FAU Institutional Repository http://purl.fcla.edu/fau/fauir This paper was submitted by the faculty of FAU’s Harbor Branch Oceanographic Institute. Notice: ©1990 Elsevier Inc. The final published version of this manuscript is available at http://www.sciencedirect.com/science/journal/03009629 and may be cited as: Peterson, M. S. (1990). Hypoxia-induced physiological changes in two mangrove swamp fishes: sheepshead minnow, Cyprinodon variegatus Lacepede and sailfin molly, Poecilia latipinna (Lesueur). Comparative Biochemistry and Physiology Part A: Physiology, 97(1), 17-21.doi:10.1016/0300-9629(90)90715-5 let 1) Compo Biochem. Physiol. Vol. 97A, No. I, pp. 17-21, 1990 0300-9629/90 $3.00 +0.00 Printedin Great Britain © 1990 Pergamon Press pic HYPOXIA-INDUCED PHYSIOLOGICAL CHANGES IN TWO MANGROVE SWAMP FISHES: SHEEPSHEAD MINNOW, CYPRINODON VARIEGATUS LACEPEDE AND SAILFIN MOLLY, POECILIA LATIPINNA (LESUEUR) MARK S. PETERSON* Harbor Branch Oceanographic Institution, Inc. Division of Marine Science 5600 Old Dixie Highway Ft. Pierce, FL 34946, USA. Telephone: (601) 325-3120 (Received 19 December 1989) Abstract-I. Laboratory measurements (30°C and 300/00 salinity) were made of plasma osmolality, plasma chloride ion concentration, hematocrit, oxygen consumption and survival of sheepshead minnow, • Cyprinodon oariegatus Lacepede and sailfin molly, Poecilia latipinna (Lesueur) under normoxic (150 mm Hg) and hypoxic (40 mm Hg) conditions. 2. Significant increases in hematocrit and reductions in oxygen consumption were documented for both species. Plasma osmolality increased in sheepshead minnows while in hypoxic conditions but plasma " chloride did not change from values in 150mm Hg in either species. There was no mortality in either species during the 24 hr hypoxia survival tests. 3. Results suggest a strong tolerance of hypoxia in both species and use of aquatic surface respiration (ASR) by P. latipinna. 4. Low-level mortality occurs in both species but severe mortality occurs only in C. tariegatus and may be due to synergistic environmental effects typical of mangrove swamp habitats. INTRODUCTION molality, plasma chloride ion concentration and oxy­ Mangrove swamps provide critical habitats for nu­ gen consumption. Finally, I investigated survival in merous resident and transient fishes (Odum et al., low-oxygen tensions in these two resident species. 1982; Thayer et aI., 1987). These habitats, however, are being modified for mosquito control or waterfowl MATERIALS AND METHODS management in a number of southeastern USA estu­ Field collections and general laboratory protocol aries (Whitman and Meredith, 1987) with resulting Fishes were collected from impounded mangrove swamps water quality deterioration and, at least in some in the Indian River Lagoon, Florida, USA. They were areas, a decrease in fish species richness (Harrington transported to the laboratory in styrofoam coolers contain­ and Harrington, 1982; Gilmore et al., 1982). The ing impoundment water where they were held at 25°C species that use these habitats must be able to endure overnight under high aeration or transferred directly into fluctuating and stressful environmental conditions for outdoor concrete vaults (900 I) when environmental temper­ extended periods of time. For example, sheepshead atures approached experimental temperatures. Experimen­ minnow and sailfin molly have been reported in tal animals were then transferred to 761 aquaria equipped hypersaline habitats (Gilmore et al., 1982) but are with individual filters, aerators and heaters. They were held good osmoregulators (Gustafson, 1981; Nordlie, in 30 ± 10/00 salinity and 30 ± 1°C under a 12L: 12D photo­ 1987). These two species and the mosquitofish, Gam­ period centered at 1230 hr for at least 7 days. Experimental animals were fed Tetramin flake food ad libitum twice daily busia affinis are also able to tolerate low-oxygen but were fasted for 24 hr prior to testing (except the survival tensions (Cech et al., 1985) while mosquitofish, other experiments). In all experiments, sex of the fishes was not mollies (Poulin et al., 1987), and striped mullet, Mugil considered. Experimental salinities were produced using cephalus (Moore, 1976) can behaviorally adjust their filtered (5 /lm) Atlantic Ocean seawater diluted with aged oxygen uptake by aquatic surface respiration (ASR). reverse osmosis water. Salinities were checked daily. The comparative eco-physiology of sheepshead minnow, Cyprinodon variegatus, and sailfin molly, Normoxic lhypoxic blood constituents experimental protocol Poecilia latipinna, from impounded mangrove Fish were netted from their experimental aquaria and swamps is unknown. The objective of this study was immediately measured to the nearest mm standard length to compare and contrast physiological changes be­ (SL). All blood samples were obtained by first blotting each tween two closely related resident species that differ individual dry and severing its caudal fin. The incision was immediately blotted and blood from the caudal artery was in their use of habitat. To do this, I documented drawn into a