Body temperatures of over-wintering Cottonmouth snakes: hibernaculum use and inter-individual variation.

ABSTRACT

Environmental temperatures affect the physiology, ecology and behavior of reptiles due to the temperature dependence of biochemical reactions and the inability of reptiles to metabolically maintain constant temperatures. Consequently, periods during which environmental temperatures are extreme or variable (e.g. winter) may be particularly stressful for reptiles. However, few studies have reported body temperatures of reptiles during the winter. We studied body temperatures of Cottonmouth snakes (Agkistrodon piscivorus) during the winter in eastern central Alabama. We compared Cottonmouth body temperatures that were measured using implanted temperature-sensitive radio transmitters to air, soil and water temperatures to determine whether snakes conformed to environmental temperatures. We also compared body temperatures among snakes to determine whether individual snakes differed in their body temperatures. Results indicate that snakes maintained mean body temperatures near the upper range of environmental temperatures. Additionally, individuals maintained different body temperatures during over-wintering. These results provide evidence that snakes select hibernacula (the sites where the snakes spend the winter) or employ thermoregulatory behaviors to maintain relatively warm winter body temperatures and that different individuals experience different body temperatures during the winter.

INTRODUCTION

Environmental temperature profoundly affects the physiology, ecology and behavior of many reptiles (Gibson and Falls 1979, Plummer 1993, Beaupre 1995, Luiselli and Akani 2002, Herczeg et al. 2008, Kapfer et al. 2008). This is due to the temperature-dependence of biochemical reactions and the inability of reptiles to metabolically maintain constant body temperatures as environmental temperatures change (Seebacher and Franklin 2005). Body temperatures affect the rates of diverse physiological processes including digestion (Wang et al., 2003), neural activity (Teng and Wilkinson, 2003), metabolism (Dorcas et al. 2004), immune function (Cooper et al. 1985), and growth and development (O'Donnell and Arnold 2005). Consequently, reptiles must maintain body temperatures within a physiologically determined range in order to maintain homeostasis. During parts of the year when environmental temperatures are mild, animals maintain stable body temperatures through behavioral thermoregulation-moving between microhabitats to achieve body temperatures within a desired range (Hertz et al., 1993). However, thermoregulation is much more difficult during periods of the year when environmental temperatures are extreme and variable (e.g. winter for temperate species, Blouin-Demers and Weatherhead 2002). During the winter, reptiles are faced with three options: conform to environmental temperatures, attempt behavioral thermoregulation, or use hibernacula (wintering sites) that offer favorable winter temperatures.

Many studies have reported body temperatures of reptiles during parts of the year when they are active (i.e. warm spring and summer months for temperate species, Plummer 1993, Blouin-Demers and Weatherhead 2002, Crane and Greene 2008, Herczeg et al. 2008, Kapfer et al. 2008). Few studies, however, have reported body temperatures of reptiles during inactive periods when environmental conditions are harsh and variable (e.g., during the winter for temperate species, Blouin-Demers and Weatherhead 2001, Gregory 1982, Himes et al., 2006) despite evidence suggesting that winter body temperatures may be directly linked to over-wintering survival. Low body temperatures during over-wintering can cause animals to freeze to death (Blem and Blem, 1995, Shine and Mason 2004). High temperatures, on the other hand, elevate metabolic rates and may cause animals to exhaust energy stores and the of starvation (Blem and Blem 1995, Blem 1997). The paucity of published data on over-wintering body temperatures of reptiles makes it difficult to determine the importance of this period in determining their physiology, behavior and survival.

In order to more fully characterize winter body temperatures of reptiles and evaluate the possibilities of winter thermoregulation and hibernaculum selection, we studied over-wintering body temperatures of free-ranging Cottonmouth snakes, Agkistrodon piscivorus (Lacepede). This species has served as a model in numerous studies of reptilian physiology, behavior and thermal biology (e.g. Blem and Blem 1995, Blem 1997, Crane and Greene 2008, Glaudas et al. 2007, Hill 2004, Hill and Beaupre 2008, McCue and Lillywhite 2002, Wharton 1969); however, to our knowledge, no study has reported over-wintering body temperatures of Cottonmouths. Specifically, we sought to address two questions about over-wintering Cottonmouths at our study site: 1) Do Cottonmouths (a) conform to dominant environmental temperatures during the winter, (b) actively thermoregulate or (c) use hibernacula with thermal properties that differ from the dominant environmental media (air, water, soil)? 2) Do different individuals experience different body temperatures during the winter? Since air, water and soil temperatures at our study site frequently drop below zero degrees Celsius during winter, we predicted that Cottonmouths would actively thermoregulate or use warmer microhabitats to avoid lethal body temperatures. Additionally, we were interested in inter-individual differences in body temperature. Such differences could point to differences in physiological states of individuals during this particularly harsh time in their life history (Gregory 1982). For example, body temperature differences among individuals can cause differences in the energetic cost of over-wintering because metabolic rates increase as a function of increasing temperature (McCue and Lillywhite 2002). The ability to maintain favorable body temperatures during the winter may be an important trait linked to over-wintering survival.

