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Animals cope with environmental perturbations through the stress response, a set of behavioural and physiological responses aimed to maintain and/or return to homeostasis and enhance fitness. Vertebrate neuroendocrine axis activation in response to environmental stressors can result in the secretion of glucocorticoids (GCs), whose acute increases may be adaptive, while chronic elevation may be detrimental. Invasive grey squirrels ( Sciurus carolinensis ) act as a stressor eliciting elevation of GCs in native red squirrels ( Sciurus vulgaris ). Here we used 6-year data of variation in faecal glucocorticoid metabolite (FGM) concentrations following invasion by grey squirrels in three red squirrel populations, to identify if red squirrels showed physiological habituation to this stressor. The decrease in FGMs over time was more pronounced shortly after invasion and at high densities of grey squirrels while it decreased less strongly and was no longer influenced by the invader density as time since invasion elapsed. At the individual level, FGMs also decreased more markedly as each red squirrel experienced prolonged contact with the invader. Our study provides compelling new data suggesting that native species in the wild can habituate to prolonged contact with invasive species, showing that they may avoid the potentially harmful effects of chronic elevations in GCs.

A wide range of species have been found to differentiate kin from nonkin. However, the ability to recognize kin, or the costs and benefits of discriminating kin from nonkin may depend on particular extrinsic environmental or intrinsic physiological conditions, resulting in context-dependent kin discrimination. North American red squirrels, Tamiasciurus hudsonicus, have previously been shown to be capable of discriminating the territorial calls of kin and nonkin. More recently, post hoc analysis of existing data suggested that kin discrimination might depend on the state of the caller (i.e. acute stress). Here we tested this hypothesis with independently collected data from a repeated-measures playback experiment and found no evidence in support of acute stress mediating kin discrimination. While squirrels were shown to register and respond to playbacks, exposure to a trapping/handling stressor in the caller had no effect on the receiver's response to the calls of kin and nonkin. Furthermore, we found no overall discrimination between the calls of kin and nonkin. These independently collected data instead suggest that the previous post hoc finding consistent with context-dependent kin discrimination was likely a spurious relationship associated with the data from which the post hoc hypothesis was generated rather than representing a real biological relationship. Our findings from this study emphasize the importance of testing post hoc hypotheses with independent experiments.

Understanding the causes and consequences of repeatable among-individual differences in behavior (i.e., animal personality) is a major area of research in behavioral and evolutionary ecology. Recently, attention has turned to understanding the processes behind changes in repeatability through ontogeny because of their implications for populations. We evaluated the relative importance of selective disappearance (i.e., differential mortality), an among-individual mechanism, in generating age-related changes in the repeatability of aggression and activity in juvenile North American red squirrels (Tamiasciurus hudsonicus). We observed age-related decreases in the repeatability of aggression across ages, arising from lower among-individual variance. Although we found evidence for directional selection on aggressiveness, it was insufficient to erode among-individual variance. Thus, ontogenetic decreases in the repeatability of aggression do not appear to be due to selective disappearance. In contrast, the repeatability of activity was higher across ages due to higher among-individual variance in activity, but there was no support for selective disappearance based on activity. Taken together, our results suggest that age-related changes in trait repeatability in red squirrels are not the result of selective disappearance and instead may be the result of within-individual developmental processes, such as individual differences in developmental trajectories.

This is a manuscript version of an article published as: Haines, J.A., Nason, S.E., Skurdal, A.M.M., Bourchier, T., Boutin, S., Taylor, R.W., McAdam, A.G., Lane, J.E., Kelley, A.D., Humphries, M.M., Gorrell, J.C., Dantzer, B., Coltman, D.W., & Hämäläinen, A. (2020). Sex- and context-specific associations between personality and a measure of fitness but no link with life history traits. Animal Behaviour, 167, 23-29. https://doi.org/10.1016/j.anbehav.2020.06.013 The pace-of-life syndrome hypothesis posits that personality traits (i.e. consistent individual differences in behaviour) are linked to life history and fitness. Specifically, fast-paced individuals are predicted to be proactive (i.e. active and aggressive) with an earlier age at first reproduction, a shorter life span and higher fecundity than slow-paced individuals. Environmental conditions and sex differences may be important in maintaining behavioural and life history variation in populations and may influence the covariance of personality with life history or lifetime fitness. However, these effects are rarely tested together. We investigated whether the occurrence of a resource pulse (called a mast year) during adulthood altered the associations between personality and life history traits or lifetime offspring production in adult North American red squirrels, Tamiasciurus hudsonicus. Despite accounting for environmental context during adulthood, we found no evidence of an overall pace-of-life syndrome in this population as personality was not associated with age at first reproduction or longevity in either sex. Males and females had similar activity levels, but females were more aggressive, potentially due to the fitness benefits of protecting their offspring from predation. In all females, regardless of mast experience, there was no association between activity and lifetime pup production, but there was a positive association between aggression and lifetime pup production. In males that experienced a mast, there was a positive association between lifetime pup production and both activity and aggression. In males that did not experience a mast, there was no association between activity and lifetime pup production, but there was a negative association between aggression and lifetime pup production. Lifetime recruit production (number of adult offspring recruited into the breeding population) in either sex was not influenced by activity or aggression regardless of mast experience. Overall, our results suggest that the infrequent occurrence of mast years may contribute to maintaining variation in personality traits in red squirrels. Pre-print version This is a manuscript version of an article published as: Haines, J.A., Nason, S.E., Skurdal, A.M.M., Bourchier, T., Boutin, S., Taylor, R.W., McAdam, A.G., Lane, J.E., Kelley, A.D., Humphries, M.M., Gorrell, J.C., Dantzer, B., Coltman, D.W., & Hämäläinen, A. (2020). Sex- and context-specific associations between personality and a measure of fitness but no link with life history traits. Animal Behaviour, 167, 23-29. https://doi.org/10.1016/j.anbehav.2020.06.013 https://viurrspace.ca/bitstream/handle/10613/23323/GorrellAB.pdf?sequence=3

Animals switch between inactive and active states, simultaneously impacting their energy intake, energy expenditure and predation risk, and collectively defining how they engage with environmental variation and trophic interactions. We assess daily activity responses to long-term variation in temperature, resources and mating opportunities to examine whether individuals choose to be active or inactive according to an optimisation of the relative energetic and reproductive gains each state offers. We show that this simplified behavioural decision approach predicts most activity variation (R2 = 0.83) expressed by free-ranging red squirrels over 4 years, as quantified through accelerometer recordings (489 deployments; 5066 squirrel-days). Recognising activity as a determinant of energetic status, the predictability of activity variation aggregated at a daily scale, and the clear signal that behaviour is environmentally forced through optimisation of gain, provides an integrated approach to examine behavioural variation as an intermediary between environmental variation and energetic, life-history and ecological outcomes.

