The Metabolic Effort and Duration of Ecdysis in Timber Rattlesnakes: Implications for Time-Energy Budgets of Reptiles.

AbstractTemperate reptiles are often considered to be low-energy systems, with their discrete use of time and energy making them model systems for the study of time-energy budgets. However, the semifrequent replacement and sloughing of the epidermis is a ubiquitous feature of squamate reptiles that is often overlooked when accounting for time and energy budgets in these animals. We used open-flow respirometry to measure both the energetic effort of ecdysis and the duration of the associated metabolic upregulation (likely related to behavioral changes often reported for animals in shed) in wild-caught timber rattlesnakes (Crotalus horridus). We hypothesized that total effort of skin biosynthesis and physical removal would be related to body mass and expected the duration of the process to remain static across individuals at a fixed temperature (25°C). We provide both the first measurements of the cost of skin biosynthesis and physical removal in a reptile and the highest-resolution estimate of process duration recorded to date. We found that skin biosynthesis, but not the cost of physical removal of the epidermis, was related to body mass. Shed cycle duration was consistent across individuals, taking nearly 4 wk from process initiation to physical removal of the outermost epidermal layer. Total energetic effort of ecdysis was of sizeable magnitude, requiring ∼3% of the total annual energy budget of a timber rattlesnake. Energetic effort for a 500-g snake was equivalent to the amount of metabolizable energy acquired from the consumption of approximately two adult mice. Ecdysis is a significant part of the time-energy budgets of snakes, necessitating further attention in studies of reptilian energetics.

Thermal sensitivity of digestion in Sceloporus consobrinus, with comments on geographic variation.

Individual variation in energetics, environment, and genetics can influence population-level processes. However, it is often assumed that locally measured thermal and bioenergetic responses apply among broadly related species. Even closely related taxa may differ in the thermal sensitivity of performance, which in turn influences population persistence, population vital rates, and the ability to respond to environmental changes. The objectives of this project were to quantify the thermal sensitivity of digestive physiology in an Sceloporus lizards, to compare closely related, but geographically distinct, populations. Sceloporus lizards are a model organism, as they are known to exhibit thermally dependent physiologies and are geographically widespread. Digestive passage time, food consumption, fecal and urate production, metabolizable energy intake (MEI), and assimilated energy (AE) were compared for Sceloporus consobrinus in Arkansas and S. undulatus in South Carolina and New Jersey. Published data were acquired for NJ and SC lizards, while original data were collected for S. consobrinus. Comparisons of digestion among populations were made at 30 °C, 33 °C, or 36 °C. Results suggest that digestive physiology differs among populations, with S. consobrinus being more efficient at warmer temperatures. In contrast, NJ and SC lizards had quicker passage times and lower fecal and urate production at 30 °C in comparison to AR. The results of the current study exemplify how closely related organisms can differ in thermal sensitivity of performance. Such data are important for understanding how individual-level processes can vary in response to climate, with implications for understanding variation in physiological traits across the range of Sceloporus lizards.

Cycling temperature treatments affect estimates of digestive performance in prairie lizards (Sceloporus consobrinus).

In nature, many organisms experience a daily range of body temperatures. Thermal performance at stable temperatures is often extrapolated to predict function in cyclical environments. However, temperature order and cyclicity may influence physiological processes. The current study compared energy intake, digestive passage time and energy budgets at a stable temperature (33°C) and two temperature cycles in lizards (Sceloporus consobrinus), to determine (1) whether stable treatments adequately project performance in a cycling environment and (2) whether temperature order influences performance. Cycles had a mean temperature of 33°C, and rotated through 30°C, 33°C and 36°C daily, with equal durations of time at each temperature but differing temperature order, with warm days and cool nights in cycle 1 and cool days and warm nights in cycle 2. For analyses, performance in the stable treatment was compared with that during cycles. If temperature is the primary factor regulating performance, then performance from the stable treatment and cycles should compare favorably. However, physiological performance varied based on temperature treatment. Energy intake and budgets were similar between the stable trial and cycle 1 but not cycle 2. However, passage time did not differ. Notably, the two cycling regimes consistently varied in performance, indicating that temperature order plays a primary role in regulating performance. Physiological data collection requires careful consideration of effects of cycling versus stable temperature treatments. Stable temperatures do not consistently represent performance in cycling regimes and consideration should be paid not only to which temperatures animals experience but also to how temperature is experienced in nature.

