Fecal Corticosterone Metabolites Research Articles

Validation of two enzyme immunoassays for non-invasive glucocorticoid measurement in a lacertid lizard (Podarcis muralis): effects of pharmacological and biological stimuli on faecal corticosterone metabolites and behaviour.

The assessment of stress-related hormone levels using non-invasive methods has gained popularity in mammal and bird welfare, yet its application in reptiles remains limited. Particularly, the exploration of physiological measures such as faecal corticosterone metabolites (FCMs) for reptilian welfare has scarcely been explored. This study aims to validate two enzyme immunoassays (5α-pregnane-3ß,11ß,21-triol-20-one and 11-oxoaetiocholanolone EIA) for monitoring FCM levels in the European common wall lizard (Podarcis muralis). We collected daily faecal samples before (baseline) and after (post-treatment phase) inducing elevated corticosterone levels using transdermal administration of corticosterone (pharmacological treatment) and handling/confinement (biological treatment). We also conducted daily behavioural observations to explore the relationship between stress-related corticosterone changes and behaviour. Although treatments induced significant increases in FCM levels, the effect was much larger in the pharmacological one. Transdermal corticosterone induced a cumulative increase in FCMs over the treatment period, with a higher response observed in females. In contrast, the biological treatment yielded smaller FCM peaks, with no significant sex differences. Overall, 5α-pregnane-3ß,11ß,21-triol-20-one EIA appeared to be more sensitive in detecting these effects. Regarding lizard behaviour, both treatments led to increased hiding and decreased basking compared to baseline. The effects were more pronounced in animals subjected to handling/confinement, despite smaller FCM increases. Our results confirm the suitability of an EIA for monitoring FCMs in both male and female common wall lizards and provide insights into the complexities of using integrated approaches to assess stress, highlighting the need for further research on direct measures to evaluate reptile welfare.

Effects of Intermittent Harem Mating on Parturition-induced Stress and Success Rate of Weaning in C57BL/6JNarl Mice.

Improving the effectiveness of mating schemes for large-scale production of mice is an ongoing challenge in animal facilities. Continuous mating, which requires fewer breeding cages than intermittent mating, has traditionally been used to take advantage of postpartum estrus for efficient production. However, the continuous mating scheme lacks flexibility because it cannot immediately accommodate the reduced needs of mice when production levels are high. In this study, we compared reproductive performance, fecal corticosterone metabolite (FCM) level as a stress indicator, and mouse mortality between the continuous trio (CT) and intermittent quad (IQ) mating schemes. The weaning rates in the IQ scheme were higher than those in the CT scheme (98.8% compared with 85.3%). The FCM levels in IQ female breeders were lower during the first 5 d after parturition than those in CT female breeders. The FCM levels in postpartum females housed with 2 adult mice were significantly higher on days 1, 3, and 5 after giving birth than those of females housed alone. This suggests that the presence of cage mates may induce stress responses in postpartum females. Increasing the individual cage area did not reduce the FCM levels of female breeders when accompanied by cage mates after parturition. In addition, the incidence of dystocia and mortality was lower in IQ breeders than in CT breeders. In summary, this breeding trial suggests that compared with the continuous mating scheme, the intermittent mating scheme improves the welfare of postpartum females with normal breeding performance in the C57BL/6JNarl production colony.

Refining pain management in mice by comparing multimodal analgesia and NSAID monotherapy for neurosurgical procedures

While neurosurgical interventions are frequently used in laboratory mice, refinement efforts to optimize analgesic management based on multimodal approaches appear to be rather limited. Therefore, we compared the efficacy and tolerability of combinations of the non-steroidal anti-inflammatory drug carprofen, a sustained-release formulation of the opioid buprenorphine, and the local anesthetic bupivacaine with carprofen monotherapy. Female and male C57BL/6J mice were subjected to isoflurane anesthesia and an intracranial electrode implant procedure. Given the multidimensional nature of postsurgical pain and distress, various physiological, behavioral, and biochemical parameters were applied for their assessment. The analysis revealed alterations in Neuro scores, home cage locomotion, body weight, nest building, mouse grimace scales, and fecal corticosterone metabolites. A composite measure scheme allowed the allocation of individual mice to severity classes. The comparison between groups failed to indicate the superiority of multimodal regimens over high-dose NSAID monotherapy. In conclusion, our findings confirmed the informative value of various parameters for assessment of pain and distress following neurosurgical procedures in mice. While all drug regimens were well tolerated in control mice, our data suggest that the total drug load should be carefully considered for perioperative management. Future studies would be of interest to assess potential synergies of drug combinations with lower doses of carprofen.

