Effects of Chiropractic on Equine Gait Kinematics, Heart Rate, And

EFFECTS OF CHIROPRACTIC ON EQUINE GAIT KINEMATICS, HEART RATE, AND

SERUM CORTISOL

JESSICA GLADNEY

(Under the Direction of Kari Turner)

ABSTRACT

Study used 18 horses to determine how chiropractic affects riding sound horses over 8 weeks. Horses divided into Chiropractic (n=6), Riding (n=6), and Sedentary (n=6) group. Riding and Chiropractic group ridden 4 days per week. Sedentary horses unridden. Kinematics collected on Day 0, Day 2, Day 14, Day 28, Day 30, Day 42, and Day 56. Chiropractic group received chiropractic while Riding and Sedentary received grooming treatment on Day 1 and Day 29.

Heart rate monitors placed on all horses during treatment. Serum cortisol collected on all PRE,

MID, and POST treatment. At the trot, swing as a percentage of stride decreased in hind limbs on

Day 56 compared to Day 28 (p=0.05), Day 14 (p=0.03), and Day 2 (p=0.02) in Chiropractic group. Cortisol is higher on Day 29 PRE vs. Day 1 PRE (P=0.03) in the Chiropractic group.

Results show chiropractic minimally effects kinematics and increases PRE cortisol levels.

INDEX WORDS: Chiropractic, Gait Kinematics, Heart rate, Serum Cortisol

EFFECTS OF CHIROPRACTIC ON EQUINE GAIT KINEMATICS, HEART RATE, AND

SERUM CORTISOL IN RIDING HORSES

JESSICA GLADNEY

B.S., UGA, 2015

A Thesis Submitted to the Graduate Faculty of The University of Georgia in Partial Fulfillment

of the Requirements for the Degree

MASTER OF SCIENCE

ATHENS, GEORGIA

Jessica Gladney

SERUM CORTISOL

JESSICA GLADNEY

Major Professor: Kari Turner Committee: Kylee Jo Duberstein Franklin West

Electronic Version Approved:

Suzanne Barbour Dean of the Graduate School The University of Georgia December 2017 ACKNOWLEDGEMENTS

Committee: Dr. Kari Turner, Dr. Kylee Jo Duberstein, Dr. Franklin West

UGA Livestock Arena Manager: Alexander Abrams

Chiropractor: Dr. Dana Peroni DVM, Covered Bridge Equine

Lameness Evaluation: Dr. Paige Williams DVM, Covered Bridge Equine

Cortisol: Dr. Clay Lents, (who was found via Dr. Michael Azain)

Undergraduate Researchers and Riders: Mary Cate Marchert, Marisa Bartholomew, Erin Jarboe,

Madisen Gloeggler, Morgan Garrick, Jessica Simons, and Kaitlyn Gilroy

This project would not be possible without every single one of the above names. Thank you, Thank you with all my heart. Thank you to every single one of my family and friends who encouraged me, supported me, and helped me along the way.

iv TABLE OF CONTENTS

Page

ACKNOWLEDGEMENTS ...... iv

LIST OF TABLES ...... vii

LIST OF FIGURES ...... viii

CHAPTER

1 INTRODUCTION ...... 1

Literature Cited ...... 3

2 REVIEW OF LITERATURE ...... 4

Alternative Therapies ...... 4

Pain and Lameness ...... 7

Treating Pain and Lameness ...... 9

Stress in Horses ...... 9

Biomechanics of the Horse ...... 15

Gait and Back Kinematics ...... 16

Literature Cited ...... 21

3 EFFECTS OF CHIROPRACTIC ON EQUINE GAIT KINEMATICS, HEART

RATE, AND SERUM CORTISOL ...... 29

Abstract ...... 30

Introduction ...... 32

Materials and Methods ...... 34

v Results ...... 41

Discussion ...... 48

Literature Cited ...... 51

4 CONCLUSIONS...... 56

Literature Cited ...... 58

REFERENCES ...... 59

APPENDICES

A Chiropractic Adjustment for Horses ...... 70

B Results Tables ...... 71

vi LIST OF TABLES

Page

Table 1: AAEP Lameness Scale ...... 8

Table 2: Treatment*Day lsmeans of Cortisol ...... 46

Table 3: Treatment*Day lsmeans of Heart Rate ...... 47

Table 4: Treatment*Day lsmeans of the Front Limb Measurements at the Walk ...... 71

Table 5: Treatment*Day lsmeans of the Back Limb Measurements at the Walk ...... 73

Table 6: Treatment*Day lsmeans of the Front Limb Measurements at the Trot ...... 74

Table 7: Treatment*Day lsmeans of the Back Limb Measurements at the Trot ...... 75

Table 8: Treatment*Day lsmeans of Suspension Time at the Trot ...... 76

Table 9: Treatment*Day lsmeans of Stride Length at the Walk and Trot ...... 77

Table 10: Treatment lsmeans of Cortisol ...... 77

Table 11: Treatment lsmeans of AVG Heart Rate ...... 78

Table 12: Treatment lsmeans of MAX Heart Rate ...... 78

Table 13: Treatment lsmeans of MIN Heart Rate ...... 78

Table 14: Treatment lsmeans of Limb Measurements at the Walk ...... 78

Table 15: Treatment lsmeans of Limb Measurements at the Trot ...... 80

Table 16: Serum Cortisol by Horse and Group on Day 1 ...... 86

Table 17: Serum Cortisol by Horse and Group on Day 29 ...... 87

vii LIST OF FIGURES

Page

Figure 1: Walk and Trot of the Horse ...... 16

Figure 2: Phases of the Trot ...... 19

Figure 3.1: Treatment*Day Lsmeans of the Front Limbs at the Walk ...... 41

Figure 3.2: Treatment*Day Lsmeans of the Hind Limbs at the Walk ...... 42

Figure 4.1: Treatment*Day Lsmeans of the Front Limbs at the Trot ...... 43

Figure 4.2: Treatment*Day Lsmeans of the Hind Limbs at the Trot ...... 44

Figure 5: Stride Length of the Walk and Trot...... 45

Figure 6.1: Treatment*Day Lsmeans of the Front Limbs at the Trot ...... 82

Figure 6.2: Treatment*Day Lsmeans of the Front Limbs at the Trot ...... 82

Figure 7.1: Treatment*Day Lsmeans of the Hind Limbs at the Trot ...... 83

Figure 7.2: Treatment*Day Lsmeans of the Hind Limbs at the Trot ...... 83

Figure 8.1: Treatment*Day Lsmeans of the Front Limbs at the Walk ...... 84

Figure 8.2: Treatment*Day Lsmeans of the Front Limbs at the Walk ...... 84

Figure 9.1: Treatment*Day Lsmeans of the Hind Limbs at the Trot ...... 85

Figure 9.2: Treatment*Day Lsmeans of the Hind Limbs at the Trot ...... 85

viii CHAPTER 1

INTRODUCTION

Lameness is the main cause of profit loss within the equine industry. But, what if owners could prevent their horses from developing musculoskeletal pain? Currently, western medicine is the ‘go-to’ option for owners in need of lameness treatment. A recent push in the industry towards alternative therapies such as chiropractic provides owners with options when traditional medicine is not an option [Sullivan et al., 2008]. Chiropractic is used in two different ways: 1) treating clinically diagnosed musculoskeletal pain alongside traditional methods and 2) treating equine athletes still in work in order to prevent a reduction in performance due to subclinical pain. While research exists to show how chiropractic affects horses suffering from clinically diagnosed back pain [Gomez Alvarez et al., 2008], more research is necessary to show the effects of chiropractic on riding sound horses. The study used eighteen horses from the UGA

Livestock Arena to determine how chiropractic affects horses in a riding program. Horses from this study were used by a university riding program where students rode the horses 4x per week.

Horses were separated into a Chiropractic (n=6), Riding (n=6), and Sedentary (n=6) group based on age, gender, lameness score, tack, and rider experience. Horses in the riding group received a grooming treatment while actively being ridden over the course of eight weeks. Sedentary horses received a grooming treatment and were not ridden over the course of this study. Horses placed in the chiropractic group underwent manual chiropractic therapy on Day 1 and on Day 29. PRE and POST kinematics for all groups were collected on Day 0, Day 2, Day 14, Day 28, Day 30,

Day 42, and Day 56. Heart rate monitors placed on the chiropractic group recorded changes in heart rate before, during, and after chiropractic treatment while monitors on the riding and sedentary group collected data before, during, and after grooming. Serum cortisol was collected via venipuncture by an experienced handler PRE, DURING, and POST treatment for all groups.

The study ended on Day 56. From this research, we hope to measure the effects of multiple chiropractic treatments on the musculoskeletal system of horses used in a regular exercise program.

2 LITERATURE CITED

1. Gomez Alvarez, C.B., L’Ami, J.J., Moffatt, D., Back, W., van Weeren, P.R. (2008).

Effect of chiropractic manipulations on the kinematics of back and limbs in horses with

clinically diagnosed back problems. Equine Vet J, 40(2): p. 153-9.

2. Sullivan, K.A., A.E. Hill, and K.K. Haussler (2008). The effects of chiropractic, massage

and phenylbutazone on spinal mechanical nociceptive thresholds in horses without

clinical signs. Equine Vet J, 40(1): p. 14-20.

3 CHAPTER 2

REVIEW OF LITERATURE

Alternative Therapies

Over the past several decades, the human model of alternative medicine has been adapted to fit the musculoskeletal and nervous systems of other species. Alternative therapies include any medicinal practices outside the realm of traditional western medicine. In conjunction with veterinary treatments, rehabilitation programs involving hydrotherapy, physiotherapy, laser therapy, electrotherapy, acupuncture, chiropractic, and massage exist to treat pain [Daglish and

Mama, 2016; Goff L., 2016]. Alternative therapies, unlike traditional treatments such as

NSAIDs, can be legally used while the horse is competing in various disciplines governed by the

United States Equestrian Federation and Federation Equestre Internationale [usef.org; fei.org].

Chiropractic

Chiropractic is defined as “high-velocity, low-amplitude thrust or impulse that moves a joint or vertebral segment beyond its physiologic range of motion, without exceeding the anatomic limit of the articulation” [Haussler, 1999]. A distinctive lack of research exists on the efficacy of chiropractic in an equine model. The use of chiropractic to treat musculoskeletal problems in horses was adapted from the human model [Haussler, 1999].