The episodic production of large seed crops by some perennial plants, is referred to as masting and is known to increase seed escape by alternately starving and swamping seed predators. These pulses of resources, however, might also act as an agent of selection on the life histories of seed predators, which could indirectly enhance seed escape by inducing an evolutionary load on seed predator populations. Lag loads in seed predators could result from mast-induced shifts in optimum phenotypes that exceed the capacity of seed predators to adaptively track optimum phenotypes through phenotypic plasticity. Alternatively, masting could generate mismatches in selection across generations, where adaptation to the parental environment leads to maladaptation in the offspring environment. Here we measured natural selection on female North American red squirrels (Tamiasciurus hudsonicus) across 28 years and five white spruce (Picea glauca) masting events. Red squirrel litter sizes were similar to optimum litter sizes during non-mast years, but were well below optimum litter sizes during resource-rich mast years. Mast events, therefore caused selection for larger litters (B = 0.25) and a lag load (L = 0.25) on red squirrels during mast years. Furthermore, we found that the annual fitness of spruce trees was negatively related to the local density of squirrels during mast years, indicating that the observed lag load on squirrels enhanced the number of spruce cones escaping squirrel predation. Although, the frequency of mast events and the demography of red squirrels were such that offspring and parents often experienced opposite environments with respect to the mast, we found no effect of environmental mismatches across generations on either offspring survival or population growth. Instead, squirrels plastically increased litter sizes in anticipation of mast events, which partially, although not completely, reduced the lag load resulting from this change in food availability. Variable selection on litter size caused by white spruce mast events, therefore, induced a lag load on the population of red squirrels that was not affected by whether individual squirrels were born during mast (matching) or non-mast (mismatching) conditions.

Acoustic signaling is an important means by which animals communicate both stable and labile characteristics. Although it is widely appreciated that vocalizations can convey information on labile state, such as fear and aggression, fewer studies have experimentally examined the acoustic expression of stress state. The transmission of such public information about physiological state could have broad implications, potentially influencing the behavior and life-history traits of neighbors. North American red squirrels (Tamiasciurus hudsonicus) produce vocalizations known as rattles that advertise territorial ownership. We examined the influence of changes in physiological stress state on rattle acoustic structure through the application of a stressor (trapping and handling the squirrels) and by provisioning squirrels with exogenous glucocorticoids (GCs). We characterized the acoustic structure of rattles emitted by these squirrels by measuring rattle duration, mean frequency, and entropy. We found evidence that rattles do indeed exhibit a “stress signature.” When squirrels were trapped and handled, they produced rattles that were longer in duration with a higher frequency and increased entropy. However, squirrels that were administered exogenous GCs had similar rattle duration, frequency, and entropy as squirrels that were fed control treatments and unfed squirrels. Our results indicate that short-term stress does affect the acoustic structure of vocalizations, but elevated circulating GC levels do not mediate such changes.

Organisms can affect one another's phenotypes when they socially interact. Indirect genetic effects occur when an individual's phenotype is affected by genes expressed in another individual. These heritable effects can enhance or reduce adaptive potential, thereby accelerating or reversing evolutionary change. Quantifying these social effects is therefore crucial for our understanding of evolution, yet estimates of indirect genetic effects in wild animals are limited to dyadic interactions. We estimated indirect phenotypic and genetic effects, and their covariance with direct effects, for the date of spring breeding in North American red squirrels (Tamiasciurus hudsonicus) living in an array of territories of varying spatial proximity. Additionally, we estimated indirect effects and the strength of selection at low and high population densities. Social effects of neighbours on the date of spring breeding were different from zero at high population densities but not at low population densities. Indirect phenotypic effects accounted for a larger amount of variation in the date of breeding than differences attributable to the among-individual variance, suggesting social interactions are important for determining breeding dates. The genetic component to these indirect effects was however not statistically significant. We therefore showcase a powerful and flexible method that will allow researchers working in organisms with a range of social systems to estimate indirect phenotypic and genetic effects, and demonstrate the degree to which social interactions can influence phenotypes, even in a solitary species.

Animals in urban areas that experience frequent exposure to humans often behave differently than those in less urban areas, such as exhibiting less vigilance or anti-predator behavior. These behavioral shifts may be an adaptive response to urbanization, but it may be costly if animals in urban areas also exhibit reduced anti-predator behavior in the presence of natural predators. In trials with only a human observer as the stimulus, urban squirrels exhibited reduced vigilance and anti-predator behavior compared to those in less urban areas. Next, we exposed squirrels in multiple urban and less urban sites to acoustic playbacks of a control stimulus (non-predatory bird calls), a natural predator (hawk), and dogs and recorded their vigilance and three different anti-predator behaviors when a human approached them while either broadcasting one of these three playbacks or no playbacks at all. Squirrels at urban sites also did not differ in their behavioral responses to the playbacks from possible predators (hawks or dogs) when they were compared to those at less urban sites exposed to these playbacks. Urban squirrels also exhibited increased vigilance and anti-predator behavior when exposed to a human paired with hawk playbacks compared to the control playbacks. Together, our results indicate that urban squirrels did perceive and assess risk to the natural predator appropriately despite exhibiting increased tolerance to humans. These results provide little support for the hypothesis that increased tolerance to humans causes animals to lose their fear of natural predators.

Environmental factors experienced during development can affect the physiology and behavior of offspring. Maternal glucocorticoids (GCs) may convert environmental cues experienced by the mother into a cue triggering adaptive developmental plasticity in offspring. In North American red squirrels (Tamiasciurus hudsonicus), females exhibit increases in GCs when conspecific density is elevated, and selection favors more aggressive and perhaps more active mothers under high density conditions. We experimentally elevated maternal GCs during gestation or early lactation to test the hypothesis that elevated maternal GCs cause shifts in offspring aggression and activity that may prepare them for high density conditions. When offspring were weaned, we measured two behavioral traits (activity and aggression) using a standardized behavioral assay. Because maternal GCs may influence offspring hypothalamic-pituitary-adrenal (HPA) axis activity and HPA axis activity may in turn affect offspring behavior, we also measured the impact of our treatments on offspring HPA axis activity (adrenal reactivity and negative feedback) and the association between offspring HPA axis activity and behavior. Increased maternal GCs during lactation, but not gestation, only slightly elevated activity levels in offspring. Offspring aggression, adrenal reactivity, and negative feedback did not differ between GC-treated and control groups. Offspring with higher adrenal reactivity did exhibit lower aggression, but the relationship between adrenal reactivity and aggression was not affected by treatment with maternal GCs. These results suggest maternal GCs during gestation or early lactation alone may not be a sufficient cue to produce changes in behavioral and physiological stress responses in offspring in natural populations.Summary StatementWe found maternal glucocorticoid levels do not influence offspring personality or HPA axis dynamics in North American red squirrels. Regardless of maternal glucocorticoid treatment, more aggressive squirrels exhibited lower adrenal reactivity.

As a response to environmental cues, maternal glucocorticoids (GCs) may trigger adaptive developmental plasticity in the physiology and behavior of offspring. In North American red squirrels (Tamiasciurus hudsonicus), mothers exhibit increased GCs when conspecific density is elevated, and selection favors more aggressive and perhaps more active mothers under these conditions. We tested the hypothesis that elevated maternal GCs cause shifts in offspring behavior that may prepare them for high-density conditions. We experimentally elevated maternal GCs during gestation or early lactation. We measured two behavioral traits (activity and aggression) in weaned offspring using standardized behavioral assays. Because maternal GCs may influence offspring hypothalamic–pituitary–adrenal (HPA) axis dynamics, which may in turn affect behavior, we also measured the impact of our treatments on offspring HPA axis dynamics (adrenal reactivity and negative feedback), and the association between offspring HPA axis dynamics and behavior. Increased maternal GCs during lactation, but not gestation, slightly elevated activity levels in offspring. Offspring aggression and adrenal reactivity did not differ between treatment groups. Male, but not female, offspring from mothers treated with GCs during pregnancy exhibited stronger negative feedback compared with those from control mothers, but there were no differences in negative feedback between lactation treatment groups. Offspring with higher adrenal reactivity from mothers treated during pregnancy (both controls and GC-treated) exhibited lower aggression and activity. These results suggest that maternal GCs during gestation or early lactation alone may not be a sufficient cue to produce substantial changes in behavioral and physiological stress responses in offspring in natural populations.