Analyzing Time-Energy Constraints to Understand the Links between Environmental Change and Local Extinctions in Terrestrial Ectotherms.

AbstractAccelerated extinction rates have prompted an increased focus on the interplay between environmental change and species response. The effects of environmental change on thermal opportunity are typically considered through a climate change context. However, habitat alteration can also have strong effects on the thermal environment. Additionally, habitat alteration is considered a leading factor of species extinction, yet few studies address the influence of habitat alteration on thermal opportunity and time-energy budgets in at-risk species. Here, we show the strong effects that habitat degradation can have on thermal opportunity, time-energy budgets, and life history demographics of local populations. In the Ozark Mountains of northern Arkansas, woody vegetation encroachment has resulted in a shift in life history traits that appears to play an important role in recent extirpations of eastern collared lizards (Crotaphytus collaris). Populations in degraded habitats experienced a decline in thermal opportunity and less time at body temperatures (time at Tb) suitable for digestion compared with those in intact habitats. We used our data to model the effect of reduced time at Tb on the net assimilated energy available for growth and reproduction. Our model predicts an ∼46% decline in the annual fecundity of individuals, which is similar to empirical observations of reproduction of C. collaris populations in degraded habitats (~49%). We conclude that C. collaris in degraded habitats experienced reduced growth and reproduction primarily as a result of constrained thermal opportunity leading to a decline in digestive processing rates. Our study applies an underappreciated approach to identify the biophysical and time-energy effects of habitat alteration.

Integrating bioenergetics and conservation biology: thermal sensitivity of digestive performance in Eastern Collared Lizards (Crotaphytus collaris) may affect population persistence.

Information on bioenergetics can provide valuable insight into the ecology, life history and population dynamics of organisms. For ectothermic animals, thermal sensitivity of digestion is an important determinant of net assimilated energy budgets. A recent study in the Ozark Mountains indicated that eastern collared lizards (Crotaphytus collaris) restricted to encroached glades (characterized by woody vegetation encroachment) experience reduced environmental heat loads and have reduced age-specific growth and reproductive rates compared to populations in intact glades. To assess the potential impact of reduced body temperatures on assimilation rates of C. collaris in encroached glades, we conducted feeding trials across four temperature treatments (28, 31, 34 and 37°C). We tested for temperature effects on voluntary feeding rates, passage times, apparent assimilated energy (AE) and metabolizable energy (ME). Passage times decreased and voluntary feeding rates increased significantly with increasing temperature. Consumption explained the majority of variance in AE and ME, followed by the effect of temperature treatments. Using data on voluntary feeding rates, passage times and ME as a function of temperature, we estimated over a 10-fold increase in predicted daily assimilated energy across temperature treatments (28°C = 0.58 kJ/day, 31°C = 1.20 kJ/day, 34°C = 4.30 kJ/day, 37°C = 7.95 kJ/day). Thus, lower heat loads in encroached glades may cause reduced body temperature and result in restricted energy assimilation rates. Our study provides a novel approach to the integration of bioenergetics and conservation and shows the efficacy of using information on digestive performance to investigate underlying mechanisms in a conservation context.

The effect of body posture on available habitat and activity-time in a lizard: Implications for thermal ecology studies.