One case study involving an 85 year old man diagnosed with neuralgic amyotrophy showed that chiropractic adjustment 3x week for 4 weeks and a home exercise program relieved

4 his pain equal to that of prescription pain medications and his pain completely disappearing after

6 months and 18 additional chiropractic adjustments [Gliedt et al., 2017]. This case study utilized several different therapies including chiropractic manipulation to the thoracic lumbar, myofascial release therapy, scapular function exercise therapy, electrical stimulation to the serratus anterior, and a home exercise program to treat the symptoms of neuralgic amyotrophy

[Gliedt et al., 2017]. Another study looking at the effects of chiropractic on cervicogenic headaches showed that the headache index improves in chiropractic patients [Chaibi et al.,

2017]. A study involving 56 pregnant women with one-sided pelvic pain showed no differences between the experiment and control group, indicating that chiropractic was not more effective than traditional medicine when treating pelvic pain [Marie et al., 2017].

Some evidence exists to suggest that chiropractic has the same effect on nociceptive thresholds as phenylbutazone and massage in horses [Sullivan et al., 2008]. Mechanical noiceptive thresholds show at what point an animal experiences pain due to pressure [Sullivan et al., 2008]. One study utilized the ProReflex system using six infared cameras to capture the horses’ movements while moving on gravel or asphalt and showed that chiropractic has positive effects on the range of motion in horses suffering from back pain [Gomez Alvarez et al., 2008].

A case study involving the use of chiropractic to treat a horse with back pain showed that treatment positively affected the horse’s soundness and that horse was able to return to full work

[Faber et al, 2003]. Kinematics of the walk was measured at a constant velocity of 1.8 m/s and the trot measured at 4 m/s on a treadmill. The horse received chiropractic adjustment every three weeks for a total of six weeks. Kinematics were collected directly before, 48 hours, 3 weeks, 7 weeks, and 36 weeks after the first treatment using the ProReflex system. Results show that the temporal patterns were not affected by the chiropractic adjustment. Chiropractic did affect the

5 lateral excursion movement patterns, intervertebral symmetry, and axial rotation of the back

[Faber et al., 2003].

Acupuncture

Acupuncture utilizes small needles inserted into the body in order to treat pain and illness

[le Jeune et al., 2016]. A study conducted on 102 horses found that 82% of lame horses tested positive for pain during an acupuncture exam [le Jeune et al., 2014]. Acupuncture acts through acupoints, neurologic, connective tissue, fascial, and neurogenic regeneration mechanisms [le

Jeune et al., 2016]. The use of acupuncture in horses with pain along acupuncture meridians on

15 horses showed that 80% of them were cured after the initial treatment [Still, 2015]. When treating heel pain, however, acupuncture did not significantly affect pain versus a control group

[Robinson and Manning, 2015].

Veterinary Outlook on Alternative Therapies

A survey conducted by the University of Edinburgh shows that, of the 127 respondents, the majority of them support the use of alternative therapies to treat musculoskeletal back pain in horses [Bergenstrahle and Nielsen, 2015]. A survey conducted by the American Association of

Equine Practitioners showed that 20% of the veterinarians who responded to the survey practice some form of alternative therapeutic medicine [le Jeune et al., 2016].

Pain and Lameness

Pain

Pain is the result of injury or damage to the body. Nociceptors react to noxious stimuli and result in nociception [Muir, 2010]. Nociception is responsible for how the body reacts to the potentially harmful stimuli [Muir, 2010]. The two overarching types of pain are acute and chronic [Daglish and Mama, 2016]. Acute pain can result from nociceptive, inflammatory, and neuropathic pain. Inflammatory pain is the result of inflammation of a joint or tissue [Muir,

2010]. An example of an acute inflammatory pain would be the sharp reaction to a burn or cut.

Neuropathic pain involves illness and injury to the nervous system [Muir, 2010]. Chronic pain is the continuation of acute pain due a lack of complete healing [Daglish and Mama, 2016]. For example, the development of osteoarthritis due to trauma could cause chronic pain. A change in behavior and posture is a common indicator of pain. Recent studies, however, indicate the humans may not be adept at recognizing overall health and behavioral signs of distress in equines [Lesimple and Hausberger, 2014; Lesimple et al., 2013]. The use of a static surface electromyogram combined with neck postures has shown success in identifying riding horses with back pain versus a sedentary control group [Lesimple et al., 2012].

Equine Lameness

Pain causes equine lameness. At the European Eventing Championships, forty-five percent of the horses were unable to compete due to injury [Munsters et al., 2013]. Within the racing industry, lameness and illness costs $6.5 billion annually [Breton et al., 2013]. Equine lameness is based on a scale of 1-5 (Table 1) [Daglish and Mama, 2016]. The scale, provided by the American Association of Equine Practitioners (AAEP), gives practitioners and researchers a

7 standard to adhere to when diagnosing, treating, and studying lameness in the equine model.

Lameness is divided into “swinging” and “supporting” limb lameness. During gait analysis at the trot, lameness increases stance duration and stride frequency due to a loss in the force a limb exerts on the ground to propel it into the next phase of the stride in a supporting limb lameness

[Buchner et al., 1995]. Certified veterinarians diagnose and treat lameness via an exam [Clayton,

2016]. According to the AAEP, a lameness exam includes its medical history, a visual appraisal, hands-on exam, hoof testers, motion over different surfaces, and joint flexions [Lameness

Exams: evaluating a lame horse. aaep.org].

Table 1 AAEP lameness scale Grade 0 Lameness not perceptible under any circumstances Grade 1 Lameness is difficult to observe and is not consistently apparent, regardless of circumstances (eg. Under saddle, circling, inclines, hard surfaces). Grade 2 Lameness is difficult to observe at a walk or when trotting in a straight line but is consistently apparent under certain circumstances (eg. Weight-carrying, circling, inclines, hard surfaces). Grade 3 Lameness is consistently observable at a trot under all circumstances. Grade 4 Lameness is obvious at a walk. Grade 5 Lameness produces minimal weight bearing in motion and/or at rest or a complete inability to move. From http://www.aaep.org/info/horse-health?publication=836 Accessed January 7th, 2017.

Based on the veterinarian’s initial diagnosis, the horse undergoes diagnostic tests to definitively diagnose a lameness. Diagnostics include radiographs, nerve and joint blocks, ultrasound, scintigraphy, blood samples, joint fluid samples, and tissue biopsies [Lameness

Exams: evaluating a lame horse. aaep.org].

8 Treating Pain and Lameness

Current Treatments

Many treatments exist to treat equine lameness. Treatments range from simply icing the injury to in-depth veterinary care and rehabilitation. Joint lameness is often treated with injections of steroids and hyaluronic acid directly into the joint [aaep.org; Bentz and Revenaugh,

2013]. Surveys show that triamcinolone acetate (Vetalog) and methylprednisolone acetate

(Depo-Medrol™) are the most common steroids used in equine medicine [Ferris et al., 2011].

Veterinary prescriptions such as Legend® (hyaluronate sodium), via intravenous injection, and

Adequan® (polysulfated glycosaminoglycan), via muscular injection, are also used with a high degree of frequency to treat arthritis [Ferris et al., 2011]. Analgesics are also administered to treat pain [Muir, 2010]. Non-steroidal anti-inflammatory medication, also known as NSAIDs, can also be used to treat chronic lameness [aaep.org]. Commonly used NSAIDs are phenylbutazone, aspirin, Banamine, Diclofenac, Firocoxib, and Ketoprofen [Muir W., 2010].

Advanced therapies such as Interlukein Receptor Antagonist Protein (IRAP), Platelet Rich

Plasma (PRP), stem cell, and shockwave therapy are also used in many cases where joint injection or NSAIDS fall short of treating the lameness [aaep.org; Bentz and Revenaugh, 2013].

While effective in most cases, these therapies are expensive and treatments such as NSAIDS are illegal to give to the horse while it is competing.

Stress in Horses

Humans and horses have a unique relationship that goes back millennia to the original domestication of equines. Every day, horses and humans interact during all aspects of the domesticated horses’ routine including feeding, grooming, exercising, and veterinary treatment.

9 This does not mean, however, that the relationship is without stress. Polar heart rate monitors attached to the horses during rest, grooming, and warm-up showed that heart rate is lowest during rest and highest during handling and riding of the horses [Kowalik et al., 2016]. Rider experience, however, has no effect on the stress level of the horses during exercise [Kang and

Yun, 2016]. The increased heart rate during grooming and saddling indicates that grooming, while a necessary part of every day interaction between horse and rider, does stress the horse to some degree [Kowalik et al., 2017]. In humans, chiropractic has been shown to help non-PTSD veterans with neck and back pain [Dunn et al., 2009]. Research also shows that cancer patients use chiropractic as a complementary and alternative medicine during cancer treatment [Kang et al., 2014]. While chiropractic is used to treat stress in humans [Jamison, 1999], no research looks at the effects of chiropractic on stress parameters in equines.

Cortisol

Cortisol is a highly studied and consistent measure of stress in horses. Cortisol is a glucocorticoid released by the hypothalamic-pituitary- adrenal axis in response to a stress variable [Mostl and Palme, 2002]. Research shows that cortisol levels increase due to physical stimuli such as exercise and non-physical stimuli such as colic and transportation [Kedzierski et al., 2014; Mair et al., 2014; Fazio et al., 2016]. The administration of dexamethasone and fluticasone propionate suppresses the response of cortisol to a stressor [Munoz et al., 2015].

During research, cortisol samples are collected during rest to establish a baseline. Cortisol is collected via multiple different methods including serum, plasma, and saliva.

10 Serum Cortisol

Release of serum cortisol is the result of stimulation of the hypothalamic-pituitary- adrenal axis [Casella et al., 2016; Mair et al., 2014]. Venipuncture is used to collect serum cortisol [Peeters et al., 2011; Casella et al., 2006; Mair et al., 2014; Pazzola et al., 2015]. Serum free cortisol is the amount of cortisol in the blood that is not bound to cortisol binding globulin and albumin. The unbound cortisol, estimated to be 10% of the total plasma cortisol, is the cortisol that binds to receptors and effects the systems response Research shows that obese horses and horses suffering from an endocrine disease have higher amounts of serum free cortisol than healthy horses [Hart et al., 2016].