A Comparison of LED with Fluorescent Lighting on the Stress, Behavior, and Reproductive Success of Laboratory Zebra Finches (Taeniopygia guttata).

There are limited evidence-based husbandry recommendations for laboratory zebra finches (Taeniopygia guttata), including appropriate light sources. Light-emitting diode (LED) technology has been shown to improve circadian regulation and reduce stress in some laboratory animal species, such as mice and rats, when compared with cool-white fluorescent (CWF) lighting, but the effects of LED lighting on zebra finches have not been published. We compared the effects of broad-spectrum, blue-enriched (6,500 Kelvin) CWF and flicker-free LED lighting on the behavior, stress, and reproductive outcomes of indoor-housed zebra finches. Using breeding pairs housed in cubicles illuminated with either CWF or LED lighting, we compared the reproductive output as determined by clutch size, hatching rate, and hatchling survival rate. We also compared the behavior of group-housed adult males, first housed under CWF followed by LED lighting, using video recordings and an ethogram. Fecal samples were collected from these males at the end of each recording period, and basal fecal corticosterone metabolite (FCM) levels were compared. A FCM assay for adult male zebra finches was validated for efficacy and accuracy using a capture-restraint acute stress response and parallelism analysis, respectively. The breeding pairs had no significant difference in the clutch size or percent hatching rate, but percent hatchling survival improved under LED with an increased proportion achieving 100% survival. There was no significant difference in FCM between the lighting treatments. However, the activity budgets of the birds were altered, with a reduction in flighted movement and an increase in enrichment manipulation under LED. Overall, these results support the use of blue-enriched, broad-spectrum flicker-free LED as a safe alternative to CWF lighting for breeding and nonbreeding indoor-housed zebra finches.

Biological validation of faecal corticosterone metabolites as a non-invasive stress assessment in translocated California valley quail (Callipepla californica).

US quail species are vulnerable to population declines as a result of climate change, habitat loss and habitat fragmentation, all of which can result in physiological stress. Additionally, population restoration techniques (PRTs), like translocations, also induce stress. Traditional assessments of avian stress hormone levels include capturing and handling birds to extract blood, methods that are inherently stressful and can compound stress analyses. However, the stress hormone corticosterone (CORT) is metabolized from the blood and excreted in faeces as faecal corticosterone metabolites (FCMs). FCMs have been used as a non-invasive measurement of stress hormone levels in a variety of species, but must be validated for each species. The objective of this study was to biologically validate the use of FCMs as a non-invasive measurement of CORT levels in California valley quail (Callipepla californica). Reference and treatment quail were acclimated for 3 weeks in an outdoor aviary. Subsequently, treatment quail were subjected to a simulated 48-h translocation, a common and stress hormone-inducing PRT. Faecal samples were collected every 4h and processed using an enzyme immunoassay. Mean FCM concentrations of treatment quail (41.50 ± 16.13ng/g) were higher than reference FCM concentrations (24.07 ± 10.4ng/g). These results biologically validate the use of FCMs as a non-invasive method to assess CORT levels in California valley quail, demonstrate diurnal variation in quail CORT levels, and confirm that quail translocations are a stress-inducing PRT. Ultimately, this research validates a new non-invasive tool for stress response measurement to advance quail research, management and conservation.

Evaluation of the stress state based on fecal corticosterone metabolite concentrations in captive penguins in Japan.

Fecal corticosterone metabolite (FCM) concentrations, which can be determined noninvasively, have recently been explored as a stress indicator in birds. In our study, we measured FCM concentrations in penguins under nonmolting or molting conditions, cool or hot season, diseased condition, and incubation period. These measurements were conducted in an aquarium that housed king penguins, gentoo penguins, and African penguins. This study aimed to investigate the validity of fecal matter as a stress indicator. Our findings revealed that FCM concentrations were significantly higher in molting individuals than in nonmolting individuals. Compared with the cool season, FCM concentrations were significantly higher in penguins housed outdoors during the hot season. However, no differences were observed in penguins housed indoors. Diseased individuals and an incubating individual showed notably higher FCM concentrations than healthy individuals. Interestingly, the FCM concentration in king penguin that underwent cataract surgery was extremely high before the surgery. However, 1 month postsurgery, it decreased to a level similar to that of healthy individuals. We observed increased FCM concentrations in penguins considered to be exposed to stressors. Notably, FCM concentration decreased after removing the stress factor. The FCM concentration was found to be consistent with the stress state of penguins, suggesting its usefulness as a stress indicator.

