Hatchling painted turtles (Chrysemys picta) survive exposure to subzero temperatures during hibernation by avoiding freezing

Hatchling painted turtles (Chrysemys picta) were placed individually into artificial nests constructed in jars of damp soil and then were cooled slowly to temperatures between-7.7 and-12.7 °C. Distinct exotherms were recorded in all jars when water in the soil began to freeze at temperatures between-0.9 and-2.4 °C. A second (animal) exotherm was subsequently detected in some of the jars when water in hatchlings also began to freeze. An animal exotherm occurred in the temperature records for all 23 hatchlings that died in tests terminating at temperatures between-7.7 and-10.8 °C, but no such exotherm was apparent in the temperature records for the 23 turtles that survived these treatments. Moreover, the 4 hatchlings that produced exotherms in tests terminating between-11.5 and-12.7 °C failed to survive, but 5 of 7 hatchlings that produced no exotherm in these tests also died. Thus, turtles that die at subzero temperatures above-11 °C apparently succumb to freezing when ice propagates across their integument from the frozen soil, but animals that die at temperatures below-11 °C generally perish from some other cause. These findings indicate that hatchling painted turtles overwintering inside their shallow, subterranean nests survive exposure to subzero temperatures by avoiding freezing instead of by tolerating freezing.     

The relationship between gut contents and supercooling capacity in hatchling painted turtles (Chrysemys picta)

Painted turtles (Chrysemys picta) typically spend their first winter of life in a shallow, subterranean hibernaculum (the natal nest) where they seemingly withstand exposure to ice and cold by resisting freezing and becoming supercooled. However, turtles ingest soil and fragments of eggshell as they are hatching from their eggs, and the ingestate usually contains efficient nucleating agents that cause water to freeze at high subzero temperatures. Consequently, neonatal painted turtles have only a modest ability to undergo supercooling in the period immediately after hatching. We studied the limit for supercooling (SCP) in hatchlings that were acclimating to different thermal regimes and then related SCPs of the turtles to the amount of particulate matter in their gastrointestinal (GI) tract. Turtles that were transferred directly from 26 degrees C (the incubation temperature) to 2 degrees C did not purge soil from their gut, and SCPs for these animals remained near -4 degrees C for the 60 days of the study. Animals that were held at 26 degrees C for the duration of the experiment usually cleared soil from their GI tract within 24 days, but SCPs for these turtles were only slightly lower after 60 days than they were at the outset of the experiment. Hatchlings that were acclimating slowly to 2 degrees C cleared soil from their gut within 24 days and realized a modest reduction in their SCP. However, the limit of supercooling in the slowly acclimating animals continued to decline even after all particulate material had been removed from their GI tract, thereby indicating that factors intrinsic to the nucleating agents themselves also may have been involved in the acclimation of hatchlings to low temperature. The lowest SCPs for turtles that were acclimating slowly to 2 degrees C were similar to SCPs recorded in an earlier study of animals taken from natural nests in late autumn, so the current findings affirm the importance of seasonally declining temperatures in preparing animals in the field to withstand conditions that they will encounter during winter.     

Influence of the nest environment on bone mineral content in hatchling painted turtles (Chrysemys picta)

We performed an experiment at a field site in north-central Nebraska to assess the role of the nest environment in inducing variation in bone mineral content in hatchling painted turtles Chrysemys picta (Schneider 1783). The contents of several newly constructed nests were manipulated by reciprocal transplant, after which the eggs were allowed to incubate for 8 wk under natural conditions. The nests were then excavated, and the eggs were brought into the laboratory to complete incubation and hatch under standard conditions of temperature and moisture. The hatchlings were killed, and their carcasses and residual yolks were analyzed separately for calcium and phosphorus. More of the random variation in carcass calcium and phosphorus was related to the nest in which eggs incubated (37% and 42%, respectively) than was associated with the clutch of origin (21% and 37%). Moreover, hatchlings from some nests contained substantially more calcium and phosphorus than did hatchlings from other nests, both in terms of the absolute amounts of the elements in their carcasses (pointing to variation in body size) and in terms of the concentrations of those elements (pointing to variation in bone density). The amounts of calcium and phosphorus in carcasses of hatchlings were positively correlated with changes in mass of their eggs during the 8 wk that the eggs incubated in nests in the field, thereby indicating that the influence of the nest environment on developing embryos probably was mediated by water exchanges experienced by the eggs. These findings indicate that developmental plasticity underlies a major fraction of the variation in mineral content of hatchling painted turtles emerging from nests in the field. Phenotypic variation attributable to plasticity consequently needs to be addressed in models for life-history evolution of painted turtles and other chelonians producing eggs with soft, flexible shells.     