Parental investment theory predicts that observed levels of parental care afforded to offspring are set by the benefits (to offspring quality and survival) relative to the costs (to parental survival or future reproduction). Although difficult to document in mammals, there is often substantial individual variation in the amount of parental care within species. We measured the impact of individual variation in maternal care (“attentiveness” towards offspring or maternal motivation) on offspring growth and survival in a wild population of North American red squirrels (Tamiasciurus hudsonicus). We used latency to return to pups following a nest intrusion as a measure of maternal attentiveness to pups. We found this behavior to be repeatable within individuals suggesting this behavior is a personality trait or a “maternal style.” In this population, postnatal growth rate is important for pup overwinter survival. Pups from large litters grew faster if they had a highly attentive mother, indicating that maternal care can mitigate the trade-off between litter size and offspring growth and potentially improve survival of pups. Additionally, more attentive mothers had slightly higher lifetime reproductive success than less attentive mothers. These results highlight important fitness effects of having a highly attentive mother and show that maternal care can alter a fundamental life history trade-off between offspring quantity and quality. It pays to be attentive to your pups as a squirrel mom. In a long-term study of a wild population of North American red squirrels, we tested whether maternal behavior was predictive of growth rate and survival of offspring. To do this, we observed maternal attentiveness towards offspring by recording the time until the mother returned to her pups following a nest intrusion by researchers. We found repeatable individual variation in maternal attentiveness suggesting consistent “maternal styles” among individuals. Mothers who returned faster to pups following a nest intrusion produced faster growing pups and were able to produce larger fast-growing litters. Over their entire lifetime, attentive mothers also had more offspring survive to adulthood.

Interactions between organisms are ubiquitous and have important consequences for phenotypes and fitness. Individuals can even influence those they never meet, if they have extended phenotypes that alter the environments others experience. North American red squirrels (Tamiasciurus hudsonicus) guard food hoards, an extended phenotype that typically outlives the individual and is usually subsequently acquired by non-relatives. Hoarding by previous owners can, therefore, influence subsequent owners. We found that red squirrels breed earlier and had higher lifetime fitness if the previous hoard owner was a male. This was driven by hoarding behaviour, as males and mid-aged squirrels had the largest hoards, and these effects persisted across owners, such that if the previous owner was male or died in mid-age, subsequent occupants had larger hoards. Individuals can, therefore, influence each other's resource-dependent traits and fitness without ever meeting, such that the past can influence contemporary population dynamics through extended phenotypes.

Free-living animals cope with environmental stressors through physiological and behavioural responses. According to the unidimensional model, these responses are integrated within a coping style: proactive individuals (bold, active-explorative and social) have a lower hypothalamic-pituitary-adrenal (HPA) axis reactivity than reactive ones (shy, less active-explorative, less social). These associations may change when individuals are exposed to human-induced rapid environmental change (HIREC), such as the introduction of invasive alien species (IAS). Here, we studied Eurasian red squirrels to investigate the relationship between personality traits and one integrated measure of HPA axis activity, both in areas uncolonized (natural populations) and colonized by an IAS, the Eastern grey squirrel (invaded populations). We expected an association between physiological and behavioural responses, and that activity, exploration and social tendency would covary, forming a behavioural syndrome in natural populations, while competition with the IAS was predicted to disrupt these associations. We used faecal glucocorticoid metabolites (FGMs) as an integrated measure of adrenocortical activity, and measured the levels of four personality traits (exploration, activity, activity-exploration and social tendency) with an open field test and a mirror image stimulation test. We found no correlation between FGMs and personality traits, neither in natural nor invaded populations. However, we found correlations among personality traits in areas without interspecific competition, indicating a behavioural syndrome, which was disrupted in invaded populations. This is one of the few studies showing that an IAS, acting as an environmental stressor, alters a native species' behavioural syndrome, but does not influence its coping style.

Background Our understanding of gut microbiota has been limited primarily to findings from human and laboratory animals, but what shapes the gut microbiota in nature remains largely unknown. To fill this gap, we conducted a comprehensive study of gut microbiota of a well-studied North American red squirrel (Tamiasciurus hudsonicus) population. Red squirrels are territorial, solitary, and live in a highly seasonal environment and therefore represent a very attractive system to study factors that drive the temporal and spatial dynamics of gut microbiota. Result For the first time, this study revealed significant spatial patterns of gut microbiota within a host population, suggesting limited dispersal could play a role in shaping and maintaining the structure of gut microbial communities. We also found a remarkable seasonal rhythm in red squirrel’s gut microbial composition manifested by a tradeoff between relative abundance of two genera Oscillospira and Corpococcus and clearly associated with seasonal variation in diet availability. Our results show that in nature, environmental factors exert a much stronger influence on gut microbiota than host-associated factors including age and sex. Despite strong environmental effects, we found clear evidence of individuality and maternal effects, but host genetics did not seem to be a significant driver of the gut microbial communities in red squirrels. Conclusion Taken together, the results of this study emphasize the importance of external ecological factors rather than host attributes in driving temporal and spatial patterns of gut microbiota in natural environment. Electronic supplementary material The online version of this article (10.1186/s40168-017-0382-3) contains supplementary material, which is available to authorized users.

Nearly all animals interact with members of their own species at some point during their lives. These behavioral interactions range from courtship, mating, and parental care to the complex cooperative behavior among related or unrelated individuals in group-living species. A number of theoretical models have attempted to explain how cooperation can evolve through natural selection. Although tremendously influential in animal behavior research, these traditional models have largely ignored individual variation in cooperative behavior and its underlying developmental and proximate mechanisms. However, a set of emerging models suggest that the evolution of cooperation can be heavily influenced by the degree of individual variation in cooperative behavior, as well as the complexity of the underlying mechanisms. Yet, while theoreticians argue the importance of studying individual variation in cooperation and the mechanisms underlying it, empiricists have not focused upon these aspects. The main objectives of our symposium at the 2017 meeting of the Society for Integrative and Comparative Biology is to establish new research avenues to study variation in cooperative behavior using both proximate and ultimate explanations and to produce a road map to study the developmental and proximate mechanisms in generating individual variation in cooperative behavior. This symposium brought together empiricists and theoreticians investigating cooperative behavior in diverse taxa and across multiple levels of analysis. Here we briefly describe the rationale for this symposium and why we thought it was needed as well as provide a brief overview of the contributions.

The National Environment Research Council (RG53472 to T.H.C-B.), the European Research Council (294494 to T.H.C-B.), the University of Zurich and the Mammal Research Institute at the University of Pretoria.

Short-term elevation of glucocorticoids (GCs) is one of the major physiological mechanisms by which vertebrates cope with challenging environmental or social factors (stressors). However, when exposure to stressors occurs repeatedly or over a prolonged period of time, animals may experience chronic elevation of GCs, which reduces the immune response efficiency and can lead to higher intensity of parasitic infection. Here, we used invasive gray squirrels Sciurus carolinensis introduced in Northern Italy and their 2 most prevalent gastrointestinal parasites, the nematode Strongyloides robustus and coccidia of the genus Eimeria, as a model to investigate relationships among macroparasite infection and concentrations of fecal glucocorticoid metabolites (FGMs), an integrated measure of circulating GCs. Our results revealed an association of FGMs with infection by St. robustus, but not with coccidia. Individuals with higher FGMs appear to be responsible for the greatest St. robustus egg shedding within gray squirrel populations, thus possibly acting as superspreaders. However, FGMs were negatively associated with adult St. robustus, suggesting that the abundance of adults of this nematode species does not induce elevation in FGMs, but is only affected by it through immune-mediated effects on its fecundity. Finally, the relationship between St. robustus (both eggs and adult parasites) and FGMs was not linear, suggesting that only high levels of physiological stress influence parasite infection. Our findings highlight that the direction and magnitude of the stress–infection relationship may depend not only on the specific host–parasite system, but also on the different life stages of the same parasite.