We addressed our questions by measuring field body temperatures of Cottonmouths from a population in eastern Alabama during the late-fall, winter, and early spring of 2005-2006. To test our prediction that snakes actively thermoregulate or use thermally favorable hibernacula rather than conforming to dominant environmental temperatures, we compared snake body temperatures (measured using temperature-sensitive radio telemetry) to air, water and soil temperatures measured with a quick-read thermometer at the site where each snake was located. To determine whether among-individual variation in body temperature existed in our study population, we compared mean body temperatures among snakes.

MATERIALS AND METHODS

We conducted this study from September 2005 to April 2006 in Tuskegee National Forest, Macon County, Alabama. This site (32[degrees]25.57'N, 85[degrees]38.39'W) consists of a series of small ephemeral ponds and larger permanent ponds that contain tree islands ranging in size from ~l[m.sup.2] to ~1000[m.sup.2]. The water table is between 10cm and 100cm below ground level on all islands, and many islands have extensive networks of passages below the soil surface that are commonly used by snakes. We excavated several burrows to determine their internal structure. Burrows consisted of one or more openings and a network of horizontal passageways that was typically located 8-15cm below the soil surface. Burrow passages were lined with a mixture of sand, tree roots, and decaying organic material and passages varied between approximately 5 cm and 25 cm in diameter.

Capture and Implantation

We recorded body temperatures of nine adult Cottonmouth snakes from 18 October 2005 to 6 April 2006. We gathered these data using implanted temperature-sensitive radio transmitters (Holohil, Ontario, Canada, model SB-2T). Transmitters allowed us to locate snakes in the field and measure body temperature at the time of location. From 15 September to 7 October of 2005 we captured nine adult Cottonmouths and brought them into the lab at Auburn University. On 10 October, we implanted a transmitter into the peritoneal cavity of each snake using the method of Reinert and Cundall (1982). We anesthetized snakes using isoflourane (Sigma) prior to surgery (mass of transmitter less than 4% of snake mass). We released snakes approximately one week after surgery.

We tracked snakes two to five times per week, between 8:00 and 18:00, from 18 October 2005 and to 6 April 2006. When we located a snake, we recorded the rate of pulses emitted by the transmitter and calculated the inter-pulse interval (IPI). IPI indicates the temperature (snake body temperature) measured by the implanted transmitter. We calculated IPI by measuring the time interval for ten pulses with a digital stopwatch and dividing this interval by ten (Hill 2004). This was repeated three times each time we located a snake, and the mean of the three IPI values was later used to calculate body temperature (see "Data analysis"). If we disturbed an animal before recording pulse rate, the data associated with that observation were discarded. After recording pulse rate, we recorded environmental temperatures with a quick-read digital thermometer (air at 1m above ground level, water at 2.5cm below surface, and soil 12cm below surface) as close as possible to the snake's location. We recorded soil temperatures by pushing the temperature probe directly into the soil adjacent to burrows. Thus the soil temperature reported is a measure of soil temperature and not a measure of the air temperature within burrows. To avoid non-independence of temperature measurements, we recorded snake body temperature no more than once per snake in any given 24-h period.

In March-April 2006, we recaptured five snakes (males = 3, females = 2) and removed transmitters. Animals were maintained in the lab after removal surgeries until incisions closed and then released. The other four snakes that were implanted (males = 2, females = 2) could not be recaptured before the batteries in their transmitters died in April 2006.

Data Analysis

We defined over-wintering period based on the frequency of snake movements between observations. We did this because foraging Cottonmouths move frequently (Hein, unpublished data) and we were interested in the period during which snakes were not actively foraging. We inferred that snakes that were not observed moving or foraging had ceased active foraging for the over-wintering period. Between 15 November 2005 and 10 March 2006, snakes moved between burrows in only 6% of observations, as opposed to 96% before and 90% after this period. Additionally, no snakes were observed foraging during this period. Only measurements from this period were included in analyses. All snakes included in analyses were observed alive in late March-early April 2006.