It is possible to induce increased levels of serum cortisol with the administration of an

ACTH or adrenocorticotropic hormone test [Monk et al., 2014]. An ACTH test involves the injection of synthetic ACTH in order to simulate the body’s response to a stressful event. This test is considered repeatable over an extended period of time in order to analyze the effects that

ACTH has on certain equine conditions [Scheidigger et al., 2016].

Research shows an increased level in serum cortisol during exercise. A study conducted on reining horses measured serum cortisol by jugular venipuncture before exercise, immediately after exercise, one-hour post exercise, two hours post exercise, and twenty-four hours post exercise with tubes containing no anticoagulant [Casella et al., 2016]. Results show a linear relationship between serum cortisol, heart rate, respiratory rate, and rectal temperature.

Researchers concluded that exercise stimulates the hypothalamic-pituitary-adrenal axis and results in peak cortisol levels immediately post exercise [Casella et al., 2016]. Results also show that while cortisol is highest immediately after exercise, it decreases rapidly post exercise

[Casella et al., 2016]. At Sartiglia tournament in Sardinia, Italy, serum cortisol was collected

from 21 competing horses. Results suggest that cortisol is lowest the day before and the day after the tournament. The highest cortisol levels occurred the day of the tournament directly after the horses completed the race [Pazzola et al., 2012].

Serum cortisol also rises in horses undergoing non-physical stimuli. One study looked at serum cortisol concentrations in horses suffering from colic [Mair et al., 2014]. This study was observational with the horses used from the Bell Equine Veterinary Clinic from January 2005 to

June 2008. The control group consisted of horses admitted for front foot lameness and the case group consisted of horses admitted to the clinic for acute colic. Results showed that horses suffering from colic with a heart rate of greater than forty-five beats per minute had a higher chance of serum cortisol concentrations of greater than 200 nmol/L. Results also showed that serum cortisol concentrations were higher in horses suffering from moderate to severe colic versus mild colic [Mair et al., 2014]. Horses also experience a rise in serum cortisol during transport despite previous transport experience [Fazio et al., 2016]. The amount of cortisol also depends on previous handling experiences. Horses that have positive experiences with their handlers during transport have lower levels of serum cortisol than horses with poor transport experiences [Fazio et al., 2016]. Exposure to the sun also increases serum cortisol in horses.

Horses left out in the sun have significantly elevated levels of cortisol over horses that are offered shade due to heat stress [Holcomb et al., 2013].

Salivary Cortisol

Salivary cortisol follows a diurnal circadian rhythm with the highest concentrations during the morning. The use of salivary cortisol as a measure of exercise-related stress in horses is frequently used in research [von Lewinski et al., 2013; Scheidegger et al., 2016; Becker-Birck et al., 2013; Schmidt et al., 2010.]. A direct enzyme immunoassay is used to measure cortisol

12 levels collected via this method [Schmidt et al., 2010]. Results showed highest concentration of cortisol at 6 AM with a decrease throughout the day and the highest total concentration in

December versus other months [Aurich et al., 2015]. Research shows that salivary cortisol follows similar patterns as serum cortisol and is collected via a metal clamp with a swab called a

Salivette [Peeters et al., 2011].

Horses experience increases in cortisol levels due to physical exertion and non-physical stimuli. During exercise, salivary cortisol is highest immediately after exercise and lowest while at rest [Kedzierski et al., 2014]. During dressage and show jumping competitions, horses experience a significant increase in salivary cortisol immediately post, 5-minutes post, and 15- minutes post riding on days 1 and 2 of a 3-day competition [Becker-Brick et al., 2013].

Hyperflexion, also known as rollkur, also significantly increases salivary cortisol levels versus a loose frame where the horses are allowed to stretch their necks [Christensen et al., 2014]. This research is particularly important to the dressage world where hyper flexion was regularly used as an alternative training method. Research also shows that salivary cortisol is significantly increased during exercise in 3-year old Warmbloods during their first 9-12 weeks of training and is higher in mares than stallions [Schmidt et al., 2010]. This increase in mares versus stallions could, however, be attributed to the fact that Warmblood stallions undergo a stallion test, which is a performance test over a certain period of time to judge their quality as a breeding animal, before they enter a training program [Schmidt et al., 2010].

Non-exercise events can also raise salivary cortisol levels in mature and young horses.

Body clipping, for example, is shown to increase salivary cortisol levels in horses undergoing this common winter grooming practice [Yarnell et al., 2013]. Foals also experience an increase

13 in salivary cortisol when exposed to stressful stimuli such as fire branding and microchipping

[Erber et al., 2012].

Plasma Cortisol

Plasma cortisol is collected via a blood tube containing an anticoagulant. A circadian rhythm exists in horses that do not suffer constant stressors from their environment. When placed in an entirely new environment, the circadian rhythm of peak cortisol in the morning from 6 a.m. to 9 a.m. and lowest from 1800 – 2100 hr., is completely disrupted [Irvine and Alexander, 1994].

Therefore, it is important to allow horses to become accustomed to a new environment or allow horses to maintain their accustomed schedule when performing research on horses. Research shows that plasma cortisol concentrations are highest immediately after exercise and lowest at rest [Kedzierski et al., 2014].

Heart Rate

Heart rate is a measurable factor when studying stress in horses. Epinephrine release due to a stressful event causes an increase in heart rate and vasodilation [Wagner A., 2010]. In a mature horse, the normal range for a horse’s heart beat is 20-40 beats per minute. During exercise, the horse’s heart can increase up to 240 beats per minute. The sympathetic nervous system controls increases in heart rate by releasing epinephrine [Erber et al., 2012].

Horse’s experience an increase in heart rate during exercise. During competition, the horse’s heart rate begins to increase while the horse is being tacked up and increases significantly during competition [Becker-Brick et al., 2013]. This increase in heart rate during competition is expected due to an increase in physical activity. While exercising, beat-to-beat

intervals and heart rate variability can be used to measure stress in horses. A decrease in beat-to- beat interval and heart rate interval indicates an increase in stress [Schmidt et al., 2010]. A study conducted on three-year old Warmbloods undergoing their first weeks of training reported a decrease in beat-to-beat interval and heart rate variability while mounting the horses and then returned to normal after the horses began exercising [Schmidt et al., 2010]. This increase indicates that mounting is considered a more stressful event to young horses than exercising with a rider. Training alters a horse’s response to stressful stimuli. Experienced horses ridden at the

French National School for Equitation were found to have to no significant changes in heart rate between riding with an audience versus without an audience [von Lewinski et al., 2013].

Heart rate and heart rate variability also increases in horses undergoing a non-exercise related event such as clipping. While heart rate increased in horses that do not stand for clipping, it decreased in horses that stand still for this grooming exercise [Yarnell et al., 2013]. Fire branding and microchipping also causes an increased heart rate in Warmblood foals [Erber et al.,

2012]. Transportation also causes an increase in heart rate and heart rate variability for the duration of transport [Schmidt et al., 2010].

Biomechanics of the Horse

Eadweard Muybridge was the first person during the 19th century to capture the horses’ gait using cameras [Clayton, 2016]. Horses have a distinct pattern of movement that is broken up into gaits. The horse has four main gaits: walk, trot, canter, and gallop. Specific breeds of horses also have several special gaits such as the tolt in Icelandic horses and the paso corto in Paso

Finos [Clayton, 2016].

15 Figure 1. Walk and Trot of the Horse From Hilary M. Clayton. HORSE SPECIES SYMPOSIUM: Biomechanics of the exercising horse. J. Anim. Sci. (2016) 94:4076-4086.

Movement during these gaits is controlled by ground reaction forces acting on the horse’s limbs to propel the horse in a particular direction [Clayton H., 2016]. The walk is a four-beat gait where four distinct hoof marks are left on the ground. During the walk, one stride would consist of ground contact in this order: right hind, right front, left hind, left front [Clayton H., 2016]. The trot is a two-beat gait where diagonal limbs contact the ground at the same time point. One stride consists of the right front, left hind in contact with the ground then the left front, right hind on the ground [Clayton, 2016].

Gait and Back Kinematics

Kinematics, also known as gait or back analysis, is defined as the study of movement

[Clayton H., 2016]. Analysis of the horse’s movement utilizes high-frame rate cameras and software to capture multiple aspects of the horse’s gait within a certain period of time. The use of these cameras allows researchers to measure changes to a horse’s gait that are not visible to the naked eye. Kinematics can be used to study rein tension, define gait quality, predict the quality of a horse as a foal, analyze movements at each gait, and test a horse’s jumping ability [Egenvall et al., 2015; Morales et al., 1998; Back et al., 1995; Willemen et al., 1997; Hodson et al., 1999;

16 Powers and Harrison., 1999]. Several aspects of the horses every day routine can affect the kinematics of the horse. A poorly fitted saddle negatively impacts communication between horse and rider and increases the risk of back pain in horses [Peham et al., 2004; Greve and Dyson,

2015]. The stride of the horse is affected by the tension a rider puts on the reins [Egenvall et al.,

2015]. The experience level of a rider alters a horse’s movement with less experienced riders hindering the natural gait and experienced riders being able to move with the horse [Greve and

Dyson, 2013]. A lack of rider balance can also lead to changes in the way a horse carries itself and increase the risk of chronic back pain [Lesimple et al., 2010]. Other studies have looked at the change in gait based on the addition of a rider [Hyun and Ryew, 2016]. Gait analysis is also used to analyze the effect that a treatment has on a horse’s gaits.

