The Journal of Experimental Biology 201, 647–654 (1998) 647 Printed in Great Britain © The Company of Biologists Limited 1998 JEB1431 BLOOD VOLUME, PLASMA VOLUME AND CIRCULATION TIME IN A HIGH- ENERGY-DEMAND TELEOST, THE YELLOWFIN TUNA (THUNNUS ALBACARES) RICHARD W. BRILL*,1, KATHERINE L. COUSINS1, DAVID R. JONES1,2, PETER G. BUSHNELL1,3 AND JOHN F. STEFFENSEN1,4 1Pelagic Fisheries Research Program, Joint Institute for Marine and Atmospheric Research, School of Ocean and Earth Science and Technology, University of Hawaii, Honolulu, HI 96822, USA, 2Department of Zoology, University of British Columbia, 6270 University Boulevard, Vancouver, British Columbia, Canada V6T 2A9, 3Department of Biological Sciences, Indiana University South Bend, 1700 Mishawaka Avenue, South Bend, IN 46634-7111, USA and 4Marine Biological Laboratory, Copenhagen University, DK-3000 Helsingør, Denmark *e-mail: [email protected] Accepted 8 December 1997: published on WWW 5 February 1998 Summary We measured red cell space with 51Cr-labeled red blood of yellowfin tuna is small, and its exact volume is not cells, and dextran space with 500 kDa fluorescein- measurable by our methods. Although blood volume is not isothiocyanate-labeled dextran (FITC-dextran), in two exceptional, circulation time (blood volume/cardiac output) groups of yellowfin tuna (Thunnus albacares). Red cell is clearly shorter in yellowfin tuna than in other active − space was 13.8±0.7 ml kg 1 (mean ± S.E.M.) Assuming a teleosts. In a 1 kg yellowfin tuna, circulation time is whole-body hematocrit equal to the hematocrit measured approximately 0.4 min (47 ml kg−1/115 ml min−1 kg−1) at the ventral aortic sampling site and no significant compared with 1.3 min (46 ml kg−1/35 ml min−1 kg−1) in sequestering of 51Cr-labeled red blood cells by the spleen, yellowtail (Seriola quinqueradiata) and 1.9 min blood volume was 46.7±2.2 ml kg−1. This is within the range (35 ml kg−1/18 ml min−1 kg−1) in rainbow trout reported for most other teleosts (30–70 ml kg−1), but well (Oncorhynchus mykiss). In air-breathing vertebrates, high below that previously reported for albacore (Thunnus metabolic rates are necessarily correlated with short alalunga, 82–197 ml kg−1). Plasma volume within the circulation times. Our data are the first to imply that a primary circulatory system (calculated from the 51Cr- similar relationship occurs in fishes. labeled red blood cell data) was 32.9±2.3 ml kg−1. Dextran space was 37.0±3.7 ml kg−1. Because 500 kDa FITC-dextran Key words: fish, yellowfin tuna, Thunnus albacares, pelagic, appeared to remain within the vascular space, these data cardiorespiratory, metabolic rate, cardiac output, red cell space, 51Cr, imply that the volume of the secondary circulatory system dextran, primary circulatory system, secondary circulatory system. Introduction The relationships between metabolic rate and the respiratory, metabolic rates of tunas include large gill surface area, high circulatory and cellular mechanisms governing oxygen cardiac output, elevated hemoglobin concentrations and the transport from the respiratory medium to the tissues in air- ability to maintain muscle temperature significantly above breathing vertebrates have been the subject of intensive scrutiny ambient (Bushnell and Jones, 1994; Dickson, 1996). Other over the last two decades (e.g. Taylor et al. 1996 and the related adaptations that could potentially allow tunas to achieve high papers in the same volume). Many of the lessons learned from rates of oxygen consumption are either a significantly elevated these studies appear to be applicable to fishes (Mathieu-Costello blood volume (Korzjynew and Nikolskria, 1951) or a short et al. 1992, 1995, 1996; Moyes et al. 1992), although a circulation time (i.e. blood volume/cardiac output) (Coulson, complete set of supporting physiological and morphometric 1986). Using radio-iodinated bovine serum albumin, Laurs et data for a given fish species is often wanting. Regardless, it is al. (1978) reported blood volumes of albacore (Thunnus clear that tunas (family Scombridae, tribe Thunnini) are high- alalunga) ranging from 82 to 197 ml kg−1. In contrast, blood energy-demand teleosts and that both their standard and volumes for 25 species of elasmobranch, teleost and holostean maximum rates of oxygen consumption exceed those of other fishes listed by Tort et al. (1991) range from 18 to 80 ml kg−1, active teleosts (e.g. salmonids) by at least fourfold (Brill, 1987; although blood volumes of most teleosts lay within a more Boggs and Kitchell, 1991; Dewar and Graham, 1994; restricted range (30–70 ml kg−1, Itazawa et al. 1983; Olson, Korsmeyer et al. 1996a). Adaptations supporting the high 1992). In albacore, elevated blood volumes would extend 648 R. W. BRILL AND OTHERS circulation times even in the face of the elevated cardiac outputs approximately 20 h before use in an experiment to allow of tuna (Lai et al. 1987; White et al. 1988; Bushnell and Brill, sufficient time for gut clearance (Magnuson, 1969). 