To determine whether snakes conformed to air, water or soil temperatures, we compared snake body temperatures to air, soil and water temperatures using ANOVA. We used calibration curves describing the relationship between IPI and temperature provided by the manufacturer to convert IPI (ms) to body temperature ([degrees]C). We then averaged body temperatures and environmental temperatures for each snake before pooling data from all snakes. We did this because we measured each snake's body temperature and associated environmental temperatures on numerous occasions (Gotelli and Ellison 2004). Averaging within replicates (snakes) allowed us to account for autocorrelation in our repeated measurements of snake body temperatures and associated environmental temperatures. Snake body temperatures and environmental temperature measurements met normality and homoskedasticity assumptions of ANOVA. Transformation of raw temperature data did not improve normality; therefore we performed analyses using untransformed data. We compared mean snake body temperatures to mean air, water and soil temperatures using Tukey's honest significant differences method for multiple comparisons (Tukey's HSD, Crawley 2007).

To determine whether snake body temperatures differed among individuals, we used analysis of covariance (ANCOVA) to regress body temperature with local air temperature. Individual body temperature measurements from snakes met normality and homoskedasticity assumptions of ANCOVA. We used ANCOVA because different snakes were observed on different days (which varied in environmental temperatures) and we suspected that environmental temperature would be an important covariate affecting snake body temperature. We chose air temperature as the covariate because it was the environmental variable most strongly correlated with snake body temperature. All statistical analyses were performed in the R statistical programming environment (R Development Core Team 2006).

RESULTS

Comparing Snake Temperatures to Environmental Temperatures

Transmitter measurements and associated environmental temperature readings yielded 79 body and environmental temperatures from 15 November 2005 to 10 March 2006. We excluded three snakes from analyses because we were unable to locate these individuals until the end of the over-wintering period. Additionally, One of these excluded snakes had a body temperature that was less than 0[degrees]C on two successive occasions. This individual never emerged from its hibernaculum in the spring leading us to believe that it died. We were left with 73 individual observations from the remaining six snakes (males = 4, females = 2, range = 9-13 observations per snake). Mean and standard deviation of body temperatures for each snake are displayed in table 1.

Table 1. Monthly mean (standard deviation) body temperatures of
Cottonmouths used in analyses.

Snake    Nov-05     Dec-05     Jan-06      Feb-06       Mar-06

1         -        23.0 (-)   8.3 (1.1)  11.7 (3.8)   21.2 (1.8)
2         -        17.3 (-)  15.7 (-)    17.6 (1.6)   23.9 (0.2)
3         -           -      22.0 (-)    22.8 (1.0)   24.1 (1.7)
4       16.5 (-)       -     10.5 (-)    11.7 (1.6)   13.2 (0.4)
5      28.0 (1.4)  24.0 (-)  21.9 (3.0)  23.0 (1.0)   25.4 (1.8)
6      21.8 (2.6)  19.5 (-)  17.5 (2.1)  18.3 (1.15)  20.0 (0.9)

* "-" indicates insufficient number of observations to estimate
mean/std. deviation

Overall, mean snake body temperature differed from two of the three environmental temperatures measured (ANOVA: F = 18.19, d.f. = 3, p < 0.001). Mean snake body temperature (20.0'C, 95%CI = 15.9-24.1 C) was higher than either water (15.4 C, 95%CI = 11.3-19.5[degrees]C) or soil (12.0[degrees]C, 95%CI = 7.9-16.1) temperatures but did not differ significantly from mean air (1 8.1[degrees]C, 95%CI = 16.4-19.8) temperature (Fig. 1). Snakes were, on average, 4.6[degrees]C warmer than water (Tukey HSD, adjusted p = 0.003) and 8.0[degrees]C warmer than soil temperatures (Tukey HSD, Bonferroni corrected p < 0.001). During the week when snakes became inactive (9-15 November 2005), mean air, water and soil temperatures were 18.1[degrees]C, I6.4[degrees]C and 8.6[degrees]C respectively. During the week when snakes were first observed above ground (10-16 March 2006), mean air water and soil temperatures were 26.7[degrees]C, 17.0[degrees]C and 14.1[degrees]C respectively.