Gait Kinematics

Studying the kinematics of the horse involves temporal, linear, and angular measurements. Temporal kinematics involves measuring the difference between time points. At the walk, temporal measurements include stride duration, stance duration, lateral step interval, diagonal step interval, and support sequences [Hodson et al., 1999]. During a study involving

Spanish bred horses, total stride, diagonal limb pairs, and individual limbs divided temporal measurements taken at the trot up [Morales et al., 1998]. Total stride measurements included overlap, inter-overlap, and suspension. Diagonal limb pair measurements included diagonal support, diagonal swing, advanced placement, and lift off. Individual limb measurements involved stride duration, limb support, phases of limb support, and swing time [Morales et al.,

1998]. The swing phase of a horse’s gait starts from the time the hoof leaves the ground until it contacts the ground again [Leach et al., 1984]. The amount of time when a limb makes contact with the ground is called the stance phase. A stride, also known as stride duration, consists of

17 both the swing and the stance phase [Leach et al., 1984]. The suspension phase of a horse’s gaits is when no limbs are in contact with the ground. During overlap, two limbs are in contact with the ground at the same time [Leach et al., 1984]. For example, the time that the right front and right hind are in contact with the ground during the canter is overlap. Advanced placement is the amount of time that one limb contacts the ground before another limb within the same stride

[Leach et al., 1984]. At the trot, advanced placement would be the amount of time that the left hind contacted the ground before the right front within the left diagonal limb pair. At the walk, advanced placement could define the amount of time between right hind and right front contact.

Conversely, advanced lift off is the time period where one limb leaves the ground before the other paired limb [Leach et al., 1984].

Figure 2. Phases of the Trot. From Leach, DH. Ormrod, K. Clayton, HM. Standardised terminology for the description and analysis of equine locomotion. Equine vet. J. (1984) 16(6), 522-528

Linear kinematics involves measuring distance and angles. Common linear measurements include overtrack and stride length [Morales et al., 1998]. Stride length is the distance that a horse travels in one stride [Leach et al., 1984].

The use of angles is also measured via kinematics. Reflective markers attached to the skin of the horse track the movement of an underlying joint. The maximum and minimum angle of a joint is calculated for each joint in the horse’s front and hind legs [Morales et al., 1998]. The angle at the point where the limb makes contact with and leaves the ground is also measured via camera for each joint [Morales et al., 1998]. Protraction is the forward movement of a limb

whereas retraction is the backward movement of a limb [Morales et al., 1998]. The use of a force plate combined with angular measurements can also be used to identify the effects of weight on each joint in the limb [Hodson et al., 2001; Hodson et al., 2000; Hodson-Tole, 2001].

Back Kinematics

Kinematics is also used to study back pain in horses. A repeatable method for measuring back kinematics in two dimensions was developed by Utrecht University [Faber, 2002]. In this model, cameras are orthogonally placed on an x-, y-, and z- axis at 2 x 4 x 2.5 m3 while the horse remained on a treadmill [Faber et al., 2002; Johnston et al., 2004]. For analyzing back kinematics, the most common system is the ProReflex that uses three or six infared cameras to capture the horses’ movements on a treadmill [Faber et al., 2002; Wennerstrand et al., 2004;

Johnston et al., 2004]. A study conducted by the Swedish University of Agricultural Sciences developed a protocol using 9 or 12mm spherical reflective markers on the thoracic, lumbar, and sacral vertebrae for using kinematics to study the back in riding horses [Faber, 2002]. This protocol was then used to develop a database containing the mean and standard deviation of morphometric and spatiotemporal variables for the kinematics of sound horses [Johnston et al.,

2004]. Research shows that horses suffering from back pain have a decreased range of motion in flexion-extension at the walk and trot, increased lateral bending at T13 vertebra at the walk and trot, differences in lumbar symmetry at the walk, axial rotation a the walk, and a shorter stride length at the walk [Wennerstrand et al., 2004]. A few studies have looked at how chiropractic affects a horse’s back [Gomez Alvarez et al., 2008; Faber et al., 2003]. It is important to note that the equine back does change over time. Type of work, saddle fit, maturity, nutrition, and the rider all affect how the back changes over an extended period of time [Greve and Dyson, 2014].

Velocity

19 Research varies on velocity in equine kinematics. Several studies do not control for velocity during gait analysis at the walk [Hodson et al., 2000; Hodson et al., 2001; Hodson-Tole,

2001; Clayton et al., 2000]. At the trot, the controlled velocity rate varies between models. One model trots the horses at a velocity of 3 m/s [Clayton et al., 1998]. Other studies allow horses to move at their preferred speed during the trot [Gomez Alvarez et al., 2008; Morales et al., 1998a;

Morales et al., 1998b]. Recently, the use of treadmills has necessitated a controlled speed at which to walk and trot horses. Treadmills, however, require a training period due to alterations in the horses’ gaits from working in unnatural conditions [Buchner et al., 1994]. At the walk, a standard velocity for horses is 1.6 m/s [Faber et al., 2002; Johnston et al., 2004]. While trotting,

4 m/s is the standard velocity for horses during kinematic analysis [Faber et al., 2002; Johnston et al., 2004].

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29 CHAPTER 3

EFFECTS OF CHIROPRACTIC ON EQUINE GAIT KINEMATICS, HEART RATE, AND

SERUM CORTISOL

Gladney, J., Turner, K., West, F., Peroni, D., Lents, C., Duberstein, KJ. To be submitted to

Journal of Equine Veterinary Medicine

30 ABSTRACT

Heart rate monitors placed on all horses during treatment. Serum cortisol collected on all PRE,

MID, and POST treatment. At the trot, swing as a percentage of stride decreased in hind limbs on

Day 56 compared to Day 28 (p=0.05), Day 14 (p=0.03), and Day 2 (p=0.02) in Chiropractic group. Cortisol is higher on Day 29 PRE vs. Day 1 PRE (P=0.03) in the Chiropractic group.

Results show chiropractic minimally effects kinematics and increases PRE cortisol levels.

31 INTRODUCTION

Maintaining soundness is of the highest priority to the owners of equine athletes. Treating musculoskeletal pain is often expensive and, in some cases, unsuccessful. This desire to treat and possibly prevent the effects of clinical and subclinical musculoskeletal pain has given birth to a wide variety of alternative therapies. One of those therapies is equine chiropractic. Chiropractic is defined as “high-velocity, low-amplitude thrust or impulse that moves a joint or vertebral segment beyond its physiologic range of motion, without exceeding the anatomic limit of the articulation” and was adapted to treat horses from a human model [Haussler, 1999].

Current studies show that the use of chiropractic as an alternative therapy could have the same effects as phenylbutazone and massage [Sullivan et al., 2008] and treat horses suffering from clinically diagnosed back pain [Gomez Alvarez et al., 2008; Faber et al, 2003]. Mechanical noiceptive thresholds show at what point an animal experiences pain due to pressure and some evidence exists to suggest that chiropractic has the same effect on nociceptive thresholds as phenylbutazone and massage in horses [Sullivan et al., 2008]. Another study using 10 warmblood horses showed that chiropractic has positive effects on the range of motion in horses suffering from back pain [Gomez Alvarez et al., 2008]. A case study also used chiropractic to treat a horse with back pain showed that treatment positively affected the horse’s soundness and that horse was able to return to full work [Faber et al, 2003]. No studies, however, exist on the effects of chiropractic in the riding sound horse.

Kinematics is defined as the study of movement, utilizes high-frame rate cameras to study a horse’s gaits, and ascertain how treatments affect those gaits [Clayton, 2016]. Kinematics has been used to study rein tension, define gait quality, predict the quality of a horse as a foal,

analyze movements at each gait, test a horse’s jumping ability, and look at how a rider effects the horse’s movement [Egenvall et al., 2015; Morales et al., 1998; Back et al., 1995; Willemen et al., 1997; Hodson et al.; 1998. Powers and Harrison, 1999; Hyun and Ryew, 2016].

While some studies exist regarding the effects of chiropractic on horses already suffering musculoskeletal pain [Gomez Alvarez et al., 2008; Faber et al, 2003;Sullivan et al., 2008], no studies exist measuring the effects of chiropractic on horses that are ridden regularly. It would be beneficial to explore the effects of chiropractic on riding sound horses considering chiropractic is often performed on equine athletes in an exercise program.

The objective of this study was to ascertain the effects of chiropractic on the riding sound horse using gait kinematics and physiological stress markers when compared to their riding and sedentary counterparts. The hypotheses is that chiropractic will positively affect the temporal and linear parameters of gait kinematics and will not be more stressful than a grooming treatment.

33 MATERIALS AND METHODS

Horses

Eighteen stock type horses ranging in age from 4-18 years, 13 geldings and 5 mares, in the care of the University of Georgia Livestock Arena were used in this study. The horses were divided into three groups of six (chiropractic, riding, and sedentary) based on age, lameness score, riding type (English/Western), rider experience (Experienced/Beginner), gender

(Mare/Gelding), living arrangements (Stalled/Pasture), and by which group of students rode the horses (Research/Class). The groups dictated which treatment the horses received. Horses in the chiropractic group received a chiropractic treatment on Day 1 and Day 29 while being ridden over the course of this study. The riding group received a grooming treatment on Day 1 and Day

29 while being ridden over the course of this study. Horses in the sedentary group received a grooming treatment on Day 1 and Day 29 and were not ridden during this study. Horses in the

Riding and Sedentary groups were groomed by experienced handlers for twenty minutes. Horse in the Chiropractic group received a chiropractic adjustment that also lasted about 20 minutes.

The Animal Care and Use Committee approved this study.

Lameness Evaluation

Before the start of the study, a licensed veterinarian performed a lameness exam on all 18 horses. An experienced handler presented each horse to the veterinarian one at a time. The veterinarian remained blind as to which horses would be placed in what group before and after the lameness exam. All horses were deemed sound to participate in this study.

34 Riding

Horses in the chiropractic and riding groups were ridden four days a week. The horses were ridden for forty minutes each day at the walk, trot, and canter. Horses in the riding and chiropractic groups were exercised before gait analysis on all days. The sedentary group was not ridden during the course of this study. On Day 1 and Day 29, horses in the riding and chiropractic groups were exercised before treatment and allowed time to rest for at least thirty minutes before receiving treatment.

Chiropractic

Horses placed in the chiropractic group received two chiropractic treatments from a veterinarian with a chiropractic license on Day 1 and Day 29. None of the horses had previously received a chiropractic adjustment. The veterinarian had no contact with the horses before the study. During treatment, the veterinarian adjusted each part of the horse by hand. No instruments were used to treat the horses. Parts of the horse examined and adjusted are as follows: TMJ

(Jaw), Occiput (Poll), Cervical (Neck), Thoracic (Upper Back), Lumbar (Lower Back), Sacrum

(Pelvic and Tail), and Legs. The cervical vertebrae consists of C1-C7. The thoracic portion includes T1-T18, right ribs, left ribs, logan, and intertransverse. The lumbar consists of L1-L6 and the sacrum includes the base, apex, Sacroiliac Joint Posterior Inferior (SIJ PI), and Sacroiliac

Joint Anterior Superior (SIJ AS), coccyx. The legs consist of the scapula, left front, right front, and hind legs.