Ectothermic animals contend with variable environmental temperature through behavioral thermoregulation, including selection of activity-times and microhabitat spaces with suitable operative temperatures. Thus, an important component to understanding the influence of temperature on animals is through the assessment of thermal constraints on time and space usage. Thermal ecologists have recognized that postural adjustments are an important part of behavioral thermoregulation. However, the impact of postural adjustments on available space and time has received little attention. We hypothesized that postural adjustments would significantly affect the thermal availability of space and time for surface activity. To test our hypothesis, we used data collected over a four-year study of the thermal ecology of Eastern Collared Lizards (Crotaphytus collaris) in Arkansas. We used a novel approach to model three distinct postures used by C. collaris, and to assess the impact of posture on available space and time. For our study species and habitat, posture had a significant impact on several indices of available space and time including: a) a 13% increase in length of the reproductive activity season, b) a 35% increase in the frequency distribution of habitat within active body temperature range and c) a 42% increase in average thermal quality index. We conclude that posture can significantly impact space and time available for surface activity in species that employ it for thermoregulation. Thus, a clearer understanding of the thermal constraints on time-space usage in ectotherms requires consideration of the impact of posture on the spatiotemporal distribution of thermally suitable microhabitats.

Animal Welfare Policy: Implementation in the Context of Wildlife Research--Policy Review and Discussion of Fundamental Issues.

The use of vertebrate animals in research and education in the United States is subject to a number of regulations, policies, and guidelines under the immediate oversight of Institutional Animal Care and Use Committees (IACUCs), which are charged with ensuring the ethical and appropriate use of the animal subjects. In almost all instances, this regulatory and oversight landscape of animal use has been developed around domesticated animals in biomedical research environments. When the research activities involve wild species, especially in their natural habitat rather than a laboratory, oversight personnel and investigators alike struggle with determining what constitutes ethical and appropriate activities. These difficulties stem from fundamental differences in biology between wild and domesticated animals and from the differences in research objectives and methods in wildlife compared with biomedical research. Here we discuss the various policies, regulations, and guidance documents for animal use in the context of wildlife research. We compare the expectations of the various oversight agencies and how these expectations are met when working with wild vertebrates. We make recommendations for how IACUCs can use available resources to ensure that activities involving wild species are conducted in compliance with existing regulations and policies and in ways that are biologically appropriate for these nondomesticated species.

Male Snakes Allocate Time and Energy according to Individual Energetic Status: Body Condition, Steroid Hormones, and Reproductive Behavior in Timber Rattlesnakes, Crotalus horridus.

Life-history theory predicts that organisms will hedge current reproductive investment against potential costs in terms of survivorship and future fecundity. However, little is known regarding the endocrine mechanisms underlying bet-hedging strategies in free-ranging male vertebrates. We examined the relationships among individual energetic status, steroid hormones, mate search, and reproductive behavior in free-ranging male timber rattlesnakes. Snakes were monitored over four active seasons in order to test two hypotheses: (1) males adjust the amount of time and energy allocated toward reproduction according to the level of individual energy stores, and (2) observed condition-dependent reproductive allocation is associated with circulating concentrations of steroid hormones (testosterone and corticosterone) thought to regulate reproductive behaviors in vertebrates. A positive relationship between body condition and testosterone was observed in both the field and the laboratory. Male mate search effort was positively correlated with both body condition and testosterone. Body condition and testosterone concentrations were negatively related to time allocated toward foraging during the breeding season. A strong effect of year was observed in the analysis of testosterone and search effort, suggesting that multiple environmental factors impact hormone production and reproductive investment. Corticosterone was not related to any measured variable. Therefore, our results did not indicate a clear role of corticosterone in mediating observed relationships between energetic status and behavior. Observed relationships are consistent with the hypothesis that males allocate time and energy toward reproduction according to individual energetic status and that testosterone plays a role in mediating the trade-off between current reproductive investment and residual reproductive value.

Natural variation in steroid hormone profiles of male Timber Rattlesnakes, Crotalus horridus, in northwest Arkansas.

