range in pheno- quently will produce more variable calves. typic values or the standard deviation) by producing Variation in calf age contributes greatly to variation different types of related calves. Using related sires in phenotype. Table 4 summarizes expected differences has relatively little effect on phenotypic variation, even in variation associated with differences in the length of when heritability is high. The production of full-sib calving season. The range in weaning weights expected calves (i.e., through embryo transfer) or clones could among steers born over a 45-d period is approximately reduce phenotypic variation for traits exhibiting much 118 kg. This difference increases by another 36 kg for genetic variation (additive and dominance variance) calves born over a 120-d calving period. The obvious way (Lamberson, 1994), but it is doubtful that either will to restrict the calving period is to restrict the breeding be practical in the near future in beef cattle. Inbreeding season. However, the real issue is the length of the helps reduce variation, but the loss of performance asso- calving period rather than the number of days bulls ciated with inbreeding is too great for this to be a useful run with the cows. tool (Brinks and Knapp, 1975). Within-family variation Sorting off extreme animals prior to sale or shipment does not appear to be affected by differences in EPD can be an effective tool to reduce phenotypic variation. between parents (Bullock et al., 2000). Unfortunately, Tables 3 and 4 illustrate the effects of sorting on wean- there are few practical tools to significantly reduce ge- ing weight variation. Just as ultrasound is used in feed- netic variation within breeds. lots to sort animals into management/marketing Genetic variation associated with breeds and mating groups, it seems possible that DNA markers might also systems also can add to variation among calves. Table 3 be used this way. For example, knowing whether cattle illustrates the expected weight variation among calves are expressing a major allele affecting marbling might from herds using what could be called “uniform” cross- help beef producers better match cattle with specific breeding vs herds using “inconsistent” crossbreeding. pricing grids, or knowing whether or not cattle are ex- Predictions in Table 3 were simulated based on perfor- pressing a major allele for tenderness might reduce the mance differences among British and Continental beef risk of guaranteeing branded products as tender. The breeds reported by the U.S. Meat Animal Research Cen- utility of using DNA markers as management/market- ter (Cundiff et al., 1986). Herds using rotational cross- ing tools will depend on the cost/benefit of the infor- breeding based on breeds with similar performance lev- mation.

Table 4. Effects of calving season length and sorting on the range in sale weights (kg)a

Calving season No sort Sort off 5% Sort off 10%

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Smith. 1999. potential epistatic interactions and genotype × environ- Technical Note: Direct genotyping of the double-muscling locus (mh) in Piedmontese and Belgian Blue cattle by fluorescent PCR. ment interactions are characterized for each QTL. To J. Anim. Sci. 77:2028–2030. avoid detrimental effects on maternal traits, genetic Ferrell, C. L., and T. G. Jenkins. 1985. Cow type and the nutritional management needs to consider multiple traits as part environment: Nutritional aspects. J. Anim. Sci. 61:725–741. of a systems approach. Structured crossbreeding, ex- Ferrell, C. L., and T. G. Jenkins. 1998a. Body composition and energy ploiting heterosis and breed strengths, should be used utilization by steers of diverse genotypes fed a high-concentrate diet during the finishing period: I. Angus, Belgian Blue, Here- to increase profitability of production systems. Manage- ford, and Piedmontese Sires. J. Anim. 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