Kinematic Analysis

Floor markers were used to set up the system the same way on Day 0, Day 2, Day 14,

Day 28, Day 30, Day 42, and Day 56. A 0.914 meter wide and 8 meter long runway was used to

walk and trot horses. Two Ethernet GigE uEyeTM camera (IDS Imaging Development System,

Obersulm, Germany) were used to record the videos at 70 frames per second. The cameras were set up perpendicular to the runway at 6.25 meters away on the left and right side. A Farmtek

MD-300 Electronic Timer (Farmtek Inc., Wylie, TX, USA) was used to measure the time it took horse and handler to travel 8 meters in order to control velocity. The target velocity of the walk was 1.6 m/s and 4 m/s at the trot. The time frame allowed for the walk is 4.84-5.16 seconds. The allotted time frame for the trot is 1.8-2.2 seconds. Horses were walked and trotted in hand for a maximum of 15 runs at the walk and trot in order to get 6 repetitions within the time frame.

Reflective tape used to calibrate the linear measurements were placed on the left and right shoulder of the horse measured 12.7 centimeters.

The same handler was used for all horses and days. Before the recorded runs, horses were walked up and down the runway to ensure the horses were comfortable with the equipment.

During the recorded runs, horses started moving 4 meters before the timer and continued moving

4 meters past the timer to ensure that movement was consistent throughout the entire run. Horses in the chiropractic and riding groups were ridden before gait analysis and allowed to rest for at least thirty minutes on each recording day. Sedentary horses were not exercised at any point of this study and were taken directly from their living arrangements to gait analysis.

The videos were downloaded and measured in Kinovea (Version 0.8.15). The following parameters were measured at the walk and trot: swing duration, stance duration, and stride duration of the left and right front limbs, as well as swing as a percent of stride duration for both front limbs. The same parameters were also measured on both hind limbs. Swing time was defined as the period of time from where the hoof left the ground until the hoof touched the ground again. Stance time was defined as the period of time from when the hoof touched the

36 ground until it lost contact with the ground again. Stride duration was the addition of swing and stance time. Swing time as a percentage stride duration was defined as swing time divided by stride duration. Stance time as a percentage stride duration was defined as stance time divided by stride duration. Suspension time was defined as the length of time all four hooves were off the ground between diagonal pairs at the trot. Stride length was defined as the linear measurement from one hoof leaving the ground until that foot touched the ground again.

Heart Rate

On Day 1 and Day 29, horses from all groups were brought into the barn for treatment one at a time. A Polar RS300x sa heart rate monitor (Polar Electro Oy, Professorintie 5, FI-

90440 Kempele, Finland) was placed on the horses using an elastic band at the heart girth.

Electrodes placed at the heart girth and withers measured the heart rate of each horse. Ultrasound gel was placed on the electrodes. The mode “Other Sport 2” was selected from the options. A timer was started when the horses’ heart rate could be read on the watch dial attached to the elastic band. At the end of the session, the monitors were stopped, removed, cleaned, and stored until the next horse. The saved data was then transferred to polarpersonaltrainer.com via usb cable. The monitors recorded time, heart rate minimum, heart rate maximum, and heart rate average.

Serum Cortisol

Collection

Blood was collected via jugular venipuncture with a vacutainer needle at 0 minutes

(PRE), 10 minutes (MID), and 20 minutes (POST). Each horse was selected randomly to prevent

the circadian rhythm from affecting the levels of serum cortisol. Two blood tubes were filled at each time point. Blood tubes were then refrigerated at 35ºF until centrifuging. Blood was centrifuged 4-6 hours after collection to separate serum. Serum was stored in a -20 freezer until cortisol analysis was performed. After centrifuging, the blood tubes were placed in a rack and the serum pipetted into 1.5mL microcentrifuge tubes labeled with the horse’s number, time point, and day. Transfer pipets (Samco Scientific, General Purpose, Lg Bulb) were used to move the serum to the 1.5mL microcentrifuge tubes and a new pipet was used for each serum transfer. The serum was then frozen at -20ºF in Styrofoam containers.

Assay

The serum cortisol was sent to the USDA (Clay Center, Nebraska 68933) for assay.

Concentrations of cortisol were determined using a commercial radioimmunoassay kit (06B-

256440; MP Biomedicals LLC., Irvine, CA). The assay was conducted according to the manufacture’s recommendations except that the standard curve ranged from 0.25 to 60 μg/dL, the sample (25 μL) was diluted in 100 μL of buffer (0.1M PBS, pH 7.5) before assay, the incubation period was 1 h, and 1 mL of ice-cold buffer was added to the tube before decanting.

A pooled sample of plasma from all horses in the study was parallel to the standard curve when serial dilutions were assayed (P = 0.93). When 70 ng of cortisol were added to the pooled plasma sample or buffer, recovery was 90% and 104%, respectively. Intra- and Interassay coefficient of variation of 2.3% and 4.5%, respectively, were calculated from a pooled sample of horse plasma that measured 1.9 μg/dL. Sensitivity of the assay (0.1 ug/dL) was defined as 90% of the binding of the zero tube.

38 Statistical Analysis

SAS version 9.4 (Cary, NC) proc mixed for a repeated measures over time model with treatment and day variables was used to analyze the data sets for kinematic, heart rate, and serum cortisol. A statistical significance of P<0.05 was set for all data.

Timeline

The timeline for the entire study is as follows:

Day -33: Horses start exercise program

Day -10: Lameness Evaluation performed on all 18 horses

PHASE 1

Day 0: Gait Analysis performed on all horses

Day 1: Chiropractic horses receive chiropractic treatment, Riding and Sedentary horses grooming treatment for twenty minutes, blood pulled PRE, MID, and POST on all horses, heart rate monitors placed on all horses for duration of treatment. Blood is spun and serum placed in tubes.

Day 2: 1-day POST gait analysis on all horses

Day 3-13: Horses ridden and handled by students

Day 14: 2-Weeks POST gait analysis performed on all horses

Day 15-27: Horses ridden and handled by students

Day 28: 4-Weeks POST gait analysis performed on all horses

PHASE 2

Day 28: PRE gait analysis performed on all horses

39 Day 29: Chiropractic horses receive chiropractic treatment, Riding and Sedentary horses grooming treatment for twenty minutes, blood pulled PRE, MID, and POST on all horses, heart rate monitors placed on all horses for duration of treatment. Blood is spun and serum placed in tubes.

Day 30: 1-day POST gait analysis on all horses

Day 31-41: Horses ridden and handled by students

Day 42: 2-Weeks POST gait analysis performed on all horses

Day 43-55: Horses ridden and handled by students

Day 56: 4-Weeks POST gait analysis performed on all horses

STUDY ENDS

40 RESULTS

Walk

Twenty-one parameters were measured from the gait analysis of the walk.

No significant changes were seen for the front limbs at the walk (Figure 3.1).

41 At the walk, hind swing as a percentage of stride increased and a corresponding decrease in stance as a percentage of stride occurred in the Riding group between Day 2 and Day 30

(p=0.05) (Figure 3.2).

42 Trot

Twenty-two parameters were measured for the gait analysis of the trot from eighteen horses.

No significant changes to the front limbs at the trot (Figure 4.1).

At the trot, swing as a percentage of stride decreased in the hind on Day 56 when compared to Day 28 (p=0.05), Day 14 (p=0.03), and Day 2 (p=0.02) in the Chiropractic group

(Figure 4.2). Hind swing as a percentage of stride decreased and the corresponding increase in stance as a percentage of stride occurred on Day 0 (p=0.04), Day 2 (p=0.01), and Day 30

(p=0.05) when compared to Day 56 in the Sedentary group (Figure 4.2).

44 In the Sedentary group, the stride length decreased on Day 30 (P=0.02) when compared to Day 0 (Figure 5).

45 Serum Cortisol

The results of the serum cortisol are listed for PRE, MID, and POST on Day 1 and Day

29 for all groups.

Table 2. Treatment*Day lsmeans of Cortisol (nmol/L) Treatment Time Point Day 1 Day 29 SEM Chiropractic PRE 63.0a 112.2b 15.6 MID 67.5 94.6 12.5 POST 82.5 90.0 11.6 Riding PRE 68.0 96.5 15.6 MID 69.1 82.8 12.4 POST 82.8 73.1 11.6 Sedentary PRE 60.9 91.1 15.6 MID 72.8 75.0 12.9 POST 77.0 70.6 11.6 PRE cortisol pulled immediately before treatment (0 Minutes), MID cortisol at midpoint of treatment (10 minutes), and POST cortisol pulled immediately post treatment (20 minutes) a,b,c Means within rows with different superscripts differ (P < 0.05) When comparing the same time point on different days, cortisol is higher on Day 29 PRE vs. Day 1 PRE (P=0.03) (Table 2).

Heart Rate

Three parameters (Min, Avg, and Max) were collected for the heart rate via the Polar

Heartrate Monitors.

Table 3. Treatment*Day lsmeans of Heart Rate (bpm) Treatment Heart Rate Day 1 Day 29 SEM Chiropractic Min 27a 34b 2.4 Avg 44 41 3.0 Max 68 73 7.6 Riding Min 31 34 2.3 Avg 39 42 2.9 Max 69 66 7.3 Sedentary Min 35 36 2.5 Avg 45 44 3.1 Max 67 80 7.7 From Polar RS300x sa heart rate monitor set at ‘Other Sport 2’. Min heart rate is the minimum heart rate for each group. Avg is the average heart rate for each group. Max heart rate is the maximum heart rate for each group. a,b,c Means within rows with different superscripts differ (P < 0.05)

When comparing the Min heart rate across Day 1 and Day 29, Chiropractic Day 1 Min is lower than Day 29 (P= 0.05) (Table 3).

47 DISCUSSION

Linear and temporal kinematics show minimal differences in sound horses as a result of chiropractic treatments. While previous research has been shown to positively alter the kinematics of horses with clinically diagnosed back pain [Sullivan et al., 2008; Gomez Alvarez et al., 2008; Faber et al, 2003], this study did not find that chiropractic treatment has any significant benefit to sound horses in a riding.