We describe the seasonal profile of circulating steroid hormones (testosterone and corticosterone) in relation to the breeding season in free ranging male Timber Rattlesnakes, Crotalus horridus, over the course of three active seasons. In addition, we examine variation in steroid concentrations across years and in relation to body condition. We found that seasonal profiles of plasma testosterone were different compared to other crotalines with similar mating patterns. Concentrations of testosterone were elevated above baseline in the three months leading up to the single late summer breeding season. Testosterone peaked in July at the onset of the breeding season and dropped to baseline during the peak months of breeding (August and September). Testosterone concentrations also varied annually. Although the exact cause of annual variation could not be established, our results indicate that weather patterns may have driven observed differences. Testosterone concentrations were positively related to body condition, indicating that testosterone production is modulated according to energetic status (particularly in the two months prior to the breeding season). Corticosterone did not vary seasonally or with any measured variable, a result similar to other studied crotalines. Our results highlight the importance of long-term descriptive studies of the regulatory mechanisms that underlie behavior and physiology in diverse taxa, as these mechanisms can vary greatly within and among populations and are valuable in elucidating the intrinsic and extrinsic sources of such variation.

Metabolic cost of venom replenishment by Prairie Rattlesnakes (Crotalus viridis viridis).

Snakes demonstrate a great deal of variation in the amount of venom injected in both predatory and defensive strikes. There is some evidence that snakes can adaptively meter venom dosage. An underlying assumption in the evolution of venom metering is that the production of venom is energetically costly. To date there has been very little research that has quantified the metabolic costs associated with venom production. We used open-flow respirometry to test for significant differences between Prairie Rattlesnakes (Crotalus v. viridis) that had venom extracted and control snakes that did not. Although previous studies demonstrated high metabolic costs for venom production, we found that snakes that had venom extracted did not have significantly higher metabolic rates than control snakes. The pattern of metabolic deviation from baseline measurements was similar for both treatments, and on average snakes that had venom extracted only exhibited a 1.1% increase over baseline compared to a 2.5% increase in control snakes. Our data suggest that venom is not energetically costly to produce and that perhaps other costs associated with venom can better explain the variability in venom expenditure.

Stable isotope tracer reveals that viviparous snakes transport amino acids to offspring during gestation.

Viviparity and placentation have evolved from oviparity over 100 times in squamate reptiles (lizards and snakes). The independent origins of placentation have resulted in a variety of placental morphologies in different taxa, ranging from simple apposition of fetal and maternal tissues to endotheliochorial implantation that is homoplasious with mammalian placentation. Because the eggs of oviparous squamates transport gases and water from the environment and calcium from the eggshell, the placentae of viviparous squamates are thought to have initially evolved to accomplish these functions from within the maternal oviduct. Species with complex placentae have also been shown to rely substantially, or even primarily, on placental transport of organic nutrients for embryonic nutrition. However, it is unclear whether species with only simple placentae are also capable of transporting organic nutrients to offspring. Among viviparous squamates, all of the snakes that have been studied thus far have been shown to have simple placentae. However, most studies of snake placentation are limited to a single lineage, the North American Natricinae. We tested the abilities of four species of viviparous snakes - Agkistrodon contortrix (Viperidae), Boa constrictor (Boidae), Nerodia sipedon (Colubridae: Natricinae) and Thamnophis sirtalis (Colubridae: Natricinae) - to transport diet-derived amino acids to offspring during gestation. We fed [(15)N]leucine to pregnant snakes, and compared offspring (15)N content with that of unlabeled controls. Labeled females allocated significantly more (15)N to offspring than did controls, but (15)N allocation did not differ among species. Our results indicate that viviparous snakes are capable of transporting diet-derived amino acids to their offspring during gestation, possibly via placentation.

Bioenergetic components of reproductive effort in viviparous snakes: costs of vitellogenesis exceed costs of pregnancy.