Individuals vary in their behavioral and physiological responses to environmental changes. These behavioral responses are often described as ‘coping styles’ along a proactive-reactive continuum. Studies in laboratory populations often, but not always, find that behavioral responses and physiological responses to stressors covary, where more proactive (more aggressive and active) individuals have a lower physiological stress response, specifically as measured by hypothalamic-pituitary-adrenal (HPA) axis activity. These studies support the possibility of hormonal pleiotropy underlying the presentation of behaviors that make up the proactive-reactive phenotype. However, recent research in wild populations is equivocal, with some studies reporting the same pattern as found in many controlled laboratory studies, whereas others do not. We tested the hypothesis that physiological and behavioral stress responses are correlated in wild adult North American red squirrels (Tamiasciurus hudsonicus). We used fecal cortisol metabolites (FCMs) as a non-invasive, integrated estimate of circulating glucocorticoids for our measurement of HPA axis activity. We found that FCM concentrations were not correlated with three measures of behavioral coping styles (activity, aggression, and docility) among individuals. This does not support the hypothesis that hormonal pleiotropy underlies a proactive-reactive continuum of coping styles. Instead, our results support the “two-tier” hypothesis that behavioral and physiological stress responses are independent and uncorrelated traits among individuals in wild populations that experience naturally varying environments rather than controlled environments. If also found in other studies, this may alter our predictions about the evolutionary consequences of behavioral and endocrine coping styles in free-living animals.Significance StatementIndividuals vary in how they respond to stressors through behavior and physiology, but we find the two responses are independent in wild animals. Many laboratory studies find links between the behavioral and physiological stress responses, however studies conducted with wild populations are less conclusive. In wild North American red squirrels, independence between the physiological response and behavioral response may allow adaptive responses to a changing environment without pleiotropic constraint.

Elevations in glucocorticoid levels (GCs) in breeding females may induce adaptive shifts in offspring life histories. Offspring produced by mothers with elevated GCs may be better prepared to face harsh environments where a faster pace of life is beneficial. We examined how experimentally elevated GCs in pregnant or lactating North American red squirrels (Tamiasciurus hudsonicus) affected offspring postnatal growth, structural size, oxidative stress levels (two antioxidants and oxidative protein damage) in three different tissues (blood, heart, liver), and liver telomere lengths. We predicted that offspring from mothers treated with GCs would grow faster but would also have higher levels of oxidative stress and shorter telomeres, which may predict reduced longevity. Offspring from mothers treated with GCs during pregnancy were 8.3% lighter around birth but grew (in body mass) 17.0% faster than those from controls, whereas offspring from mothers treated with GCs during lactation grew 34.8% slower than those from controls and did not differ in body mass around birth. Treating mothers with GCs during pregnancy or lactation did not alter the oxidative stress levels or telomere lengths of their offspring. Fast-growing offspring from any of the treatment groups did not have higher oxidative stress levels or shorter telomere lengths, indicating that offspring that grew faster early in life did not exhibit oxidative costs after this period of growth. Our results indicate that elevations in maternal GCs may induce plasticity in offspring growth without long-term oxidative costs to the offspring that might result in a shortened lifespan.Summary StatementWe show that experimental increases in glucocorticoids in breeding female North American red squirrels affects offspring postnatal growth but not levels of oxidative damage and antioxidants or telomere lengths.

Comparative studies aid in our understanding of specific conditions favoring the initial evolution of different types of social behaviors, yet there is much unexplained intraspecific variation in the expression of social behavior that comparative studies have not yet addressed. The proximate causes of this individual variation in social behavior within a species have been examined in some species but its fitness consequences have been less frequently investigated. In this study, we quantified the fitness consequences of variation in the sociality of prairie voles (Microtus ochrogaster). We characterized sociality of voles in semi-natural enclosures using an automated behavioral tracking system paired with social network analyses to quantify the degree of spatial and temporal co-occurrence of different voles. We then assessed the relationship between sociality with mating success (number of different conspecifics with which an individual produced offspring) and reproductive success (total number of offspring surviving to first capture). We measured the number of social connections each individual had with all voles and only with opposite-sex voles by calculating unweighted degree through social network analyses. Both female and male voles varied in the number of social connections they had with all conspecifics and with opposite-sex conspecifics. Voles with an intermediate number of social connections with voles of both sexes had higher mating success overall. In our analyses that considered all social connections with voles of both sexes, voles with an intermediate number of social connections produced more offspring. Males with a very high or low number of social connections also had the lowest average body mass. Overall, our results suggest some limit on the fitness benefits of sociality. Although there was substantial individual-variation in our measure of vole social behavior, intermediate levels of social connections may be most favorable.

Museum genomics has transformed the field of collections-based research, opening up a range of new research directions for paleontological specimens as well as natural history specimens collected over the past few centuries. Recent work demonstrates that it is possible to characterize epigenetic markers such as DNA methylation in well preserved ancient tissues. This approach has not yet been tested in traditionally prepared natural history specimens such as dried bones and skins, the most common specimen types in vertebrate collections. In this study, we developed and tested methods to characterize cytosine methylation in dried skulls up to 76 years old. Using a combination of ddRAD and bisulphite treatment, we characterized patterns of cytosine methylation in two species of deer mouse (Peromyscus spp.) collected in the same region in Michigan in 1940, 2003, and 2013-2016. We successfully estimated methylation in specimens of all age groups, although older specimens yielded less data and showed greater interindividual variation in data yield than newer specimens. Global methylation estimates were reduced in the oldest specimens (76 years old) relative to the newest specimens (1-3 years old), which may reflect post-mortem hydrolytic deamination. Methylation was reduced in promoter regions relative to gene bodies and showed greater bimodality in autosomes relative to female X chromosomes, consistent with expectations for methylation in mammalian somatic cells. Our work demonstrates the utility of historic specimens for methylation analyses, as with genomic analyses; however, studies will need to accommodate the large variance in the quantity of data produced by older specimens.

Phenotypic plasticity—one individual’s capacity for phenotypic variation under different environments—is critical for organisms facing fluctuating conditions within their lifetime. North American red squirrels (Tamiasciurus hudsonicus) experience drastic among-year fluctuations in conspecific density. This shapes juvenile competition over vacant territories and overwinter survival. To help young cope with competition at high densities, mothers can increase offspring growth rates via a glucocorticoid-mediated maternal effect. However, this effect is only adaptive under high densities, and faster growth often comes at a cost to longevity. While experiments have demonstrated that red squirrels can adjust hormones in response to fluctuating density, the degree to which mothers differ in their ability to regulate glucocorticoids across changing densities remains unknown—little is known about within-individual plasticity in endocrine traits relative to among-individual variation. Findings from our reaction norm approach revealed significant individual variation not only in a female red squirrel’s mean endocrine phenotype, but also in endocrine plasticity in response to changes in local density. Future work on the proximate and ultimate drivers of variation in the plasticity of endocrine traits and maternal effects is needed, particularly in free-living animals experiencing fluctuating environments.