[FIGURE 1 OMITTED]

Comparing Body Temperatures Among Individuals

Body temperatures differed among the snakes over the entire over-wintering period (Fig. 2, ANCOVA: F = 27.97, d.f. = 4, p < 0.001, intercept and 95% CI of warmest and coolest snakes = 18.0[degrees]C, 13.3-22.6[degrees]C; 7.7[degrees]C, 3.1-12.4[degrees]C). The difference between intercepts for the warmest and coolest individuals was 10.3[degrees]C. The uppermost two regression lines in Fig. 2 represent body temperatures from the two females (intercept and 95%CI = 18.0[degrees]C, 13.3-22.6[degrees]C; 16.7[degrees]C, 11.8-21.6[degrees]C), and the lower three lines represent body temperatures of the three males (intercept and 95%CI = 13.3[degrees]C, 10.7-15.8[degrees]C; 13.1[degrees]C, 8.5-17.7[degrees]C; 7.7[degrees]C, 3.1-12.4[degrees]C). Body temperatures of five of the six individuals were best fitted by a common slope of 0.38 (95% CI = 0.26-0.50). One individual could not be compared using this analysis because the relationship between this snake's body temperature and air temperature differed from that displayed by all other snakes (this snake was consistently located in an inundated burrow). Consequently, we could not use ANCOVA to compare the intercept of this snake to intercepts of the other snakes. Regression analysis indicated a strong relationship between water temperature and the body temperature of this snake (F = 13.33, p=0.003, [r.sup.2] = 0.55, slope and 95%CI = 1.24, 0.58-1.91).

[FIGURE 2 OMITTED]

DISCUSSION

Several authors have suggested that snakes engage in thermoregulatory behavior or select hibernacula during the winter (Prior and Weatherhead 1996, Harvey and Weatherhead 2006, Schuett et al. 2006). This is not surprising given that environmental temperatures drop to lethal levels during winter in many areas (Shine and Mason 2004). The winter basking behavior observed in some species is an example of winter thermoregulation (Schuett et al. 2006). However, we did not observe any snakes above ground during the over-wintering period, which led us to believe that snakes remained within burrows. The Cottonmouth population at our site is known to number at least in the hundreds (C. Guyer unpublished data), and many snakes are typically observed moving and basking above ground during late spring, summer and early fall. Because we did not observe any individual above ground during the over-wintering period, we do not believe that snakes in our population engaged in behavioral thermoregulation outside of their burrows during winter. Rather, Cottonmouths in our study population engaged in within-burrow thermoregulatory behaviors or selected burrows that offered particularly warm winter temperatures.

We acknowledge that any conclusions about over-wintering behavior drawn from this study must be tentative due to the small number of snakes used. However, our study provides a window into over-wintering thermal biology of Cottonmouths, an aspect of their life history that has not been previously described. Although snakes were below ground during all telemetry observations, snake body temperatures were warmer than soil or water temperatures. This result is surprising; since snakes were located in burrows in the soil, we expected their body temperatures to closely reflect soil temperatures. Additionally, snake body temperatures were relatively insensitive to fluctuations in air, water and soil temperatures. The shallow slope of regression lines of snake body temperature on air temperature suggest that snakes were buffered against fluctuations in air temperatures (regressions between body temperatures of these snakes and soil and water temperatures yielded similarly shallow slopes of 0.56, and 0.57 respectively).

Snake body temperatures could have differed from the environmental temperatures that we measured for several reasons. While Cottonmouths are not known to engage in thermogenic behavior such as shivering, some snake species do generate heat by rapidly contracting muscles (e.g. Pythons, Harlow and Grigg 1984). It is also possible that snakes were buffered from extreme temperatures by over-wintering in communal burrows. Although none of the animals that we radio-tracked were located in burrows with one another, it is possible that our study animals shared burrows with other snakes. Finally, the burrows used by our study individuals may have been warmer than the surrounding soil and water. When we excavated burrows in April 2006, we found abundant organic material within burrows. Decomposing organic material may have created localized hot spots that were warmer than the surrounding soil. Clearly, these data emphasize the need for a more comprehensive understanding of how snakes in this and other populations regulate their body temperatures during winter.