Moving beyond the mean: an analysis of faecal corticosterone metabolites shows substantial variability both within and across white-tailed deer populations.

Glucocorticoid (GC) levels have significant impacts on the health and behaviour of wildlife populations and are involved in many essential body functions including circadian rhythm, stress physiology and metabolism. However, studies of GCs in wildlife often focus on estimating mean hormone levels in populations, or a subset of a population, rather than on assessing the entire distribution of hormone levels within populations. Additionally, explorations of population GC data are limited due to the tradeoff between the number of individuals included in studies and the amount of data per individual that can be collected. In this study, we explore patterns of GC level distributions in three white-tailed deer (Odocoileus virginianus) populations using a non-invasive, opportunistic sampling approach. GC levels were assessed by measuring faecal corticosterone metabolite levels ('fCMs') from deer faecal samples throughout the year. We found both population and seasonal differences in fCMs but observed similarly shaped fCM distributions in all populations. Specifically, all population fCM cumulative distributions were found to be very heavy-tailed. We developed two toy models of acute corticosterone elevation in an effort to recreate the observed heavy-tailed distributions. We found that, in all three populations, cumulative fCM distributions were better described by an assumption of large, periodic spikes in corticosterone levels every few days, as opposed to an assumption of random spikes in corticosterone levels. The analyses presented in this study demonstrate the potential for exploring population-level patterns of GC levels from random, opportunistically sampled data. When taken together with individual-focused studies of GC levels, such analyses can improve our understanding of how individual hormone production scales up to population-level patterns.

Physiological stress in eastern black rhinoceros (Diceros bicornis michaeli) as influenced by their density, climatological variables and sexes

AbstractIt is important to understand the physiological stressors in animals especially for threatened species or intensively managed to improve their conservation and optimise their reproduction. We sought to understand changes in stress hormones (faecal glucocorticoid metabolites) in black rhinoceros (Diceros bicornis michaeli) in relation to population density and sex (intrinsic factors) and plant minerals, rainfall and land surface temperature (extrinsic factors). We used non‐invasive faecal sampling techniques on animals of known sex, age and dominance in seven populations of contrasting population densities over 1 year. We measured variability in faecal corticosterone metabolites through radioimmunoassay and related them to population density, sex and faecal calcium, phosphorus, copper, zinc and potassium as characteristic of plant minerals, rainfall and temperature. We used linear mixed models (LMM) to analyse the data. We did not detect a significant relationship between physiological stress parameters and population density. However, we have indications that stress levels increased as rainfall and temperature increased and were correlated negatively and positively with concentration of faecal phosphorus and copper respectively; we found higher stress levels in females than in males. These results suggest that both intrinsic and extrinsic factors explain the variation in physiological stress observed in black rhinoceros.

Reaction to Novel Objects and Fecal Glucocorticoid Metabolite Levels in Two Species of Nocturnal Geckos.

Many reptiles are maintained in captivity and heavily traded, although welfare measures for many species are not well established and are under-researched compared to other animals. In this study, we focused on two of these species: crested geckos (Correlophus ciliatus) and leopard geckos (Eublepharis macularius). To better interpret their behavior in captivity, the individual reaction to novel objects and the fecal glucocorticoid metabolite levels were measured in an attempt to identify the potential correlation between them. Also, we explored if some characteristic of the objects (e.g., color, shape, or smell) resulted in being more attractive to some species and/or individuals. Equivalent responses to different objects were not obtained for all the geckos, the behavioral response being highly individual and context-dependent, although modulated by the species. Individuals which manipulated earlier and interacted longer with novel objects showed lower basal fecal corticosterone metabolite (FCM) levels. Differences according to the species suggested that crested geckos have significantly greater and more variable FCM levels than leopard geckos. Our results can help to understand the reaction of geckos to novelty and have the potential to serve in their welfare assessment, although more studies are needed to proper establish welfare protocols.