Effects of temperature and moisture during incubation on carcass composition of hatchling snapping turtles (Chelydra serpentina)

Summary Flexible-shelled eggs of common snapping turtles (Chelydra serpentina) were incubated on each of two substrates (vermiculite, sand) at each of three temperatures (26.0°C, 28.5°C, 31.0°C) and three moisture regimes (wet, intermediate, dry). Embryos developing in cool, wet environments mobilized the largest amounts of protein from their yolk and attained the largest size before hatching, whereas turtles developing in warm, dry environments mobilized the smallest quantities of protein and were the smallest in body size at hatching. Embryos on wet substrates mobilized more lipid from their yolk than did embryos on dry media, but ambient temperature had no demonstrable influence on patterns of lipid mobilization. The total reserve of neutral lipid available in residual yolk plus carcass to sustain neonates in the interval prior to the beginning of feeding was largest in hatchlings from dry environments and smallest in animals from wet environments, but was unaffected by temperature during incubation. Hydration of tissues in hatchlings was higher when incubation was in cool, moist conditions than when incubation was in warm, dry settings, thereby indicating that some of the effects of moisture and temperature on mobilization of nutrients by embryos may be mediated by differences in intracellular water. Patterns of response to temperature and moisture recorded for turtles emerging from eggs on sand were similar to those recorded for hatchlings on vermiculite, so no important conclusion would have been affected by incubating eggs on one medium instead of the other.     

Temperature during embryonic and juvenile development influences growth in hatchling snapping turtles,Chelydra serpentina

Embryonic temperature influenced subsequent growth in juvenile snapping turtles, Chelydra serpentina: incubation temperatures of 24 and 26.5°C enhanced growth relative to a temperature of 29°C. Although embryonic temperature normally determines gonadal sex in this species, experimental manipulations revealed that temperature effects on growth were independent of sex. Ambient temperature also affected growth: juvenile turtles grew slowly in a cool (19°C) versus a warm (28°C) environment. In a parallel experiment, turtles from different embryonic temperatures displayed different patterns of temperature choice in response to nutritional status or time of day. We tentatively conclude that embryonic temperature has both direct and indirect (i.e., through temperature choice) effects on growth in snapping turtles.     

Aerial and aquatic oxygen uptake by freely-diving snapping turtles (Chelydra serpentina)

Summary Aerial oxygen consumption of unrestrained, freely-diving warm-and cold-acclimated snapping turtles, Chelydra serpentina, was measured at 10, 20, and 30°C. Also, simultaneous determinations of aerial and aquatic oxygen uptake by voluntarilydiving animals were made at 4 and 20°C. The standard rates of aerial oxygen consumption are equivalent in cold-and warm-acclimated animals in water and in cold-acclimated ones in air; these rates are all lower than those of warm-acclimated animals in air. Thus either cold acclimation or voluntary submergence reduces the standard metabolic rate of snapping turtles but the effects are not additive. Aquatic oxygen uptake during voluntary submergence is more important at low than at moderate temperatures and probably contributes significantly to gas exchange in these animals as they overwinter beneath the ice of ponds and streams.     