A second important observation gleaned from our study is that mean body temperatures of snakes differed from one another. Differences in over-wintering body temperatures may lead to important differences in the costs and consequences of overwintering experienced by different individuals. McCue and Lillywhite (2002) described the relationship between temperature and metabolic rates of Cottonmouths. Following their equations, we estimated an approximately 4.5 fold difference in over-wintering energy consumption between the coolest and warmest snakes (15kJ and 69kJ respectively) in our study. These differences may represent non-trivial differences in the energetic costs of over-wintering among individuals. Blem (1997) and Wharton (1969) identified lipid reserves as an important factor affecting survival of Cottonmouths through over-wintering periods. Blem (1997) suggested that animals might die during warm winters as a result of exhausting lipid reserves. Because body temperature is positively correlated with metabolic rate and consequently, energy consumption, staying warm during the winter may carry a significant energetic cost. Conversely, staying too cold may also carry significant costs. One of the snakes in our study exhibited body temperatures below 0[degrees]C on two occasions in February. We believe this individual died from exposure to lethal low temperatures.

These data represent the first report of Cottonmouth thermal behavior during the winter. The potential costs and benefits of high and low body temperatures during over-wintering and inter-individual variation in over-wintering body temperatures have been under-appreciated in past work. Though limited by the small number of individuals included, our study highlights these important aspects of thermal biology and draws attention to the need for future studies of over-wintering thermoregulation in ectotherms. More complete characterization of over-wintering thermal behavior in this and other species may help to determine the importance of winter thermoregulation to the survival and fitness of temperate reptiles.

ACKNOWLEDGEMENTS

This research was funded by an Auburn University Undergraduate Research Fellowship and Funds for Excellence grant to AMH. State collecting permits were acquired prior to the initiation of this study. The study was conducted in compliance with institutional animal care and use guidelines (Auburn University IACUC# 2004-0668, 2006-0960). We would like to thank M. Mendonca and S. Hoss for technical assistance and J. Gillooly, M. McCoy, A. Ophir and two anonymous reviewers for helpful comments on earlier drafts of this manuscript.

LITERATURE CITED

Beaupre, S. J. 1995. Effects of geographically variable thermal environment on bioenergetics of Mottled Rock Rattlesnakes. Ecology 76: 1655--1665.

Blem, C. R. 1997. Lipid reserves of the Eastern Cottonmouth (Agkistrodon piscivorus) at the northern edge of its range. Copeia 1997: 53-59.

Blem, C. R., and L. B. Blem. 1995. The Eastern Cottonmouth (Agkistrodon piscivorus) at the northern edge of its range. Journal of Herpetology 29: 391-398.

Blouin-Demers, G., and P. J. Weatherhead. 2001. Thermal ecology of black rat snakes (Elaphe obsoleta) in a thermally challenging environment. Ecology 82: 3025--3043.

Blouin-Demers, G. and P. J. Weatherhead. 2002. Habitat-specific behavioural thermoregulation by black rat snakes (Elaphe obsoleta obsoleta). Oikos 97: 59-68.

Cooper, E. L., A. E. Klempau, and A. G. Zapata. 1985. Reptilian immunity. Pp. 600--676, In C. Gans, F. Billett, and P. Maderson (Eds.). Biology of the Reptilia Vol. 14, Development A. Wiley and Sons, New York, USA, 744 pp.

Crane, A. L., and B. D. Greene. 2008. The effect of reproductive condition on thermoregulation in female Agkistrodon piscivorus near the northwestern range limit. Herpetologica 64: 156-167.

Crawley, M. J. 2007. The r book. Wiley, New York, New York, USA.

Dorcas, M. E., W. A. Hopkins, and J. H. Roe. 2004. Effects of body mass and temperature on standard metabolic rate in the eastern diamondback rattlesnake (Crotalus adamanteus). Copeia: 145-151.

Gotelli, N. J., and A. M. Ellison. 2004. A primer of ecological statistics. Sinauer, Sunderland, Massachusets, USA.

Gibson, A. R., and J. B. Falls. 1979. Thermal biology of the common garter snake Thamnophis sirtalis, I. Temporal variation, environmental effects, and sex differences. Oecologia 43: 79--97.

Glaudas, X., K. M. Andrews, J. D. Willson, and J. W. Gibbons. 2007. Migration patterns in a population of cottonmouths (Agkistrodon piscivorus) inhabiting an isolated wetland.Journal of Zoology 271: 19-124.

Gregory, P. T. 1982. Reptilian hibernation. In C. Gans and F. H. Pough [eds.], Biology of the reptilia, Vol. 13. Physiology C, 53-140. Wiley, New York, New York, USA.

Harlow, P., and G. Grigg. 1984. Shivering thermogenesis in brooding diamond python, Python spilotes spilotes. Copeia 4:959-965.

Harvey, D. S., and P. J. Weatherhead. 2006. Hibernation site selection by eastern massasauga rattlesnakes (Sistrurus catenatus catenatus) near their northern range limit. Journal of Herpetology 40: 66--73.