Physiological and behavioural adjustment of a wild rodent to laboratory conditions

Wild animals are brought to captivity for different reasons, for example to be kept in zoos and rehabilitation centres, but also for basic research. Such animals usually undergo a process of adjustment to captive conditions. While this adjustment occurs on the behavioural and the physiological level, those are usually studied separately. The aim of this study was to assess both the physiological and behavioural responses of wild wood mice, Apodemus sylvaticus, while adjusting to laboratory conditions. Over the course of four weeks, we measured in wild-caught mice brought to the laboratory faecal corticosterone metabolites and body mass as physiological parameters, stereotypic behaviour and nest-quality, as welfare-linked behavioural parameters, and four personality measures as additional behavioural parameters. The results of our study indicate that mice exhibited an adjustment in both behaviour and physiology over time in the laboratory. While the hormonal stress response decreased significantly, body mass and the proportion of stereotypic behaviours showed a tendency to increase over time. The slight increase of stereotypic behaviours, although not statistically significant, suggests the development of repetitive and non-functional behaviours as a response to laboratory conditions. However, we suggest that those behaviours might have been used by animals as a coping strategy to decrease the physiological stress response. Other behavioural parameters measured, such as boldness and nestbuilding behaviour were stable over time. The information obtained in the present study hints at a complex interplay between behavioural and physiological adjustments of wild animals to laboratory conditions, which should be considered when intending to use wild animals in experimental research.

Evidence-based severity assessment of the forced swim test in the rat.

The forced swim test (FST) is a traditional assay, which has been used for more than 40 years to assess antidepressant effects of novel drug candidates. In recent years, a debate about the test has focused on the assumption that the FST is highly aversive and burdening for the animals because of the earlier anthropomorphic interpretation and designation as a "behavioral despair test". The Directive 2010/63/EU and the German Animal Welfare law require a prospective severity classification of the planned experimental procedures. Still, an objective examination of the animals' burden in this test has not been performed yet. To fill this gap, we conducted an evidence-based severity assessment of the forced swim test in rats according to a 'standard protocol' with a water temperature of 25°C. We examined parameters representing the physiological and the affective state, and natural as well as locomotion-associated behaviors in three separate experiments to reflect as many dimensions as possible of the animal's condition in the test. Hypothermia was the only effect observed in all animals exposed to the FST when using this standard protocol. Additional adverse effects on body weight, food consumption, and fecal corticosterone metabolite concentrations occurred in response to administration of the antidepressant imipramine, which is frequently used as positive control when testing for antidepressant effects of new substances. We conclude that this version of the FST itself is less severe for the animals than assumed, and we suggest a severity classification of 'moderate' because of the acute and short-lasting effects of hypothermia. To refine the FST according to the 3Rs, we encourage confirming the predictive validity in warmer water temperatures to allow the rats to maintain physiological body temperature.

Reduction in Cold Stress in an Innovative Metabolic Cage Housing System Increases Animal Welfare in Laboratory Mice.

Housing in metabolic cages can induce a pronounced stress response. Metabolic cage systems imply housing mice on metal wire mesh for the collection of urine and feces in addition to monitoring food and water intake. Moreover, mice are single-housed, and no nesting, bedding, or enrichment material is provided, which is often argued to have a not negligible impact on animal welfare due to cold stress. We therefore attempted to reduce stress during metabolic cage housing for mice by comparing an innovative metabolic cage (IMC) with a commercially available metabolic cage from Tecniplast GmbH (TMC) and a control cage. Substantial refinement measures were incorporated into the IMC cage design. In the frame of a multifactorial approach for severity assessment, parameters such as body weight, body composition, food intake, cage and body surface temperature (thermal imaging), mRNA expression of uncoupling protein 1 (Ucp1) in brown adipose tissue (BAT), fur score, and fecal corticosterone metabolites (CMs) were included. Female and male C57BL/6J mice were single-housed for 24 h in either conventional Macrolon cages (control), IMC, or TMC for two sessions. Body weight decreased less in the IMC (females-1st restraint: -6.94%; 2nd restraint: -6.89%; males-1st restraint: -8.08%; 2nd restraint: -5.82%) compared to the TMC (females-1st restraint: -13.2%; 2nd restraint: -15.0%; males-1st restraint: -13.1%; 2nd restraint: -14.9%) and the IMC possessed a higher cage temperature (females-1st restraint: 23.7 °C; 2nd restraint: 23.5 °C; males-1st restraint: 23.3 °C; 2nd restraint: 23.5 °C) compared with the TMC (females-1st restraint: 22.4 °C; 2nd restraint: 22.5 °C; males-1st restraint: 22.6 °C; 2nd restraint: 22.4 °C). The concentration of fecal corticosterone metabolites in the TMC (females-1st restraint: 1376 ng/g dry weight (DW); 2nd restraint: 2098 ng/g DW; males-1st restraint: 1030 ng/g DW; 2nd restraint: 1163 ng/g DW) was higher compared to control cage housing (females-1st restraint: 640 ng/g DW; 2nd restraint: 941 ng/g DW; males-1st restraint: 504 ng/g DW; 2nd restraint: 537 ng/g DW). Our results show the stress potential induced by metabolic cage restraint that is markedly influenced by the lower housing temperature. The IMC represents a first attempt to target cold stress reduction during metabolic cage application thereby producing more animal welfare friendlydata.