Physiological responses to supercooling and hypoxia in the hatchling painted turtle, Chrysemys picta

We investigated physiological responses to supercooling in hatchling painted turtles (Chrysemys picta) which remain in their natal nests over winter and therefore may become exposed to subzero temperatures. These turtles are freeze tolerant but also must rely on supercooling to survive exposure to the lower temperatures occurring in nests during winter. We compared whole-body concentrations of lactate, glucose, glycerol, and ATP in turtles chilled at 0 degrees C, -4 degrees C, or -6 degrees C for 5 days, or at 6 degrees C for 19 days. In a companion experiment, we measured metabolite concentrations in turtles exposed to a hypoxic environment for 1 day, 4 days, or 8 days. Supercooling and hypoxia exposure were both associated with an increase in concentrations of lactate and glucose and a decrease in glycerol concentrations (albeit no change in the ATP pool), suggesting that supercooling induces functional hypoxia. We conclude that hypoxia tolerance may be an important pre-adaptation for surviving exposure to subzero temperatures in hatchling C. picta.     

Cardiovascular correlates of exercise in snapping turtles, Chelydra serpentina

1. Heart rate increased with a rise in body temperature (10-30 degrees C) and with induced physical exercise in snapping turtles. 2. Maximum heart rate increment occurred at 30 degrees C. 3. Standard oxygen pulse did not change with a rise in temperature. 4. Oxygen pulse during exercise and oxygen pulse increment were maximal at 10 degrees C and minimal at 20 degrees C. 5. The increase in heart rate with exercise accounted for only 9-22% of the increase in oxygen transport during activity; the remainder was provided by a rise in cardiac stroke volume and/or A-V difference.     

The physiology of hibernation among painted turtles: the Eastern painted turtle Chrysemys picta picta.

Eastern painted turtles (Chrysemys picta picta) from Connecticut were submerged at 3 degrees C in normoxic and anoxic water to simulate potential respiratory environments within their hibernacula. Those in normoxic water could survive submergence for at least 150 d, while those in anoxic water could survive for a maximum of about 125 d. Turtles in normoxic water developed a slight metabolic acidosis as plasma lactate accumulated to about 50 mM in 150 d, while anoxic turtles developed a severe lactic acidosis as plasma lactate reached about 200 mM in 125 d; there was no respiratory acidosis in either group. Plasma [Na+] changed little in either group, [Cl-] fell by about one-third in both, and [K+] increased by about fourfold in anoxic turtles but only slightly in those in normoxic water. Total plasma magnesium and calcium increased profoundly in anoxic turtles but moderately in those in normoxic water. Consideration of charge balance indicates that all major ions were measured in both groups. Plasma glucose remained unchanged in anoxic turtles until after about 75 d of submergence, when it increased and continued to increase with the duration of anoxia, with much variation among individuals; glucose remained unchanged throughout in turtles in normoxic water. Hematocrit doubled in 150 d in turtles in normoxic water; in anoxic turtles, an initial increase was no longer significant by day 100. Plasma osmolality increased markedly in anoxic turtles, largely because of accumulation of lactate, but anoxic turtles only gained about half the mass of turtles in normoxic water, who showed no increase in osmolality. The higher weight gain in the latter group is attributed to selective perfusion and ventilation of extrapulmonary gas exchange surfaces, resulting in a greater osmotic influx of water. The physiologic responses to simulated hibernation of C. picta picta are intermediate between those of Chrysemys picta bellii and Chrysemys picta dorsalis, which correlates with the severity of the winter each subspecies would be expected to encounter.     

Environmental modulation of calcium and phosphorus metabolism in embryonic snapping turtles (Chelydra serpentina)

Summary Eggs of common snapping turtles (Chelydra serpentina) were incubated on wet (–150 kPa water potential) and dry (–950 kPa) substrates in a laboratory study assessing the effects of the hydric environment on patterns of mobilization of calcium and phosphorus by developing embryos. We found that embryos developing in wet environments withdrew nutrients from their yolk faster, grew more rapidly, and incubated longer than embryos exposed to dry environments. Turtles developing in both environments absorbed calcium from the yolk at similar rates and depleted the yolk of almost its entire reserve of calcium prior to hatching. Calcium withdrawn from the yolk was supplemented with calcium mobilized from the eggshell, but embryos in wet environments obtained substantially more calcium from the eggshell than did those in dry settings. Embryos obtained all of the phosphorus used in skeletogenesis from the yolk, but those incubating in wet environments mobilized phosphorus from this compartment more rapidly than did those in dry settings. Exposing embryonic snapping turtles to wet environments apparently allows them to make more efficient use of the transitory source of calcium in the eggshell than is possible in dry environments. However, the residual yolk in hatchlings from both wet and dry environments contains too little calcium to support the growth of hard and soft tissues in neonates at rates similar to those characterizing the growth phase of development in embryogenesis.     