Juvenile survival to first breeding is a key life-history stage for all taxa. Survival through this period can be particularly challenging when it coincides with harsh environmental conditions such as a winter climate or food scarcity, leading to highly variable cohort survival. However, the small size and dispersive nature of juveniles generally make studying their survival more difficult. In territorial species, a key life-history event is the acquisition of a territory. A territory is expected to enhance survival, but how it does so is not often identified. We tested how the timing of territory acquisition influenced the winter survival of juvenile North American red squirrels Tamiasciurus hudsonicus, hereafter red squirrels, and how the timing of this event mediated the sources of mortality. We hypothesized that securing a territory prior to when food resources become available would reduce juvenile susceptibility to predation and climatic factors overwinter. Using 27 years of data on the survival of individually marked juvenile red squirrels, we tested how the timing of territory acquisition influenced survival, whether the population density of red squirrel predators and mean temperature overwinter were related to individual survival probability, and if territory ownership mediated these effects. Juvenile red squirrel survival was lower in the years of high predator abundance and in colder winters. Autumn territory owners were less susceptible to lynx Lynx canadensis and possibly mustelid Mustela and Martes spp., predation. Autumn territory owners had lower survival in colder winters, but surprisingly non-owners had higher survival in cold winters. Our results show how the timing of a life-history event like territory acquisition can directly affect survival and also mediate the effects of biotic and abiotic factors later in life. This engenders a better understanding of the fitness consequences of the timing of key life-history events.

Parental investment theory predicts that observed levels of parental care afforded to offspring are set by the benefits (to offspring quality and survival) relative to the costs (to parental survival or future reproduction). Although difficult to document in mammals, there is often substantial individual-variation in the amount of parental care within species. We measured the impact of individual variation in maternal care (“attentiveness” towards offspring or maternal motivation) on offspring growth and survival in a wild population of North American red squirrels (Tamiasciurus hudsonicus). We used latency to return to pups following a nest intrusion as a measure of maternal attentiveness to pups. We found this behavior to be repeatable within individuals suggesting this behavior is a personality trait or a “maternal style”. In this population, postnatal growth rate is important for pup overwinter survival. Pups from large litters grew faster if they had a highly attentive mother, indicating that maternal care behavior can mitigate the trade-off between litter size and offspring growth and potentially improve survival of pups. Additionally, more attentive mothers had slightly higher lifetime reproductive success than less attentive mothers. These results highlight important fitness effects of having a highly attentive mother and show that maternal care behavior can alter a fundamental life history trade-off between offspring quantity and quality.Lay SummaryIt pays to be attentive to your pups as a squirrel mom. In a long-term study of a wild population of North American red squirrels, we observed repeatable individual variation in maternal attentiveness towards offspring. Mothers who returned faster to pups following a nest intrusion produced faster growing pups and were able to produce larger fast-growing litters. Over their entire lifetime, attentive mothers also had more offspring recruit into the breeding population.

Summary One of the outstanding questions in evolutionary biology is the extent to which mutually beneficial interactions and kin selection can facilitate the evolution of cooperation by mitigating conflict between interacting organisms. The indirect fitness benefits gained from associating with kin are an important pathway to conflict resolution, 1 but conflict can also be resolved if individuals gain direct benefits from cooperating with one another (e.g., mutualism or reciprocity). 2 Because of the kin-structured nature of many animal societies, it has been difficult for previous research to assess the relative importance of these mechanisms. 3 , 4 , 5 However, one area that might allow for the relative roles of kin selection and mutualistic benefits to be disentangled is in the resolution of conflict over territorial space. 6 Although much research has focused on group-living species, the question of how cooperation can first be favored in solitary, territorial species remains a key question. Using 22 years of data from a population of North American red squirrels, we assessed how kinship and familiarity with neighbors affected fitness in a territorial mammal. Although living near kin did not enhance fitness, familiarity with neighbors increased survival and annual reproductive success. These fitness benefits were strong enough to compensate for the effects of aging later in life and have potential consequences for the evolution of senescence. We suggest that such substantial fitness benefits provide the opportunity for the evolution of cooperation between adversarial neighbors, offering insight into the role that mutually beneficial behaviors might play in facilitating and stabilizing social systems.

There is tremendous diversity in ageing rates and lifespan not only among taxa but within species, and particularly between the sexes. Women often live longer than men, and considerable research on this topic has revealed some of the potential biological, psychological and cultural causes of sex differences in human ageing and lifespan. However, sex differences in lifespan are widespread in nonhuman animals suggesting biology plays a prominent role in variation in ageing and lifespan. Recently, evolutionary biologists have borrowed techniques from biomedicine to identify whether similar mechanisms causing or contributing to variation in ageing and lifespan in humans and laboratory animals also operate in wild animals. Telomeres are repetitive noncoding DNA sequences capping the ends of chromosomes that are important for chromosomal stability but that can shorten during normal cell division and exposure to stress. Telomere shortening is hypothesized to directly contribute to the ageing process as once telomeres shorten to some length, the cells stop dividing and die. Men tend to have shorter telomeres and faster rates of telomere attrition with age than women, suggesting one possible biological cause of sex differences in lifespan. In this issue of Molecular Ecology, Watson et al. (2017) show that telomere lengths in wild Soay sheep are similar between females and males near the beginning of life but quickly diverge with age because males but not females showed reduced telomere lengths at older ages. The authors further show that some of the observed sex difference in telomere lengths in old age may be due to male investment in horn growth earlier in life, suggesting that sexually dimorphic allocation to traits involved in sexual selection might underlie sex differences in telomere attrition. This study provides a rare example of how biological mechanisms potentially contributing to sex differences in lifespan in humans may also operate in free-living animals. However, future studies using a longitudinal approach are necessary to confirm these observations and identify the ultimate and proximate causes of any sex differences in telomere lengths. Collaborations between evolutionary biologists and gerontologists are especially needed to identify whether telomere lengths have a causal role in ageing, particularly in natural conditions, and whether this directly contributes to sex differences in lifespan.

The phenotype of parents can have long-lasting effects on the development of offspring as well as on their behaviour, physiology, and morphology as adults. In some cases, these changes may increase offspring fitness but, in others, they can elevate parental fitness at a cost to the fitness of their offspring. Here, we show that in Kalahari meerkats (Suricata suricatta) the circulating glucocorticoid hormones of pregnant females affect the growth and cooperative behaviour of their offspring. We performed a 3-year experiment in wild meerkats to test the hypothesis that stress-mediated maternal effects reduce the potential for offspring to reproduce directly and therefore cause them to exhibit more cooperative behaviour. Daughters (but not sons) born to mothers treated with cortisol during pregnancy were smaller early in life and exhibited significantly more of two types of cooperative behaviour (pup rearing and feeding) once they were adults compared to offspring from control mothers. They also had lower measures of stress hormones as they aged, which could explain the observed increases in cooperative behaviour. Because early growth is a crucial determinant of fitness in female meerkats, our results indicate that stress-mediated maternal effects may reduce the fitness of offspring but may elevate parental fitness as a consequence of increasing the cooperative behaviour of their daughters. Significance Statement Maternal characteristics can have a profound influence on offspring, which are referred to as maternal effects. It is widely assumed that selfish maternal effects, where mothers modify offspring to benefit their own fitness and decrease the fitness of offspring, are rare but this assumption is rarely tested. We show that a stress-mediated maternal effect in meerkats altered the growth, behaviour, and stress physiology of daughters in such a way that benefitted mothers at the expense of their daughters. Daughters from stressed mothers grew more slowly early in life, which should reduce their future reproductive opportunities, but exhibited more cooperative behaviour that should directly benefit their mother. This highlights the important role of sociality in the evolution of maternal effects.