Acclimation and Blood Sampling: Effects on Stress Markers in C57Bl/6J Mice.

Blood sampling in rodents is common practice in scientific studies. Some of the refined methods widely used are the puncture of the saphenous vein or tail vein, or even tail docking. The handling needs of these different blood sampling methods are different and can directly affect stress, increasing the variability of the study. Moreover, there is less aversion and stress if the animal is accustomed to the environment, handling and technique. Therefore, our study aimed to assess the influence of these three blood sampling techniques (saphenous puncture, tail vein puncture and tail vein docking) and the use of previous acclimation on different indicators of animal stress, assessing blood glucose concentrations and faecal corticosterone metabolites (FCMs). Twenty-four young adult male and female C57Bl6/J mice were divided in three groups by sampling method: tail docking (TD), saphenous vein puncture (SV) and caudal vein puncture (CV) groups. All mice were studied with and without acclimation, which was performed during 9 consecutive days. The results showed that both males and females present very similar responses to the different handling and sampling methods without significant differences. Nevertheless, acclimation in all sampling methods decreased glucose and FCM levels significantly. The method that obtained the lowest glucose and FCM levels with significance was saphenous vein puncture. Therefore, we can say that it causes less stress when performing prior acclimation, even when this involves greater handling of the animal. Our results contribute to refinement within the 3R concept and could serve researchers to programme and select a good handling technique and a welfare-friendly blood sampling method for their experiments.

Anticoagulant rodenticides are associated with increased stress and reduced body condition of avian scavengers in the Pacific Northwest

Anticoagulant rodenticides (AR) have been used globally to manage commensal rodents for decades. However their application has also resulted in primary, secondary, and tertiary poisoning in wildlife. Widespread exposure to ARs (primarily second generation ARs; SGARs) in raptors and avian scavengers has triggered considerable conservation concern over their potential effects on populations. To identify risk to extant raptor and avian scavenger populations in Oregon and potential future risk to the California condor (Gymnogyps californianus) flock recently established in northern California, we assessed AR exposure and physiological responses in two avian scavenger species (common ravens [Corvus corax] and turkey vultures [Cathartes aura]) throughout Oregon between 2013 and 2019. AR exposure was widespread with 51% (35/68) of common ravens and 86% (63/73) of turkey vultures containing AR residues. The more acutely toxic SGAR brodifacoum was present in 83% and 90% of AR exposed common ravens and turkey vultures. The odds of AR exposure in common ravens were 4.7-fold higher along the coastal region compared to interior Oregon. For common ravens and turkey vultures that were exposed to ARs, respectively, 54% and 56% had concentrations that exceeded the 5% probability of toxicosis (>20 ng/g ww; Thomas et al., 2011), and 20% and 5% exceeded the 20% probability of toxicosis (>80 ng/g ww; Thomas et al., 2011). Common ravens exhibited a physiological response to AR exposure with fecal corticosterone metabolites increasing with sum ARs (ΣAR) concentrations. Both female common raven and turkey vultures’ body condition was negatively correlated with increasing ΣAR concentrations. Our results suggest avian scavengers in Oregon are experiencing extensive AR exposure and the newly established population of California condors in northern California may experience similar AR exposure if they feed in southern Oregon. Understanding the sources of ARs across the landscape is an important first step in reducing or eliminating AR exposure in avian scavengers.