Research studies show that back pain in horses participating in a riding program is far more prevalent than originally thought and the onset of back pain in equine athletes leads to a reduction in performance [Mothershead et al., 2017; Sukovaty et al., 2017; Fantini and Palhares,

2011]. A social media survey shows that 42% of equine veterinarians see at least one case per week of horses suffering from back pain [Bergenstrahle and Neilsen, 2016]. One study using 20 polo ponies showed that 19 of them showed symptoms of back pain while still being ridden and another study showed that 14 of 19 lesson horses examined for this study suffered from severe back problems [Biermann et al., 2014; Lesimple et al., 2010]. Other studies indicate that people are not adept at recognizing overall health and behavioral signs of distress in equines [Lesimple et al., 2014. Lesimple et al., 2013].

This study was performed to see if chiropractic could alleviate the back pain in working horses. While the riding program in this study was not heavy enough to induce back pain in horses still considered riding sound, as evidenced by the lack of kinematic changes in the riding group from Day 0 to Day 56, future studies could look at horses under more serious work. From this study, we can conclude that chiropractic does not improve gait quality in sound horses not experiencing back pain. This conclusion is supported by the fact the chiropractic horses did not

show any improvement throughout this study over Day 0. The temporal and linear measurements taken in this study may not be sensitive enough to detect subclinical changes in horses. Future studies could utilize more sensitive kinematic systems, force plate kinetics, and pressure algometry.

All horses showed numerical increase in cortisol levels on Day 29 PRE vs Day 1 PRE, whereas the chiropractic group was significant. The change in cortisol was accompanied by a change in the MIN heart rate but not AVG and MAX heart rate. Studies measuring serum cortisol in horses show an increase in concentrations from non-exercise stimuli including horses suffering moderate to severe colic versus mild colic, undergoing transportation, and heat stress

[Mair et al, 2014; Fazio et al., 2016; Holcomb et al., 2014]. In Mair et al., horses suffering from colic with a heart rate of greater than forty-five beats per minute had a higher chance of serum cortisol concentrations of greater than 200 nmol/L [Mair et al, 2014]. In a study that looked at the effects physical factors and season on serum cortisol concentrations in horses, healthy horses averaged between 1-11 mg/dl (27.59-303.49 nmol/L) [Hart et al., 2016]. Serum cortisol concentrations in horses averaged 188.81±51.46 nmol/L at rest and increased to 356.98±55.29 nmol/L after ACTH challenge [Peeters et al., 2011]. During the ACTH challenge, serum cortisol concentrations increased after 10 minutes and peaked at 96±16.7 minutes [Peeters et al., 2011].

In horses undergoing transport, serum cortisol increased from 123.22 nmol/L before transport to

283.75 nmol/L after 45 minutes of transport [Fazio et al., 2016]. In the Chiropractic group, the serum cortisol increased by 49.2 nmol/L from Day 1 PRE to Day 29 PRE. These results indicate that the Chiropractic group was more stressed than the other groups but not to the same degree that transport or an ACTH challenge elevates stress markers. It is important to note that while serum cortisol was elevated in the Chiropractic group, cortisol levels are still within normal

49 range for a horse. Research shows that motor units of a neuron react to the anticipation of pain and actual pain versus a control [Tucker et al., 2012]. Research also shows that rats find the anticipation of stress more stressful than an acute stressor [Yamamotová et al., 2000]. Future studies could examine behavioral and physical indicators of stress in horses undergoing chiropractic to more accurately measure the level of stress that the horse feels due to treatment.

In conclusion, our results show that chiropractic adjustment has minimal effects on horses who do not suffer from back pain and that the anticipation of chiropractic treatment might cause a degree of stress.

The researchers claim no bias in this study.

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55 CHAPTER 4

CONCLUSIONS

Horses and humans have a unique relationship. That relationship has evolved from prey, their domestication around 4000 BCE, and through the Ages to today [Johns, 2006]. Today, horses work, compete, and provide companionship to humans all over the world. As part of this unique relationship, an entire industry exists around the well-being of the horse. One small part of this industry is the use of chiropractic to treat clinical and subclinical pain.

Chiropractic was brought to the United State in 1895 but dates back to 2700 BCE [Nesci,

2017]. Today, chiropractic is used to treat musculoskeletal pain by adjusting the spine and pelvis

[Nesci, 2017]. Anyone suffering from joint pain, muscle soreness, nerve pain, etc. can seek treatment from a doctor of chiropractic. A recent survey showed that the majority of respondents believe that chiropractic is an effective measure for treating neck and back pain [Weeks et al.,

2015]. In humans, chiropractors attend four years of chiropractic school in order to be certified to treat patients [Nesci, 2017]. In 1897, Palmer School of Chiropractic opened and was the first chiropractic college in the United States [Chiropractic, 2016]. Anyone wishing to treat patients via chiropractic can attend an accredited chiropractic college and become certified.

In horses, chiropractic evolved from the human model in the late 20th century [Haussler,

1999]. If owners of equine athletes wish to treat their horse with chiropractic, the AAEP recommends seeking out a veterinarian with certification in chiropractic care from the American

Veterinary Chiropractic Association and is licensed to practice in the state of residence [Harman

56 and Pascoe, 2001; Yates, 2017]. It may also be beneficial to set up a program with a veterinarian that takes the horses specific needs into account.

Treating this pain is beneficial to owners of equine athletes, both professional and amateur, because it prevents the loss of a valuable, and often well-loved, competition horse.

Research also shows that chronic discomfort and pain can cause aggression in horses [Fureix et al., 2010]. With this in mind, chiropractic treatment, with the approval of the attending veterinarian, has the potential to prevent or reduce aggression in horses caused by certain types of pain. Future studies could look at how chiropractic affects horses with aggressive tendencies who are diagnosed with chronic pain. This could prevent injury to the owners and handlers of aggressive horses.

Treating clinical and subclinical musculoskeletal pain in animals, and especially in horses, is difficult and complex. Previous studies show that chiropractic can benefit horses with debilitating back pain [Gomez Alvarez et al., 2008]. The results of this study, however, showed that horses in a light riding program do not benefit from chiropractic in an objectively measurable way. This research study also showed that the anticipation of chiropractic does cause a slight degree of stress to horses. While future studies with horses in more intense exercise may have different results, it is important to remember that chiropractic is just one of the multitude of alternative and traditional therapies that can be used to treat the equine athlete.

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APPENDIX A

70 APPENDIX B

Table 4. Treatment*Day lsmeans of the Front Limbs Measurements at the Walk

Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing Time C Left 0.41 0.40 0.40 0.40 0.41 0.41 0.40 0.009

(seconds) Right 0.41 0.40 0.40 0.40 0.39 0.40 0.40 0.009

R Left 0.42 0.42 0.41 0.42 0.42 0.41 0.42 0.009

Right 0.42 0.41 0.41 0.41 0.41 0.41 0.41 0.009

S Left 0.42 0.41 0.41 0.41 0.42 0.42 0.41 0.009

Right 0.41 0.40 0.40 0.41 0.42 0.41 0.41 0.009

Stance Time C Left 0.71 0.71 0.71 0.69 0.69 0.70 0.70 0.02

(seconds) Right 0.72 0.71 0.71 0.70 0.69 0.70 0.70 0.02

R Left 0.71 0.72 0.71 0.70 0.70 0.71 0.69 0.02

Right 0.72 0.72 0.72 0.70 0.69 0.70 0.71 0.02

S Left 0.71 0.68 0.69 0.70 0.69 0.66 0.68 0.02

Right 0.71 0.68 0.71 0.69 0.70 0.71 0.70 0.02

Stride C Left 1.12 1.11 1.11 1.09 1.10 1.11 1.10 0.03

Duration Right 1.11 1.11 1.10 1.10 1.09 1.10 1.10 0.02

(seconds) R Left 1.14 1.13 1.12 1.11 1.11 1.12 1.11 0.03

Right 1.14 1.13 1.12 1.11 1.10 1.11 1.12 0.02

S Left 1.13 1.09 1.10 1.11 1.11 1.05 1.09 0.03

Right 1.10 1.08 1.10 1.09 1.12 1.12 1.09 0.02

71 Swing C Left 0.37 0.36 0.36 0.37 0.37 0.37 0.36 0.007

(as a % of Right 0.38 0.36 0.36 0.36 0.36 0.36 0.37 0.006

stride R Left 0.37a,b 0.37a,b 0.36a 0.37a,b 0.37a,b 0.37a,b 0.38b 0.007

duration) Right 0.37 0.36 0.36 0.37 0.37 0.37 0.37 0.006

S Left 0.37 0.38 0.37 0.37 0.38 0.42 0.38 0.007

Right 0.38a,b 0.37a 0.36b 0.37a,b 0.37a,b 0.37a,b 0.37a,b 0.006

Stance C Left 0.63 0.64 0.64 0.63 0.63 0.63 0.64 0.004

(as a % of Right 0.67 0.64 0.64 0.64 0.64 0.64 0.63 0.008

stride R Left 0.63a,b 0.63a,b 0.64a 0.63a,b 0.63a,b 0.63a,b 0.62b 0.004

duration) Right 0.63 0.64 0.64 0.63 0.63 0.63 0.63 0.008

S Left 0.63 0.63 0.63 0.63 0.63 0.63 0.62 0.004

Right 0.66a,b 0.63a 0.64b 0.63a,b 0.63a,b 0.63a,b 0.63a,b 0.008

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 5. Treatment*Day lsmeans of the Back Limbs Measurements at the Walk

Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing C Left 0.42 0.40 0.41 0.40 0.41 0.4 0.41 0.01