Herczeg, G., A. Herrero, J. Saarikivi, A. Gonda, M. Jantti, and J. Merila. 2008. Experimenta support for the cost--benefit model of lizard thermoregulation: the effects of predation risk and food supply. Oecologia 155: 1-10.

Hertz, P. E., R. B. Huey., and R. D. Stevenson. 1993. Evaluating temperature regulation by field active ectotherms: The fallacy of the inappropriate question. The American Naturalist 142: 796-818.

Hill, J. G. III. 2004. Natural history of the western cottonmouth (Agkistrodon piscivorus leucostoma) from an upland lotic population in the Ozark Mountains of northwest Arkansas. Ph.D. Dissertation. University of Arkansas, Fayetteville, Arkansas, USA.

Hill, J. G. and S. J. Beaupre. 2008. Body size, growth, and reproduction in a population of Western Cottonmouths (Agkistrodon piscivorus leucostoma) in the Ozark Mountains of northwestern Arkansas. Copeia 2008: 105-114.

Himes, J. G., L. M. Hardy, D. C. Rudolph, and S.J. Burgdorf. 2006. Body temperature variation of the Louisiana Pine Snake (Pituophis ruthveni) in a longleaf pine ecosystem. Herpetological Natural History 9: 117-126.

Kapfer, J. M., M. J. Pauers, D. M. Reineke, J. R. Coggins, and R Hay. 2008. Environmental behavioral, and habitat variables influencing body temperature in radio-tagged Bullsnakes, Pituophis catenifer sayi.. Journal of Thermal Biology 33: 174-179.

Luiselli, L., and G. C. Akani. 2002. Is thermoregulation really unimportant for tropical reptiles? Comparative study of four sympatric snake species from Africa. Acta Oecologica 23: 59--68.

McCue, M., and H. B. Lillywhite. 2002. Oxygen consumption and the energetics of island dwelling Florida cottonmouth snakes. Physiological and Biochemical Zoology 75: 165--178.

O'Donnell, R. P., and S. J. Arnold. 2005. Evidence for selection on thermoregulation: Effects of temperature on embryo mortality in the garter snake Thamnophis elegans. Copeia 4: 930-934.

Plummer, M. V. 1993. Thermal ecology of arboreal green snakes (Opheodrys aestivus). Journal of Herpetology 27: 254--260.

Prior, K. A., and P. J. Weatherhead. 1996. Habitat features of Black Rat Snake hibernacula in Ontario. Journal of Herpetology 30: 211--218.

R Development Core Team. 2006. R: A language and environment for statistical computing. Austria: R Foundation for Statistical Computing.

Reinert, H. K., and D. Cundall. 1982. An improved surgical implantation method for radio tracking snakes. Copeia 1988: 964--978.

Seebacher, F., and C. E. Franklin. 2005. Physiological mechanisms of thermoregulation in reptiles: A review. Journal of Comparative Biochemistry and Physiology B 175: 533-541.

Schuett, G. W., R. A. Repp, E.N. Taylor, D. F. DeNardo, R. L. Earley, E. A. Van Kirk, and W. J. Murdoch. 2006. Winter profile of plasma sex steroid levels in free-living male western diamond-backed rattlesnakes, Crotalus atrox (Serpentes: Viperidae). General and Comparative Endocrinology 149: 72-80.

Shine, R., and R. T. Mason. 2004. Patterns of mortality in a cold-climate population of garter snakes (Thamnophis sirtalis parietalis). Biological Conservation 120: 201--210.

Teng, H., and R. S. Wilkinson. 2003. 'Delayed' endocytosis is regulated by extracellular Ca2+ in snake motor boutons. Journal of Physiology 551: 103-114.

Wang, T., M. Zaar, S. Ardvenson, C. Vedel-Smith, and J. Overgaard. 2003. Effects of temperature on metabolic response to feeding in Python molurus. Comparative Biochemistry and Physiology Part A 133: 519-527.

Wharton, C. H. 1969. The cottonmouth moccasin on Sea Horse Key, Florida. Bulletin of the Florida State Museum 14: 227--272.

Andrew M. Hein (1), (2) and Craig Guyer (1)

(1) Department of Biological Sciences, Auburn University, Auburn, Alabama 36849, and

(2) Department of Zoology, University of Florida, Gainesville, Florida 32611-8525

Correspondence: Hein, Andrew M. ([e-mail])