The power of a touch: Regular touchscreen training but not its termination affects hormones and behavior in mice.

Touchscreen-based procedures are increasingly used in experimental animal research. They not only represent a promising approach for translational research, but have also been highlighted as a powerful tool to reduce potential experimenter effects in animal studies. However, to prepare the animals for a touchscreen-based test, an often time-consuming training phase is required that has itself been shown to cause increased adrenocortical activity and anxiety-like behavior in mice. While these findings point at a potentially negative effect of touchscreen training at first glance, results have also been discussed in light of an enriching effect of touchscreen training. The aim of the present study was therefore to shed more light on recently reported touchscreen training effects, with a particular focus on the termination of the training routine. Specifically, we investigated whether the termination of regular touchscreen training could constitute a loss of enrichment for mice. Thus, we assessed fecal corticosterone metabolites (FCMs), exploratory-, anxiety-like and home cage behavior in touchscreen-trained mice in comparison to food restricted and ad libitum fed mice, as a restricted diet is an integral part of the training process. Furthermore, we compared these parameters between mice that were continuously trained and mice whose training was terminated 2 weeks earlier. Our results confirm previous findings showing that a mild food restriction increases the animals' exploratory behavior and shifts their activity rhythm. Moreover, touchscreen training was found to increase FCM levels and anxiety-like behavior of the mice. However, no effect of the termination of touchscreen training could be detected, a finding which contradicts the enrichment loss hypothesis. Therefore, we discuss two alternative explanations for the findings. Yet, the current state of knowledge is not sufficient to draw final conclusions at this stage. In compliance with the refinement endeavors for laboratory animals, further research should assess the severity of touchscreen procedures to ensure a responsible and well-founded use of animals for experimental purposes.

Individualized Housing Modifies the Immune-Endocrine System in CD1 Adult Male Mice.

In the last years, different research groups have made considerable efforts to improve the care and use of animals in research. Mice (Mus musculus) are the most widely used animal species in research in the European Union and are sociable and hierarchical creatures. During experiments, researchers tend to individualize males, but no consideration is given to whether this social isolation causes them stress. The aim of this study was, therefore, to explore whether 4 weeks of social isolation could induce changes in different physiological parameters in adult Crl:CD1(ICR) (CD1) males, which may interfere with experimental results. Body weight, blood cells, and fecal corticosterone metabolites levels were the analyzed parameters. Blood and fecal samples were collected at weeks 1 and 4 of the experimental procedure. Four weeks of single housing produced a significant time-dependent decrease in monocytes and granulocytes. Fecal corticosterone metabolite levels were higher in single-housed mice after 1 week and then normalized after 4 weeks of isolation. Body weight, red blood cells, and platelets remained unchanged in both groups during this period. We can, therefore, conclude that social isolation affects some immune and endocrine parameters, and that this should be taken into account in the interpretation of research data.

Is chronic stress a causal mechanism for small mammal population cycles? Reconciling the evidence.

Chronic stress has long been hypothesized to play a role in driving population cycles. Christian (1950) hypothesized that high population density results in chronic stress and mass "die-offs" in small mammal populations. Updated variations of this hypothesis propose that chronic stress at high population density may reduce fitness, reproduction, or program aspects of phenotype, driving population declines. We tested the effect of density on the stress axis in meadow voles (Microtus pennsylvanicus) by manipulating population density in field enclosures over three years. Using fecal corticosterone metabolites as a non-invasive measure of glucocorticoid (GC) concentrations, we found that density alone was not associated with GC differences. However, we found that the seasonal relationship of GC levels differed by density treatment, with high-density populations having elevated GC levels early in the breeding season and decreasing towards late summer. We additionally tested hippocampal glucocorticoid receptor and mineralocorticoid receptor gene expression in juvenile voles born at different densities, with the hypothesis that high density may reduce receptor expression, altering negative feedback of the stress axis. We found that females had marginally higher glucocorticoid receptor expression at high density, no effect in males, and no detectable effect of density on mineralocorticoid receptor expression in either sex. Hence, we found no evidence that high density directly impairs negative feedback in the hippocampus, butrather female offspring may be better equipped for negative feedback. We compare our findings with prior studies to attempt to disentangle the complicated relationship between density, seasonality, sex, reproduction and the stress axis.

Ablation of Gabra5 Influences Corticosterone Levels and Anxiety-like Behavior in Mice.