Time Right 0.41 0.42 0.41 0.41 0.41 0.41 0.41 0.01

(seconds) R Left 0.43 0.43 0.42 0.42 0.43 0.42 0.42 0.01

Right 0.42 0.41 0.42 0.41 0.42 0.42 0.42 0.01

S Left 0.42 0.40 0.41 0.41 0.42 0.42 0.42 0.01

Right 0.41 0.40 0.41 0.41 0.41 0.41 0.40 0.01

Stance C Left 0.71 0.71 0.70 0.70 0.68 0.69 0.70 0.02

72 Time Right 0.71 0.70 0.71 0.68 0.69 0.70 0.70 0.02

(seconds) R Left 0.72 0.71 0.71 0.70 0.69 0.70 0.70 0.02

Right 0.73 0.73 0.71 0.71 0.71 0.71 0.70 0.02

S Left 0.71 0.70 0.70 0.70 0.70 0.70 0.70 0.02

Right 0.723 0.70 0.71 0.70 0.71 0.72 0.70 0.02

Stride C Left 1.13 1.11 1.11 1.09 1.09 1.11 1.11 0.02

Duration Right 1.13 1.11 1.11 1.09 1.09 1.11 1.11 0.02

(seconds) R Left 1.15 1.13 1.13 1.11 1.11 1.12 1.11 0.02

Right 1.15 1.14 1.12 1.12 1.12 1.13 1.11 0.02

S Left 1.14 1.09 1.10 1.11 1.12 1.12 1.10 0.02

Right 1.14 1.09 1.11 1.11 1.12 1.13 1.09 0.02

Swing C Left 0.37 0.36 0.37 0.37 0.37 0.37 0.37 0.005

(as a % of Right 0.37 0.37 0.37 0.37 0.37 0.37 0.37 0.004

stride R Left 0.38 0.37 0.37 0.38 0.38 0.38 0.38 0.005

duration) Right 0.37a,b 0.36a 0.37a,b 0.37a,b 0.37a,b 0.37a,b 0.37b 0.004

S Left 0.37 0.37 0.37 0.37 0.38 0.38 0.38 0.005

Right 0.36 0.36 0.37 0.37 0.37 0.36 0.36 0.004

Stance C Left 0.63 0.64 0.63 0.63 0.63 0.63 0.63 0.005

(as a % of Right 0.63 0.63 0.63 0.63 0.63 0.63 0.63 0.004

stride R Left 0.62 0.63 0.63 0.63 0.62 0.62 0.62 0.005

duration) Right 0.63a,b 0.64a 0.63a,b 0.63a,b 0.63a,b 0.63a,b 0.63b 0.004

S Left 0.63 0.63 0.63 0.63 0.62 0.63 0.62 0.005

Right 0.64 0.64 0.64 0.63 0.63 0.64 0.64 0.004

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 6. Treatment*Day lsmeans of the Front Limb Measurements at the Trot

Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing Time C Left 0.38 0.37 0.36 0.36 0.36 0.36 0.35 0.01

(seconds) Right 0.37 0.37 0.36 0.35 0.35 0.35 0.35 0.01

R Left 0.39 0.38 0.37 0.37 0.37 0.37 0.37 0.01

Right 0.38 0.38 0.37 0.36 0.37 0.37 0.37 0.01

S Left 0.36 0.36 0.35 0.35 0.35 0.34 0.35 0.01

Right 0.36 0.36 0.35 0.36 0.36 0.35 0.35 0.01

Stance Time C Left 0.24 0.24 0.23 0.23 0.23 0.23 0.24 0.008

(seconds) Right 0.24 0.24 0.23 0.23 0.23 0.23 0.24 0.008

R Left 0.24 0.24 0.23 0.22 0.23 0.23 0.23 0.008

Right 0.25 0.24 0.23 0.23 0.23 0.23 0.24 0.008

S Left 0.24 0.24 0.24 0.23 0.24 0.23 0.24 0.008

Right 0.24 0.24 0.23 0.23 0.23 0.24 0.24 0.008

Stride C Left 0.62 0.61 0.58 0.58 0.59 0.59 0.60 0.02

Duration Right 0.62 0.60 0.59 0.58 0.58 0.59 0.59 0.02

(seconds) R Left 0.63 0.62 0.60 0.59 0.60 0.60 0.60 0.02

Right 0.63 0.62 0.60 0.60 0.60 0.60 0.60 0.02

S Left 0.60 0.59 0.58 0.59 0.59 0.58 0.59 0.02

Right 0.60 0.60 0.58 0.58 0.59 0.59 0.60 0.02

Swing C Left 0.61 0.60 0.61 0.61 0.60 0.61 0.60 0.006

Right 0.61 0.60 0.61 0.60 0.60 0.60 0.59 0.007

74 (as a % of R Left 0.62 0.62 0.62 0.63 0.62 0.62 0.61 0.006

stride Right 0.61 0.61 0.62 0.61 0.61 0.61 0.61 0.007

duration) S Left 0.61 0.60 0.59 0.60 0.60 0.60 0.59 0.006

Right 0.60 0.61 0.60 0.61 0.61 0.60 0.59 0.007

Stance C Left 0.39 0.40 0.39 0.39 0.40 0.40 0.41 0.006

(as a % of Right 0.40 0.40 0.39 0.40 0.40 0.40 0.41 0.007

stride R Left 0.39 0.39 0.39 0.38 0.38 0.39 0.39 0.006

duration) Right 0.39 0.39 0.39 0.39 0.39 0.39 0.39 0.007

S Left 0.40 0.40 0.41 0.40 0.40 0.41 0.41 0.006

Right 0.40 0.40 0.40 0.39 0.39 0.40 0.41 0.007

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 7. Treatment*Day lsmeans of the Back Limb Measurements at the Trot

Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing Time C Left 0.39 0.39 0.38 0.37 0.37 0.36 0.37 0.01

(seconds) Right 0.38 0.38 0.37 0.37 0.37 0.36 0.36 0.01

R Left 0.4 0.39 0.38 0.38 0.39 0.39 0.38 0.01

Right 0.39 0.39 0.38 0.37 0.38 0.37 0.37 0.01

S Left 0.38 0.38 0.36 0.37 0.37 0.36 0.36 0.01

Right 0.38 0.37 0.36 0.36 0.37 0.36 0.36 0.01

Stance Time C Left 0.23 0.22 0.22 0.22 0.22 0.22 0.23 0.008

(seconds) Right 0.23 0.23 0.22 0.22 0.23 0.22 0.24 0.009

R Left 0.24 0.23 0.22 0.23 0.22 0.22 0.23 0.008

75 Right 0.24 0.23 0.22 0.23 0.23 0.24 0.23 0.009

S Left 0.23 0.23 0.22 0.22 0.22 0.23 0.23 0.008

Right 0.23 0.23 0.22 0.23 0.23 0.23 0.24 0.009

Stride C Left 0.62 0.61 0.60 0.59 0.60 0.59 0.59 0.02

Duration Right 0.62 0.61 0.59 0.59 0.59 0.59 0.60 0.02

(seconds) R Left 0.63 0.62 0.60 0.61 0.61 0.61 0.61 0.02

Right 0.63 0.62 0.61 0.61 0.61 0.61 0.60 0.02

S Left 0.61 0.60 0.59 0.59 0.60 0.58 0.59 0.02

Right 0.61 0.60 0.58 0.60 0.60 0.59 0.60 0.02

Swing C Left 0.62a,b 0.63a 0.63a,b 0.63a,b 0.62a,b 0.62a,b 0.62b 0.006

(as a % of Right 0.62a,b 0.63a 0.63a 0.63a 0.62a,b 0.62a,b 0.61b 0.006

Stride R Left 0.63 0.63 0.63 0.62 0.64 0.63 0.63 0.006

Duration) Right 0.62a 0.63a,b 0.63b 0.62a,b 0.62a,b 0.61a 0.61a 0.006

S Left 0.63 0.63 0.62 0.62 0.63 0.61 0.61 0.006

Right 0.62a 0.62a 0.62a 0.61a,b 0.61a,b 0.61a,b 0.60b 0.006

Stance C Left 0.38a,b 0.37a 0.37a,b 0.37a,b 0.38a,b 0.38a,b 0.38b 0.006

(as a % of Right 0.38a,b 0.37a 0.37a 0.37a 0.38a,b 0.38a,b 0.39b 0.006

Stride R Left 0.37 0.37 0.37 0.38 0.36 0.37 0.37 0.006

Duration) Right 0.38a 0.37a,b 0.37b 0.38a,b 0.38a,b 0.39a 0.39a 0.006

S Left 0.37a,b 0.38a 0.38a,b 0.38a,b 0.37b 0.39a,b 0.39b 0.006

Right 0.38a 0.38a 0.38a 0.39a,b 0.39a,b 0.39a,b 0.40b 0.006

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 8. Treatment*Day lsmeans of Suspension Time at the Trot (seconds)

76 Gait Group Day 0 Day 2 Day 14 Day 28 Day 30 Day 42 Day 56 SEM

Trot Chiropractic 0.05 0.05 0.05 0.05 0.05 0.05 0.05 0.004

Riding 0.05 0.05 0.05 0.05 0.05 0.05 0.05 0.004

Sedentary 0.05 0.05 0.05 0.05 0.05 0.05 0.04 0.004

a,b,c Means within rows with different superscripts differ (P <

0.05)

Table 9. Treatment*Day lsmeans of Stride Length at the Walk and Trot (centimeters)

Day

Gait Group 0 2 14 28 30 42 56 SEM

Walk C 158.62 158.35 153.5 155.62 158.94 159.77 152.25 5.3143

R 157.64 159.25 155.54 155.99 154.16 158.37 154.13 5.3143

S 154.57 161.53 148.97 153.67 156.8 162.09 150.89 5.3143

Trot C 210.39 208 190.79 206.8 190.7 203.97 203.76 9.5834

R 207.36 211.14 200.14 212.31 198.54 208.95 210.36 9.5834

S 220.27a 209.03a,b 208.24a,b 178.9b 194.1a,b 206.5a,b 207.76a,b 9.5834

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 10. Treatment lsmeans of Cortisol on all days (nmol/L) Treatment Estimate SEM Chiropractic 84.5 5.51 Riding 78.7 5.56 Sedentary 74.6 5.56 PRE cortisol pulled immediately before treatment (0 Minutes), MID cortisol at midpoint of treatment (10 minutes), and POST cortisol pulled immediately post treatment (20 minutes)

77 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 11. Treatment lsmeans of AVG Heart Rate on all days (bpm) Treatment Estimate SEM Chiropractic 42 2.1 Riding 41 2.1 Sedentary 44 2.2