Glucocorticoids (GCs) are involved in the regulation of an animal’s energetic state. Under stressful situations, they are part of the neuroendocrine response to cope with environmental challenges. Animals react to aversive stimuli also through behavioral responses, defined as coping styles. Both in captive and wild populations, individuals differ in their behavior along a proactive–reactive continuum. Proactive animals exhibit a bold, active-explorative and social personality, whereas reactive ones are shy, less active-explorative and less social. Here, we test the hypothesis that personality traits and physiological responses to stressors covary, with more proactive individuals having a less pronounced GC stress response. In wild populations of invasive gray squirrels Sciurus carolinensis, we measured fecal glucocorticoid metabolites (FGMs), an integrated measure of circulating GCs, and 3 personality traits (activity, sociability, and exploration) derived from open field test (OFT) and mirror image stimulation (MIS) test. Gray squirrels had higher FGMs in Autumn than in Winter and males with scrotal testes had higher FGMs than nonbreeding males. Personality varied with body mass and population density. Squirrels expressed more activity-exploration at higher than at lower density and heavier squirrels had higher scores for activity-exploration than animals that weighed less. Variation in FGM concentrations was not correlated with the expression of the 3 personality traits. Hence, our results do not support a strong association between the behavioral and physiological stress responses but show that in wild populations, where animals experience varying environmental conditions, the GC endocrine response and the expression of personality are uncorrelated traits among individuals.

Acoustic signaling is an important means by which animals communicate both stable and labile characteristics. Although it is widely appreciated that vocalizations can convey information on labile state, such as fear and aggression, very few studies have experimentally examined the acoustic expression of short-term stress state. The transmission of such information about physiological state could have broad implications, potentially allowing other individuals to modify their behavior or life history traits in response to this public information. North American red squirrels (Tamiasciurus hudsonicus) produce vocalizations known as rattles that advertise territorial ownership. We examined the influence of changes in physiological stress state on rattle acoustic structure through the application of a stressor (trapping and handling the squirrels) and by provisioning squirrels with exogenous glucocorticoids (GCs). We characterized the acoustic structure of rattles emitted by these squirrels by measuring rattle duration, mean frequency, and entropy. Our results provide mixed evidence that rattles show a “stress signature”. When squirrels were trapped and handled, they produced rattles that were longer in duration with a higher frequency and increased entropy. However, squirrels that were administered exogenous GCs had similar rattle duration, frequency, and entropy as squirrels that received control treatments and unmanipulated (unfed) squirrels. Our results indicate that short-term stress does affect the acoustic structure of vocalizations, but elevated circulating GC levels are not solely responsible for such changes.

Interactions between organisms are ubiquitous and have important consequences for phenotypes and fitness. Individuals can even influence those they never meet, if they have extended phenotypes which mean the environments others experience are altered. North American red squirrels (Tamiasciurus hudsonicus) guard food hoards, an extended phenotype that typically outlives the individual and is almost always inherited by non relatives. Hoarding by previous owners can therefore influence subsequent owners. We found that red squirrels bred earlier and had higher lifetime fitness if the previous owner was a male. This was driven by hoarding behaviour, as males and mid-aged squirrels had the largest hoards, and these effects persisted across owners, such that if the previous owner was male or died in mid-age subsequent occupants had larger hoards. Individuals can, therefore, influence each other’s resource dependent traits and fitness without meeting via extended phenotypes, and so the past can influence contemporary population dynamics.

It is well established that circulating maternal stress hormones (glucocorticoids, GCs) can alter offspring phenotype. There is also a growing body of empirical work, within ecology and evolution, indicating that maternal GCs link the environment experienced by the mother during gestation with changes in offspring phenotype. These changes are considered to be adaptive if the maternal environment matches the offspring's environment and maladaptive if it does not. While these ideas are conceptually sound, we lack a testable framework that can be used to investigate the fitness costs and benefits of altered offspring phenotypes across relevant future environments. We present error management theory as the foundation for a framework that can be used to assess the adaptive potential of maternal stress hormones on offspring phenotype across relevant postnatal scenarios. To encourage rigorous testing of our framework, we provide field‐testable hypotheses regarding the potential adaptive role of maternal stress across a diverse array of taxa and life histories, as well as suggestions regarding how our framework might provide insight into past, present, and future research. This perspective provides an informed lens through which to design and interpret experiments on the effects of maternal stress, provides a framework for predicting and testing variation in maternal stress across and within taxa, and also highlights how rapid environmental change that induces maternal stress may lead to evolutionary traps.

Organisms can affect one another’s phenotypes when they socially interact. Indirect genetic effects occur when an individual’s phenotype is affected by genes expressed in another individual. These heritable effects can enhance or reduce adaptive potential, thereby accelerating or reversing evolutionary change. Quantifying these social effects is therefore crucial for our understanding of evolution, yet estimates of indirect genetic effects in wild animals are limited to dyadic interactions. We estimated indirect phenotypic and genetic effects, and their covariance with direct effects, for the date of spring breeding in North American red squirrels (Tamiasciurus hudsonicus) living in an array of territories of varying spatial proximity. Additionally, we estimated variance parameters and the strength of selection at low and high population densities. Social effects of neighbours on the date of spring breeding were weak at low, but stronger at high population densities. Indirect phenotypic effects accounted for a larger amount of variation in the date of breeding than direct differences among-individuals, although the genetic component to these indirect effects was not statistically significant. Nevertheless, the estimated effect size was large enough to suggest that indirect genetic effects could alter evolutionary change, resulting in less change at high densities despite stronger selection. Despite the difficulty in estimating them precisely, indirect genetic effects have clear potential to alter evolutionary trajectories in any natural systems where organisms interact.

Hormones such as glucocorticoids (colloquially referred to as “stress hormones”) have important effects on animal behavior and life history traits, yet most of this understanding has come through correlative studies. While experimental studies offer the ability to assign causality, there are important methodological concerns that are often not considered when manipulating hormones, including glucocorticoids, in wild animals. In this study, we examined how experimental elevations of cortisol concentrations in wild North American red squirrels (Tamiasciurus hudsonicus) affected their hypothalamic–pituitary–adrenal (HPA) axis reactivity, and life history traits including body mass, litter survival, and adult survival. The effects of exogenous cortisol on plasma cortisol concentrations depended on the time between treatment consumption and blood sampling. In the first nine hours after consumption of exogenous cortisol, individuals had significantly higher true baseline plasma cortisol concentrations, but adrenal gland function was impaired as indicated by their dampened response to capture and handling and to injections of adrenocorticotropic hormone compared to controls. Approximately 24 hours after consumption of exogenous cortisol, individuals had much lower plasma cortisol concentrations than controls, but adrenal function was restored. Corticosteroid binding globulin (CBG) concentrations were also significantly reduced in squirrels treated with cortisol. Despite these profound shifts in the functionality of the HPA axis, squirrel body mass, offspring survival, and adult survival were unaffected by experimental increases in cortisol concentrations. Our results highlight that even short-term experimental increases in glucocorticoids can affect adrenal gland functioning and CBG concentrations but without other side-effects.