1992; Jones et al. 1993), implying that the former is more Anesthesia and surgical procedures were as described important than the latter for achieving exceptionally high previously (Bushnell and Brill, 1992). Briefly, fish were dip- metabolic rates. However, based on the dilution of iodocyanin netted from their holding tank and immediately placed in a green dye (‘cardio-green’), Bushnell (1988) estimated blood plastic bag containing 1 g l−1 of tricaine methanosulfonate volumes in skipjack (Katsuwonus pelamis) and yellowfin (MS222) buffered with an equal molar concentration of sodium (Thunnus albacares) tunas to be approximately 50 ml kg−1. If bicarbonate. After initial anesthesia, fish were rushed into the correct, the impact of these data on our concepts of circulation laboratory, placed ventral-side up in a soft chamois sling and time, oxygen and substrate delivery, and their relationship to ventilated with oxygenated sea water (temporarily chilled to metabolic rate is considerable because of the interrelationship 21–22 °C) containing 0.1 g l−1 of buffered tricaine between these variables (Coulson et al. 1977; Coulson and methanosulfonate. A 20 gauge, 3.2 cm Instyle Vialon Herbert, 1984; Coulson, 1986; Bushnell and Brill, 1992; intravenous catheter (Becton Dickinson Vascular Access, Korsmeyer et al. 1996a,b). Sandy, Utah, USA) was introduced into the ventral aorta under Olson (1992), in his excellent and comprehensive review on manometric guide and connected to a 20 cm length of the blood volumes of fishes, identified two trends. First, polyethylene tubing (PE 160). Fish were turned upright, a 20 Osteichthyes have the lowest blood volumes of any vertebrate gauge hypodermic needle was placed into the neural canal and, second, estimated blood volumes depend on the immediately posterior to the skull and used to inject 0.1–0.2 ml techniques employed. Larger blood volumes are usually doses of 4 % lidocaine. This procedure blocks spinal motor measured with plasma volume indicators (e.g. Evans Blue or nerves and prevents excessive tail movements, but leaves all radio-iodinated albumins) than are obtained with 51Cr-labeled cranial nerves and cardiorespiratory function intact (Bushnell red blood cells. Problems associated with dyes or protein and Brill, 1991). Fish were then placed in front of a pipe markers arise because they may not remain within the primary delivering approximately 30–35 l min−1 of oxygenated sea circulatory system, instead entering the secondary circulatory water and were thus able to set their own ventilation volume system (a parallel ‘lymph-like’ circulatory system in teleosts; by adjusting mouth gape. Throughout the duration of the Steffensen and Lomholt, 1992; Olson, 1996), the interstitial experiment, fish were kept sedated by repeated 0.1–0.3 ml space or even be excreted. The use of 51Cr-labeled red blood intramuscular injections of the steroid anesthetic Saffan cells can, however, also result in an overestimate of red cell (alphaxalone, Glaxovet, Harefield, Uxbridge, UK), a highly space and blood volume because labeled red blood cells may effective anesthetic in fish (Oswald, 1978). At the conclusion be selectively sequestered by the spleen (Duff et al. 1987). of an experiment, animals were killed with an overdose of Moreover, the accuracy of both techniques is dependent on sodium pentobarbital, injected via the indwelling ventral aortic measured hematocrit because these data are needed to calculate catheter, and weighed. Data were obtained from seven fish blood volume from the measured plasma volume or red cell using 51Cr-labeled red blood cells (body mass 0.750–2.420 kg) space (Jones, 1970; Fairbanks et al. 1996). It is generally and from seven fish using 500 kDa FITC-dextran (body mass assumed that whole-body hematocrit and hematocrit measured 0.865–3.325 kg). at the sampling site (usually a large blood vessel) are the same, although they can differ considerably (Albert, 1971; Gingerich Measurement of red cell space and Pityer, 1989; Olson, 1992; Fairbanks et al. 1996). Red blood cells labeled with 51Cr (Amersham Canada Consequently, we decided to quantify red cell space and Limited, Oakville, Ontario, Canada) were used to measure red plasma volume of yellowfin tuna using two independent cell space. To avoid any complications due to blood clotting, techniques. The former was measured directly using 51Cr- fish were given 1000 i.u. of sodium heparin (0.1 ml of labeled red blood cells and the latter by dilution of large 10 000 i.u. ml−1) via the ventral aortic catheter approximately (500 kDa) fluorescein-isothiocyanate-labeled dextran (FITC- 1 h after catheterization. Ten minutes later, 2 ml of blood was dextran). We chose 500 kDa FITC-dextran as a marker in an removed. The blood was mixed with 10 ml of tuna saline attempt specifically to reduce complications associated with (1.17 % NaCl), centrifuged (approximately 50 g for 2 min), and non-binding of a dye to plasma proteins and extravasation the supernatant discarded.