From Polar RS300x sa heart rate monitor set at ‘Other Sport 2’. Avg is the average heart rate for each treatment group. x,y,z Means within columns within variable differ (P < 0.05)

Table 12. Treatment lsmeans of MAX Heart Rate (bpm) Treatment Estimate SEM Chiropractic 70 5.4 Riding 67 5.2 Sedentary 73 5.5 From Polar RS300x sa heart rate monitor set at ‘Other Sport 2’. Max heart rate is the maximum heart rate for each treatment group. x,y,z Means within columns within variable differ (P < 0.05)

Table 13. Treatment lsmeans of MIN Heart Rate (bpm) Treatment Estimate SEM Chiropractic 30x 1.7 Riding 32x,y 1.7 Sedentary 35y 1.7 From Polar RS300x sa heart rate monitor set at ‘Other Sport 2’. Min heart rate is the minimum heart rate for each group. x,y,z Means within columns within variable differ (P < 0.05)

Table 14. Treatment*Day lsmeans of the Limb Measurements at the Walk

78 Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing Time C Front 0.41 0.40 0.40 0.40 0.40 0.40 0.40 0.008

(seconds) Hind 0.42 0.40 0.41 0.40 0.41 0.41 0.41 0.009

R Front 0.42 0.42 0.41 0.41 0.41 0.41 0.42 0.008

Hind 0.43 0.41 0.42 0.41 0.42 0.42 0.42 0.009

S Front 0.42 0.41 0.40 0.41 0.42 0.42 0.41 0.008

Hind 0.42 0.40 0.41 0.41 0.42 0.41 0.41 0.009

Stance Time C Front 0.71 0.71 0.71 0.70 0.69 0.70 0.70 0.02

(seconds) Hind 0.71 0.71 0.70 0.69 0.68 0.70 0.70 0.02

R Front 0.72 0.72 0.71 0.70 0.70 0.70 0.70 0.02

Hind 0.72 0.72 0.71 0.70 0.70 0.70 0.69 0.02

S Front 0.71 0.68 0.69 0.70 0.69 0.66 0.68 0.02

Hind 0.72 0.69 0.70 0.70 0.70 0.71 0.69 0.02

Stride C Front 1.11 1.11 1.11 1.1 1.09 1.1 1.1 0.02

Duration Hind 1.13 1.11 1.11 1.09 1.09 1.11 1.11 0.02

(seconds) R Front 1.14 1.13 1.12 1.11 1.12 1.11 1.11 0.02

Hind 1.15 1.13 1.13 1.11 1.12 1.13 1.11 0.02

S Front 1.12 1.09 1.1 1.1 1.11 1.08 1.09 0.02

Hind 1.14 1.09 1.11 1.11 1.12 1.12 1.1 0.02

Swing C Front 0.37 0.36 0.36 0.37 0.37 0.36 0.36 0.005

(as a % of Hind 0.37 0.36 0.37 0.37 0.37 0.37 0.37 0.003

stride R Front 0.37 0.36 0.36 0.37 0.37 0.37 0.37 0.005

duration) Hind 0.37ab 0.37a 0.37ab 0.37ab 0.38b 0.37ab 0.38ab 0.003

S Front 0.38 0.37 0.37 0.37 0.37 0.39 0.37 0.005

79 Hind 0.37 0.37 0.37 0.37 0.37 0.37 0.37 0.003

Stance C Front 0.65 0.64 0.64 0.63 0.63 0.64 0.64 0.005

(as a % of Hind 0.63 0.64 0.63 0.63 0.63 0.63 0.63 0.004

stride R Front 0.63 0.64 0.64 0.63 0.63 0.63 0.63 0.005

duration) Hind 0.63ab 0.63a 0.63ab 0.63ab 0.62b 0.63ab 0.62ab 0.004

S Front 0.65 0.63 0.63 0.63 0.63 0.63 0.63 0.005

Hind 0.63 0.63 0.63 0.63 0.63 0.63 0.63 0.004

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 15. Treatment*Day lsmeans of the Limb Measurements at the Trot

Day

Measurement Group Side 0 2 14 28 30 42 56 SEM

Swing Time C Front 0.37 0.37 0.36 0.35 0.35 0.35 0.35 0.01

(seconds) Hind 0.39 0.39 0.37 0.37 0.37 0.36 0.36 0.01

R Front 0.38 0.38 0.37 0.37 0.37 0.37 0.37 0.01

Hind 0.39 0.39 0.38 0.38 0.38 0.38 0.38 0.01

S Front 0.36 0.36 0.35 0.36 0.36 0.35 0.35 0.01

Hind 0.38 0.37 0.36 0.36 0.37 0.36 0.36 0.01

Stance Time C Front 0.24 0.24 0.23 0.23 0.23 0.23 0.24 0.008

(seconds) Hind 0.23 0.23 0.22 0.22 0.22 0.22 0.23 0.008

R Front 0.24 0.24 0.23 0.23 0.23 0.23 0.23 0.008

Hind 0.24 0.23 0.22 0.23 0.23 0.23 0.23 0.008

S Front 0.24 0.24 0.23 0.23 0.23 0.23 0.24 0.008

Hind 0.23 0.23 0.22 0.22 0.23 0.23 0.24 0.008

Stride C Front 0.62 0.61 0.59 0.58 0.59 0.59 0.6 0.02

Duration Hind 0.62 0.61 0.6 0.59 0.6 0.59 0.6 0.02

(seconds) R Front 0.63 0.62 0.6 0.6 0.6 0.6 0.6 0.02

Hind 0.63 0.62 0.6 0.61 0.61 0.61 0.61 0.02

S Front 0.6 0.6 0.58 0.59 0.59 0.58 0.59 0.02

Hind 0.61 0.6 0.59 0.59 0.6 0.59 0.6 0.02

Swing C Front 0.61 0.6 0.61 0.61 0.6 0.6 0.59 0.007

(as a % of Hind 0.62ab 0.63a 0.63a 0.63a 0.62ab 0.62ab 0.61b 0.005

stride R Front 0.61 0.61 0.61 0.62 0.62 0.61 0.61 0.007

duration) Hind 0.62 0.63 0.63 0.62 0.63 0.62 0.62 0.005

S Front 0.6 0.6 0.6 0.61 0.6 0.6 0.59 0.007

Hind 0.62a 0.62a 0.62ab 0.62ab 0.62a 0.61ab 0.6b 0.005

Stance C Front 0.39 0.40 0.39 0.39 0.40 0.40 0.40 0.007

(as a % of Hind 0.38ab 0.37a 0.37a 0.37a 0.38ab 0.38ab 0.39b 0.005

stride R Front 0.39 0.39 0.39 0.38 0.38 0.39 0.39 0.007

duration) Hind 0.38 0.37 0.37 0.38 0.37 0.38 0.38 0.005

S Front 0.40 0.40 0.40 0.39 0.40 0.40 0.42 0.007

Hind 0.38a 0.38a 0.38ab 0.38ab 0.38a 0.39ab 0.40b 0.005

C denotes Chiropractic Group, R denotes Riding group, S denotes Sedentary group Chiropractic performed on Day 1 and Day 29 a,b,c Means within rows with different superscripts differ (P < 0.05)

Table 16. Serum Cortisol by Horse and Group on Day 1 Name Treatment Day Nmol/L Day Nmol/L Day Nmol/L H1 S 1PRE 60.5 1MID N/A 1POST 67.5 H2 R 1PRE 50.8 1MID N/A 1POST 101.8 H3 C 1PRE 162 1MID 113.5 1POST 101.8 H4 R 1PRE 51 1MID 54.2 1POST 65.6 H5 S 1PRE 35.8 1MID 34.3 1POST 41 H6 S 1PRE 47 1MID 60.7 1POST 63 H7 C 1PRE 26.6 1MID 67.4 1POST 83.3 H8 C 1PRE 25.4 1MID 46 1POST 47.1 H9 C 1PRE 82.1 1MID 65 1POST 91.6 H10 R 1PRE 71.9 1MID 54.6 1POST 59.7 H11 C 1PRE 34.1 1MID 67.4 1POST 104.5 H12 S 1PRE 90.9 1MID 93 1POST 108.2 H13 R 1PRE 90.7 1MID 87.5 1POST 110.6 H14 C 1PRE 47.6 1MID 45.7 1POST 66.5 H15 R 1PRE 75.4 1MID 90.2 1POST 88.7 H16 S 1PRE 54.5 1MID 100.9 1POST 107.1 H17 R 1PRE 67.7 1MID 59 1POST 70.3 H18 S 1PRE 77 1MID 75.1 1POST 75.4 Shows the raw data for serum cortisol levels of each horse at PRE,MID, and POST treatment C denotes Chiropractic group, R denotes Riding group, S denotes Sedentary group

86 Table 17. Serum Cortisol by Horse and Group on Day 29 Name Treatment Day Nmol/L Day Nmol/L Day Nmol/L H1 S 29PRE 95.7 29MID 80 29POST 75.2 H2 R 29PRE 75.1 29MID 81.8 29POST 85.7 H3 C 29PRE 47.3 29MID 42.8 29POST 41.6 H4 R 29PRE 125.4 29MID 135.8 29POST 118 H5 S 29PRE 149.3 29MID 138.2 29POST 134.6 H6 S 29PRE 68.8 29MID 50.1 29POST 53 H7 C 29PRE 96.3 29MID 88.4 29POST 87.4 H8 C 29PRE 62.4 29MID 48.6 29POST 44.7 H9 C 29PRE 152.1 29MID 150 29POST 130 H10 R 29PRE 70.1 29MID 72.2 29POST 61.1 H11 C 29PRE 127.4 29MID 109 29POST 93.6 H12 S 29PRE 51.5 29MID 33.7 29POST 36.8 H13 R 29PRE 155.9 29MID 104.4 29POST 81.7 H14 C 29PRE 187.8 29MID 128.6 29POST 124.5 H15 R 29PRE 86.3 29MID 59.3 29POST 51.3 H16 S 29PRE 106.6 29MID 80.7 29POST 67.6 H17 R 29PRE 66.2 29MID 43.3 29POST 40.7 H18 S 29PRE 74.8 29MID 67.2 29POST 56.6 Shows the raw data for serum cortisol levels of each horse at PRE,MID, and POST treatment C denotes Chiropractic group, R denotes Riding group, S denotes Sedentary group