Activation of the hypothalamic-pituitary-adrenal (HPA) axis culminates in the release of glucocorticoids (henceforth CORT), which have wide-reaching physiological effects. Three hypotheses potentially explain seasonal variation in CORT. The enabling hypothesis predicts that reproductive season CORT exceeds post-reproductive season CORT because CORT enables reproductive investment. The inhibitory hypothesis predicts the opposite because CORT can negatively affect reproductive function. The costs of reproduction hypothesis predicts that HPA axis condition declines over and following the reproductive season. We tested these hypotheses in wild male red-backed voles (Myodes rutilus) during the reproductive and post-reproductive seasons. We quantified CORT levels in response to restraint stress tests consisting of three blood samples (initial, stress-induced, and recovery). Mineralocorticoid (MR) and glucocorticoid (GR) receptor mRNA levels in the brain were also quantified over the reproductive season. Total CORT (tCORT) in the initial and stress-induced samples were greater in the post-reproductive than in the reproductive season, which supported the inhibitory hypothesis. Conversely, free CORT (fCORT) did not differ between the reproductive and post-reproductive seasons, which was counter to both the enabling and inhibitory hypotheses. Evidence for HPA axis condition decline in CORT as well as GR and MR mRNA over the reproductive season (i.e. costs of reproduction hypothesis) was mixed. Moreover, all of the parameters that showed signs of declining condition over the reproductive season did not also show signs of declining condition over the post-reproductive season suggesting that the costs resulting from reproductive investment had subsided. In conclusion, our results suggest that different aspects of the HPA axis respond differently to seasonal changes and reproductive investment.

Invasive alien species can cause extinction of native species through processes including predation, interspecific competition for resources or disease‐mediated competition. Increases in stress hormones in vertebrates may be associated with these processes and contribute to the decline in survival or reproduction of the native species. Eurasian red squirrels (Sciurus vulgaris) have gone extinct across much of the British Isles and parts of Northern Italy following the introduction of North American invasive grey squirrels (Sciurus carolinensis). We extracted glucocorticoid metabolites from faecal samples to measure whether the presence of the invasive species causes an increase in physiological stress in individuals of the native species. We show that native red squirrels in seven sites where they co‐occurred with invasive grey squirrels had glucocorticoid concentrations that were three times higher than those in five sites without the invasive species. Moreover, in a longitudinal study, stress hormones in native red squirrels increased after colonisation by grey squirrels. When we experimentally reduced the abundance of the invasive grey squirrels, the concentration of faecal glucocorticoid metabolites in co‐occurring red squirrels decreased significantly between pre‐ and postremoval periods. Hence, we found that the invasive species acts as a stressor which significantly increases the concentrations of glucocorticoids in the native species. Given that sustained elevations in glucocorticoids could reduce body growth and reproductive rate, our results are consistent with previous studies where the co‐occurrence of the invasive grey squirrel was associated with smaller size and lower reproductive output in red squirrels.

SYNOPSIS Hormonal pleiotropy occurs when a part of the endocrine system (e.g., hormone concentrations) influences the expression of two or more phenotypes. Although hormonal pleiotropy may have similar evolutionary consequences as genetic pleiotropy, most conceptual and empirical work on its putative evolutionary consequences to date has focused on identifying whether the different components of an endocrine axis (titer, receptor expression, etc.) that affect trait expression are themselves able to evolve independently from one another. This is important because if these different components evolve together, the expression of two traits affected by the same hormone may be yoked and evolve non-independently. Here, we first describe methodological approaches used to identify how hormonal pleiotropy could cause the co-evolution of performance and life history traits. We then focus on a similar but less studied concept about how hormonal pleiotropy can affect phenotypic responses to selection. If the expression of two traits is affected by the same hormone, the magnitude of the phenotypic response to selection may be exacerbated or retarded compared to the absence of this hormonal pleiotropy. We use classical concepts from quantitative genetics to discuss an approach for identifying whether hormonal pleiotropy has such evolutionary consequences using data collected from longitudinal studies of wild animals. We develop a simple quantitative genetics model to derive predictions about the conditions under which hormonal pleiotropy would affect the response to selection. We focus on performance and life history traits and how the effects of hormonal pleiotropy on the evolution of these traits depend upon the genetic correlations between the hormone and traits as well as the direction and strength of selection on the two traits. Finally, we review the literature for examples that have estimated these model parameters to characterize the studies that have or have not found support for these model predictions.

The age trajectory of reproductive performance of many iteroparous species features an early - life increase in performance followed by a late - life senescent decline. The largest contribution of lifetime reproductive success is therefore gained at the age at which reproductive performance peaks. Using long term data on North American red squirrels we show that the environmental conditions individuals encountered could cause variation among individuals in the “height” and timing of this peak, contributing to life history variation and fitness in this population that experiences irregular resource pulses. As expected, high peak effort was positively associated with lifetime reproductive output up to a high level of annual effort. Furthermore, individuals that matched their peak reproductive effort to an anticipated resource pulse gained substantial fitness benefits through recruiting more offspring over their lifetime. Individual variation in peak reproductive effort thus has strong potential to shape life history evolution by facilitating adaptation to fluctuating environments.

Individuals often interact more closely with some members of the population (e.g. offspring, siblings or group members) than they do with other individuals. This structuring of interactions can lead to multilevel natural selection, where traits expressed at the group-level influence fitness alongside individual-level traits. Such multilevel selection can alter evolutionary trajectories, yet is rarely quantified in the wild, especially for species that do not interact in clearly demarcated groups. We quantified multilevel natural selection on two traits, postnatal growth rate and birth date, in a population of North American red squirrels (Tamiasciurus hudsonicus). The strongest level of selection was typically within-acoustic social neighbourhoods (within 130m of the nest), where growing faster and being born earlier than nearby litters was key, while selection on growth rate was also apparent both within-litters and within-study areas. Higher population densities increased the strength of selection for earlier breeding, but did not influence selection on growth rates. Females experienced especially strong selection on growth rate at the within-litter level, possibly linked to the biased bequeathal of the maternal territory to daughters. Our results demonstrate the importance of considering multilevel and sex-specific selection in wild species, including those that are territorial and sexually monomorphic.Data archival: the data set is archived on Dryad (info XXX), with a five-year embargo from the date of publication.

Maternal stress can prenatally influence offspring phenotypes and there are an increasing number of ecological studies that are bringing to bear biomedical findings to natural systems. This is resulting in a shift from the perspective that maternal stress is unanimously costly, to one in which maternal stress may be beneficial to offspring. However, this adaptive perspective is in its infancy with much progress to still be made in understanding the role of maternal stress in natural systems. Our aim is to emphasize the importance of the ecological and evolutionary context within which adaptive hypotheses of maternal stress can be evaluated. We present five primary research areas where we think future research can make substantial progress: (1) understanding maternal and offspring control mechanisms that modulate exposure between maternal stress and subsequent offspring phenotype response; (2) understanding the dynamic nature of the interaction between mothers and their environment; (3) integrating offspring phenotypic responses and measuring both maternal and offspring fitness outcomes under real-life (either free-living or semi-natural) conditions; (4) empirically testing these fitness outcomes across relevant spatial and temporal environmental contexts (both pre- and post-natal environments); (5) examining the role of maternal stress effects in human-altered environments-i.e., do they limit or enhance fitness. To make progress, it is critical to understand the role of maternal stress in an ecological context and to do that, we must integrate across physiology, behavior, genetics, and evolution.

Summary Many aspects of the social environment affect hypothalamic-pituitary-adrenal (HPA) axis function and increase circulating glucocorticoid concentrations. In this review, we examine the relationships between the social environment and the function of the HPA axis in vertebrates. First, we explore the effects of the social environment on glucocorticoid secretion in territorial (primarily non-social) species, with an emphasis on the effects of variation in population density, as modified by environmental factors such as predation risk and food availability. In general, high population density or frequent territorial intrusions are associated with increased glucocorticoid secretion in a wide range of taxa, including mammals, birds, fish and reptiles, although there is considerable variability across species. Second, we consider the effects of social interactions and dominance rank on glucocorticoid secretion in social species, mostly in birds and mammals. We review studies that have detected an association between social status and glucocorticoid levels – sometimes with higher glucocorticoid levels in low-ranking individuals, and sometimes with higher glucocorticoid levels in dominant individuals. The relationship between dominance and glucocorticoid levels varies among species, populations and years, in a manner that depends on the stability of the social hierarchy, environmental conditions, the type of breeding system, and the manner in which high rank is obtained and maintained. Finally, we discuss the concept of allostasis and consider interactions between social effects and other environmental factors, noting that there is relatively little research on these interactions to date. For both non-social and social species, we identify priorities of future research. These priorities include more complete descriptions of HPA function that move beyond measurements of basal glucocorticoid concentrations (which will generally require field experiments), to studies that examine organizational effects of social stressors, that directly test the relationship between HPA function and fitness, and that examine how glucocorticoid responses affect population dynamics. Although several lines of evidence suggest that glucocorticoid responses can affect the fitness of individuals and therefore can alter the dynamics of populations, the effect of glucocorticoid responses on population dynamics remains essentially unstudied.