Avenues of extrapulmonary oxygen uptake in western painted turtles (Chrysemys picta belli) at 10 degrees C

The major avenues of extrapulmonary oxygen uptake were determined on submerged western painted turtles (Chrysemys picta bellii) at 10 degrees C by selectively blocking one or more potential pathways for exchange. Previous work indicated that the skin, the cloaca, and the buccopharyngeal cavity can all contribute significantly in various species of turtles. O(2) uptake was calculated from the rate of fall in water P(O(2)) in a closed chamber. Two series of experiments were conducted: in Series 1, each of the potential avenues was mechanically blocked either singly or in combination; in Series 2, active cloacal and buccal pumping were prevented pharmacologically using the paralytic agent rocuronium. In addition in Series 2, N(2)-breathing preceded submergence in some animals and in one set of Series 2 experiments arterial blood was sampled and analyzed for pH, lactate, P(O(2)), and P(CO(2)). Results in both Series 1 and Series 2 revealed that prevention of cloacal and/or buccopharyngeal exchange did not significantly affect total O(2) uptake. Interfering with skin diffusion in Series 1, however, significantly reduced O(2) uptake by 50%. N(2)-breathing prior to submergence in Series 2 did not affect O(2) uptake in paralyzed turtles but significantly increased uptake in unparalyzed turtles without catheters. Blood analysis revealed that all submerged turtles developed lactic acidosis, but the rate of rise in lactate was significantly lower in paralyzed animals. We conclude that passive diffusion through the integument is the principal avenue of aquatic O(2) uptake in this species.     

Availability of water affects organ growth in prenatal and neonatal snapping turtles (Chelydra serpentina)

We manipulated the amount of water that was available to prenatal and neonatal snapping turtles (Chelydra serpentina) in order to assess the impact of water on growth by different organs in these animals. Three treatments were used: (1) turtles that completed their incubation on a wet substrate, (2) turtles that completed their incubation on a dry substrate, and (3) turtles that spent a few days in water after completing incubation on a dry substrate. Turtles hatching on a dry substrate (treatment 2) were smaller than animals in the other two treatments (which did not differ in size), so data for mass of different organs were adjusted by ANCOVA to remove effects of body size. Scaled masses of liver, stomach, lungs, kidneys, and small intestine did not differ between turtles emerging in wet environments and those hatching in dry environments, but hearts of turtles hatching in dry settings were substantially larger than those of animals hatching in wet ones. Thus, the mass of most organs in turtles developing in wet and dry environments scaled to body size, whereas the heart was hypertrophied in embryos developing in dry environments. Turtles that spent a few days in water after hatching from eggs in dry environments grew rapidly in size, and the increase in body size was accompanied by disproportionately rapid growth in the liver, stomach, lungs, kidneys, and small intestine. The heart did not increase in size during this period, despite the substantial increase in body mass over that at hatching. The enlarged heart of turtles hatching on dry substrates may have been caused by a circulatory hypovolemia late in incubation; the rapid growth of organs other than the heart when these animals were placed in water may reflect a release from constraints on growth once circulatory volume was restored. Accepted: 2 November 1999     

Accumulation of lactate by frozen painted turtles (Chrysemys picta) and its relationship to freeze tolerance