Stress responses are activated by the hypothalamic-pituitary-adrenal axis (HPA axis), culminating in the release of glucocorticoids. During prolonged periods of secretion of glucocorticoids or inappropriate behavioral responses to a stressor, pathologic conditions may occur. Increased glucocorticoid concentration is linked to generalized anxiety, and there are knowledge gaps regarding its regulation. It is known that the HPA axis is under GABAergic control, but the contribution of the individual subunits of the GABA receptor is largely unknown. In this study, we investigated the relationship between the α5 subunit and corticosterone levels in a new mouse model deficient for Gabra5, which is known to be linked to anxiety disorders in humans and phenologs observed in mice. We observed decreased rearing behavior, suggesting lower anxiety in the Gabra5-/- animals; however, such a phenotype was absent in the open field and elevated plus maze tests. In addition to decreased rearing behavior, we also found decreased levels of fecal corticosterone metabolites in Gabra5-/- mice indicating a lowered stress response. Moreover, based on the electrophysiological recordings where we observed a hyperpolarized state of hippocampal neurons, we hypothesize that the constitutive ablation of the Gabra5 gene leads to functional compensation with other channels or GABA receptor subunits in this model.

High-quality habitats and refuges from tourism reduce individual stress responses in a forest specialist

Context The assessment of suitable habitat for a species is often evaluated based merely on where the species is most abundant. However, a variety of environmental factors such as temperature, resources and human disturbances might force animals to avoid otherwise high-quality habitats. Additionally, individuals’ capacities to cope with such factors – as reflected in physiological stress responses – may also contribute to avoidance. Combining measures of stress response of individual animals with habitat suitability models may contribute essential insights to the development of species conservation plans. Aims We used a landscape-level approach to assess whether habitat quality, abiotic factors and human disturbance affect the physiological stress response of a forest specialist, the capercaillie (Tetrao urogallus), and to what extent these relationships are determined by variation between individuals. Methods We applied a non-invasive approach by collecting 1096 droppings across the entire species range in two adjacent national parks spanning the border between the Czech Republic and Germany. We measured faecal corticosterone metabolites (FCMs) to evaluate the physiological stress response. To test the influences of temperature, habitat quality, season and sex, as well as forest management and tourism intensity on FCM levels, we used generalised additive mixed effect models (GAMMs). The same models were applied for a reduced subset of randomly selected samples that were attributed to individuals via DNA profiling using 10 microsatellite loci. This approach allowed us to control for possible individual differences (through repeated measures within individual genotypes). Key results Both models revealed decreasing FCM levels with increasing habitat quality and showed no relationship between increasing forestry intensity and FCMs. When accounting for individual identity, we found a significant positive effect from increasing tourism intensity, although this relationship was not significant in the full dataset. Conclusions Our results highlight three conclusions for the conservation of capercaillie: (1) high-quality habitats reduce stress levels; (2) tourism-free refuge areas are important, and (3) individuality appears to be a strong determinant of response to environmental factors. Implications Generalising management recommendations from observations for single individuals to an entire population may be inappropriate.

Hidden but Potentially Stressed: A Non-Invasive Technique to Quantify Fecal Glucocorticoid Levels in a Fossorial Amphisbaenian Reptile.

To understand wildlife responses to the changing environment, it is useful to examine their physiological responses and particularly their endocrine status. Here, we validated an enzyme immunoassay (EIA) to non-invasively quantify fecal corticosterone metabolites (FCM) in the fossorial amphisbaenian reptile Trogonophis wiegmanni from North Africa. We supplemented animals assigned to the treatment group with corticosterone dissolved in oil applied non-invasively on the skin for several days, while control groups received the oil-alone solution. Fresh feces were collected at the end of the supplementation period, and FCM levels were quantified by an EIA. Basal FCM levels were similar for both treatments and increased at the end of the test, but FCM increased significantly more in corticosterone-treated animals. A further examination of FCM levels in a wild population of this amphisbaenian did not find overall sexual, size or seasonal differences but showed a high range of variation among individuals. This suggests that different uncontrolled intrinsic or local environmental variables might increase the circulating glucocorticoid levels of different individuals. Our results confirmed the suitability of EIA for analyzing physiological changes in FCM in this amphisbaenian species. This technique may be useful for understanding and remediating the little-explored potential stressors of the soil environment that may negatively affect the health state of fossorial reptiles.