heparinized micro-capillary tube and cen­ hypoxia-induced changes in hematocrit, plasma os- trifuged for 4 min at 13,460g in an International Micro­ capillary Centrifuge (Model MB) for hematocrit (%) determination. All fish were processed within a 30 min *Present address: Department of Biological Sciences, PO period and individual blood collection was completed within Drawer GY, Mississippi State University, Mississippi I min to reduce handling effects on blood constituents State, MS 39762, USA. (Robertson et al., 1987). All individuals were killed between 17 COP 97A/J-8 18 MARK S. PETERSON 0800 and 0900 hr (Peterson and Gilmore, 1988). Plasma monitored at I, 2, 4, 6, 8, 10, 12 and 24 hr intervals. Death osmolality (rrrOsrn/kgj was determined on a 10 III sample was established when the fish did not move its operculum with a Wescor Vapor Pressure Osmometer (Model 5500). for I min. Throughout this portion of the experiments, Plasma chloride ion concentration (meq/l) was determined water flow rates (121.1 ± 9.0 ml/min) flushed the 0.851 from a 10111 sample on a Buchler Digital Chloridometer tubes, on average, every 7.02 min. Sheepshead minnows (Model 4-2500). Sheepshead minnows (n = 50) were held (n = 5) were held in the experimental conditions for under the above-mentioned conditions for 11.5 ± 0.5 days 19.0 ± 1.0 days and sailfin molIies (n = 5) for 31.0 ± 2.1 and sailfin molIies (n = 30) for 7.0 days prior to experimen­ days. tation. Hypoxic conditions were produced by bubbling nitrogen gas directly into the aquarium for 3.0 hr. thus Statistical treatments gradualIy reducing the oxygen tension to hypoxic conditions Fish size, osmolality and chloride ion concentration (all (38.5 ± 2.6 mm Hg; about 26% saturation; 1.6 ppm). Fish log 10 transformed) and hematocrits (arcsine transformed) were kept at the new Po, tension for 2.5 hr. Experimental Po, were analysed by Po, treatment by analysis of variance tensions were determined and monitored by injecting a (ANOVA). Oxygen consumption rates were analysed by a single water sample into a calibrated Radiometer PHM­ paired t-test (alpha = 0.05) whereas oxygen consumption 73/D616/E5046 oxygen analyzer system. The experimental rates vs wet weight were examined by linear regresion. [j is aquaria contained floating styrofoam slabs in order to the mean difference between paired values and only applies reduce surface breathing by sailfin molIies. to the paired t-test. Oxygen consumption rates RESULTS SmalI flow-through respirometers (modified 125 ml Erlen­ meyer flasks) equipped with two glass tubes, one that Normoxic /hypoxic blood constituents alIowed inflowing water to enter near the bottom of the flask while the other tube alIowed water to pass out of the flask Individual sheepshead minnow sizes ranged from near the bottom of the stopper (top of flask) were used. 24 to 40 and 31 to 45 mm SL for the normoxic and •I Fifteen respirometers were connected by vinyl tubing to a hypoxic experiments, respectively. Sailfin molly PVC manifold that was equipped with nylon valves for ranged from 36 to 54 and 32 to 39 mm SL. There were controlIing water flow rates (26.7 ± 12.4ml/min for min­ no significant size differences between the normoxic nows; 21.6± 8.7 for molIies). Due to flow rates and and hypoxic individuals (ANOY A; P > 0.05). respirometer size, steady state was achieved rapidly (Propp Significantly (ANOY A; P < 0.05) elevated hem­ et al., 1982). Water ofthe desired oxygen tension entered the manifold from a 456 I headbox via an acrylic chamber atocrits were documented for both species under (300 m!) which was used to obtain an initial water sample hypoxic conditions (Fig. I). Only sheepshead min­ with a needle and syringe. Final water samples were taken nows exhibited significantly (ANOYA; P < 0.01) with a needle and syringe from a piece of vinyl tubing elevated plasma osmolality, whereas there were no attached to the glass tube leaving each respirometer. differences in plasma chloride ion concentration for Desired oxygen tensions (mm Hg) were produced by either species (P < 0.05) (Fig. I). passing water from a headbox through an in-line oxygen stripper (Cameron, 1986) made of PVC (152.4 em x 7.94 em Oxygen consumption rates i.d.) and filIed with marbles to increase surface area for nitrogern gas diffusion. Regulation of the counter flows For the size range examined, there was no statisti­ brought about the desired oxygen levels. cally significant relationship between wet weight and Routine oxygen consumption rates (mg 02/g/hr) were oxygen consumption rates of sheepshead minnow calculated by the equation V02 = (PO,i - Po,,) (a) (1.428) (1.63-3.21 g ww) in either normoxic or hypoxic con­ V/ww (g) (Lampert, 1984), where (Po,,) = oxygen tension of ditions (ANOY A; P > 0.05). However, sailfin molly inhalent water; (Po,,) = oxygen tension of exhalent water; (0.75-1.84 g ww) exhibited significant (P < 0.05) a = solubility coefficient (from Cameron, 1986); 1.428 con­ weight-dependent respiration in normoxic but not in verts rnl/l to mg/l oxygen; V = flow rate (ml/hr); and hypoxic conditions (P > 0.05).