Hatchling painted turtles (Chrysemys picta) survived freezing at -2 degrees C for 4 d, few recovered from freezing lasting 6 d, and none survived being frozen for 8 d. Whole-body glucose and lactate were low in animals that had not been subjected to cold and ice but increased precipitously in animals that were frozen for 2 d. Both metabolites continued to increase, but at a somewhat lower rate, in animals frozen for 4, 6, or 8 d. The increase in whole-body lactate reflects a reliance by frozen hatchlings on anaerobiosis, whereas the increase in glucose presumably results from mobilization of glycogen reserves to support anaerobic metabolism. Mortality of frozen hatchlings is correlated with the increase in whole-body lactate. Factors that may contribute to the observed correlation include a compromised capacity for individual organs to cope with the lactic acidosis that accompanies anaerobic metabolism and organ-specific depletion of energy reserves. Individual organs must rely on buffering and glucose reserves available in situ because blood of frozen hatchlings does not circulate. Thus, buffer from the shell cannot be transported to other organs, lactate cannot be sequestered in the shell, and glucose mobilized from liver glycogen is not available to supplement glucose reserves of other tissues. This integrated suite of physiological disruptions may limit tolerance of freezing to conditions with little or no ecological relevance.     

Herpesvirus-like infection in a painted turtle (Chrysemys picta)

A painted turtle (Chrysemys picta) which died in captivity had marked necrosis in the liver and lungs with numerous intranuclear inclusion bodies in hepatocytes and respiratory epithelial cells. Electron microscopy revealed herpesvirus-like particles in cells in affected tissues.     

Wetness of the nest environment influences cardiac development in pre- and post-natal snapping turtles (Chelydra serpentina)

We dissected hearts from near-term embryos and hatchlings of common snapping turtles (Chelydridae: Chelydra serpentina) whose eggs had incubated on wet or dry substrates, and then dried and individually weighed the heart and yolk-free carcass from each animal. Hearts and carcasses of prenatal and neonatal animals grew at different rates, and the patterns of growth by both heart and carcass differed between wet and dry environments. Hearts grew faster, both in actual mass and in mass adjusted for variation in body size, in embryos and hatchlings whose eggs were incubated on dry substrates than in animals whose eggs were held on wet media. This finding is consistent with a hypothesis that embryos incubating in dry settings experience hypovolemia secondary to dehydration and that enlargement of the heart compensates, in part, for the associated increase in viscosity of the blood. Embryonic turtles seemingly exhibit the same plasticity and response that would be expected from other vertebrate ectotherms subjected to the physiological challenges associated with desiccation and an associated reduction in blood volume.     

Ultrastructural morphology of the shell and shell membrane of eggs of common snapping turtles (Chelydra serpentina)

Common snapping turtles (Chelydra serpentina) lay nearly spherical, flexible-shelled eggs having an outer mineral layer composed of calcium carbonate in the aragonite form. The mineral layer is arranged into loosely organized groups of nodular shell units, with numerous spaces (or pores) between adjacent shell units. Shell units are structurally complex, consisting of an inner tip that is morphologically distinct from the main body of the shell unit. Contained within an intact shell unit at the interface of the tip and the main part of the shell unit is the central plaque, an apparent modification of the shell membrane that may serve to nucleate calcification of shell units during shell formation. The tips of shell units are firmly attached to a single, multilayered shell membrane throughout much of incubation. The calcareous layer begins to detach from the shell membrane about half-way through incubation, and changes in shell morphology attending this detachment indicate that snapping turtles may use the shell as a source of calcium during embryogenesis. The arrangement of the mineral layer into groups of shell units, the large number of spaces between shell units, and little or no interlocking of crystallites of adjacent shell units apparently are factors contributing to the ability of these eggs to swell as they absorb water.     

Metabolic rate of feeding and fasting juvenile midland painted turtles, Chrysemys picta marginata

1. Resting metabolic rates at 25 degrees C were determined for juvenile midland painted turtles that had recently been fed or fasted for 1, 2, 4, 6, 10, 14 or 19 days. 2. Recently fed turtles had an oxygen consumption rate of 211 microliter O2/g/hr. This decreased by 32% on the first day of the fast and by 69% by the 19th day. 3. Mass of the turtles (4.91-14.30 g) did not affect the rate of oxygen consumption (VO2).