Reproductive effort has been defined as the proportion of an organism's energy budget that is allocated to reproduction over a biologically meaningful time period. Historically, studies of reproductive bioenergetics considered energy content of gametes, but not costs of gamete production. Although metabolic costs of vitellogenesis (MCV) fundamentally reflect the primary bioenergetic cost of reproductive allocation in female reptiles, the few investigations that have considered costs of reproductive allocation have focused on metabolic costs of pregnancy (MCP) in viviparous species. We define MCP as energetic costs incurred by pregnant females, including all costs of maintaining gestation conditions necessary for embryogenesis. MCP by our definition do not include fetal costs of embryogenesis. We measured metabolic rates in five species of viviparous snakes (Agkistrodon contortrix, Boa constrictor, Eryx colubrinus, Nerodia sipedon, and Thamnophis sirtalis) during vitellogenesis and pregnancy in order to estimate MCV and MCP. Across all species, MCV were responsible for 30% increases in maternal metabolism. Phylogenetically-independent contrasts showed that MCV were significantly greater in B. constrictor than in other species, likely because B. constrictor yolk energy content was greater than that of other species. Estimates of MCP were not significantly different from zero in any species. In viviparous snakes, MCV appear to represent significant bioenergetic expenditures, while MCP do not. We suggest that MCV, together with yolk energy content, represent the most significant component of reptilian reproductive effort, and therefore deserve greater attention than MCP in studies of reptilian reproductive bioenergetics.

Residual yolk energetics and postnatal shell growth in Smooth Softshell Turtles, Apalone mutica.

We examined functions of residual yolk (RY) in hatchling Smooth Softshell Turtles (Apalone mutica). Removal of RY did not affect survival, shell growth, or resting metabolic rates of turtles for 40 d after hatching. Our estimates of metabolic rate suggest that RY can fuel maintenance and activity metabolism for approximately 25 days. A. mutica absorb more than 1g of water in the first 2 weeks of life, which appears to be the basis of post-hatch shell expansion rather than yolk-provisioned growth. Post-hatch growth may be limited by the magnitude of RY remaining at hatching, but RY protein and lipid proportions do not differ from those of freshly-laid eggs. In addition, A. mutica did not use RY to fuel nest emergence. Our results suggest that RY does not fulfill several hypothetical functions in A. mutica, including postnatal growth, catabolic fuel for nest emergence, and long-term nutritional sustenance for maintenance, activity, or hibernation. Instead, A. mutica appear to absorb most yolk prior to hatching, and are left with a minimum of RY. Variation in RY mass with incubation regime in other species suggests that mothers may overprovision their eggs to ensure successful development across a diversity of possible incubation conditions.

Effects of body mass, meal size, fast length, and temperature on specific dynamic action in the timber rattlesnake (Crotalus horridus).

Detailed analysis of animal energy budgets requires information on the cost of digestion (specific dynamic action [SDA]), which can represent a significant proportion of ingested energy (up to 30% in infrequent feeders). We studied the effects of snake mass, temperature (25 degrees and 30 degrees C), fasting time (1 and 5 mo), and prey size (10%-50% of snake mass) on SDA in 26 timber rattlesnakes (Crotalus horridus). We used flow-through respirometry to measure hourly CO(2) production rates (VCO2) for 1 d before and up to 17 d after feeding. Crotalus horridus, like previously studied viperids and boids, show large and ecologically relevant increases in metabolism due to feeding. Depending on treatment and individual, VCO2 increased to 2.8-11.8 times the resting metabolic rate within 12-45 h postfeeding and decreased to baseline within 4.3-15.4 d. Significant effects of snake mass, meal mass, and fast length were detected. Increased temperature decreased the time required to complete the process but had little effect on total energy expended on SDA. Energy expended on SDA increased with increasing fast length, snake mass, and prey mass. Considering all of our data, we found that a simple allometric relationship explained 96.7% of the variation in total CO(2) production during SDA. Calculations suggest that energy devoted to SDA may approach 20% of the total annual energy budget of snakes in nature. Discrepancies between our data and some previous studies draw attention to the fact that the measurement, expression, and analysis of SDA may be sensitive to several methodological and statistical assumptions.