The relative scarcity of studies at the intersection of behavioral and population ecology is surprising given the presumed importance of behavior in density-dependent population regulation. Here we tested whether North American red squirrels (Tamiasciurus hudsonicus) adjust their behavior in response to local population density and whether they use rates of territorial vocalizations in their local neighborhood to assess density. We examined these relationships using 18 years of live trapping and 20 years of behavioral data that were collected across natural variation in local population density. To disentangle the effects of population density on behavior from those due to changes in per capita food abundance or changes in the frequency of antagonistic interactions with neighbors, we also experimentally manipulated population density with long-term food supplementation as well as perceived population density with long-term playbacks of territorial vocalizations. The frequency with which squirrels emitted territorial vocalizations was positively associated with local population density. In contrast, antagonistic physical interactions observed between squirrels and territorial intrusions were rare and the frequency of intrusions was weakly and negatively, not positively, associated with population density. Squirrels experiencing naturally and experimentally high density conditions spent less time in the nest and feeding but more time being vigilant. Similar density-dependent changes in behavior were observed in response to our manipulations of perceived population density, indicating that vocalization rates and not physical interactions or food abundance were the mechanism by which squirrels assessed and responded behaviorally to changes in local density.

Consistent individual differences in behaviour, and behavioural correlations within and across contexts, are referred to as animal personalities. These patterns of variation have been identified in many animal taxa and are likely to have important ecological and evolutionary consequences. Despite their importance, genetic and environmental sources of variation in personalities have rarely been characterized in wild populations. We used a Bayesian animal model approach to estimate genetic parameters for aggression, activity and docility in North American red squirrels (Tamiasciurus hudsonicus). We found support for low heritabilities (0.08-0.12), and cohort effects (0.07- 0.09), as well as low to moderate maternal effects (0.07-0.15) and permanent environmental effects (0.08-0.16). Finally, we found evidence of a substantial positive genetic correlation (0.68) and maternal effects correlation (0.58) between activity and aggression providing evidence of genetically based behavioural correlations in red squirrels. These results provide evidence for the presence of heritable variation in red squirrel behaviour, but also emphasize the role of other sources of variation, including maternal effects, in shaping patterns of variation and covariation in behavioural traits.

Life-history traits describe parameters associated with growth, size, survival, and reproduction. Life-history variation is a hallmark of biological diversity, yet researchers commonly observe that one of the major axes of life-history variation after controlling for body size involves trade-offs among growth, reproduction, and longevity. This persistent pattern of covariation among these specific traits has engendered a search for shared mechanisms that could constrain or facilitate production of variation in life-history strategies. Endocrine traits are one candidate mechanism that may underlie the integration of life history and other phenotypic traits. However, the vast majority of this research has been on the effects of steroid hormones such as glucocorticoids and androgens on life-history trade-offs. Here we propose an expansion of the focus on glucocorticoids and gonadal hormones and review the potential role of insulin-like growth factor-1 (IGF-1) in shaping the adaptive integration of multiple life-history traits. IGF-1 is a polypeptide metabolic hormone largely produced by the liver. We summarize a vast array of research demonstrating that IGF-1 levels are susceptible to environmental variation and that IGF-1 can have potent stimulatory effects on somatic growth and reproduction but decrease lifespan. We review the few studies in natural populations that have measured plasma IGF-1 concentrations and its associations with life-history traits or other characteristics of the organism or its environment. We focus on two case studies that found support for the hypothesis that IGF-1 mediates adaptive divergence in suites of life-history traits in response to varying ecological conditions or artificial selection. We also examine what we view as potentially fruitful avenues of research on this topic, which until now has been rarely investigated by evolutionary ecologists. We discuss how IGF-1 may facilitate adaptive plasticity in life-history strategies in response to early environmental conditions and also how selection on loci controlling IGF-1 signaling may mediate population divergence and eventual speciation. After consideration of the interactions among androgens, glucocorticoids, and IGF-1 we suggest that IGF-1 be considered a suitable candidate mechanism for mediating life-history traits. Finally, we discuss what we can learn about IGF-1 from studies in free-ranging animals. The voluminous literature in laboratory and domesticated animals documenting relationships among IGF-1, growth, reproduction, and lifespan demonstrates the potential for a number of new research questions to be asked in free-ranging animals. Examining how IGF-1 mediates life-history traits in free-ranging animals could lead to great insight into the mechanisms that influence life-history variation.

Stress responses play a key role in allowing animals to cope with change and challenge in the face of both environmental certainty and uncertainty. Measurement of glucocorticoid levels, key elements in the neuroendocrine stress axis, can give insight into an animal's well-being and can aid understanding ecological and evolutionary processes as well as conservation and management issues. We give an overview of the four main biological samples that have been utilized [blood, saliva, excreta (feces and urine), and integumentary structures (hair and feathers)], their advantages and disadvantages for use with wildlife, and some of the background and pitfalls that users must consider in interpreting their results. The matrix of choice will depend on the nature of the study and of the species, on whether one is examining the impact of acute versus chronic stressors, and on the degree of invasiveness that is possible or desirable. In some cases, more than one matrix can be measured to achieve the same ends. All require a significant degree of expertise, sometimes in obtaining the sample and always in extracting and analyzing the glucocorticoid or its metabolites. Glucocorticoid measurement is proving to be a powerful integrator of environmental stressors and of an animal's condition.

Fisher's principle explains that population sex ratio in sexually reproducing organisms is maintained at 1 : 1 owing to negative frequency-dependent selection, such that individuals of the rare sex realize greater reproductive opportunity than individuals of the more common sex until equilibrium is reached. If biasing offspring sex ratio towards the rare sex is adaptive, individuals that do so should have more grandoffspring. In a wild population of North American red squirrels (Tamiasciurus hudsonicus) that experiences fluctuations in resource abundance and population density, we show that overall across 26 years, the secondary sex ratio was 1 : 1; however, stretches of years during which adult sex ratio was biased did not yield offspring sex ratios biased towards the rare sex. Females that had litters biased towards the rare sex did not have more grandoffspring. Critically, the adult sex ratio was not temporally autocorrelated across years, thus the population sex ratio experienced by parents was independent of the population sex ratio experienced by their offspring at their primiparity. Expected fitness benefits of biasing offspring sex ratio may be masked or negated by fluctuating environments across years, which limit the predictive value of the current sex ratio.

In many cooperatively breeding animal societies, breeders outlive non-breeding subordinates, despite investing heavily in reproduction [1-3]. In eusocial insects, the extended lifespans of breeders arise from specialized slowed aging profiles [1], prompting suggestions that reproduction and dominance similarly defer aging in cooperatively breeding vertebrates, too [4-6]. Although lacking the permanent castes of eusocial insects, breeders of vertebrate societies could delay aging via phenotypic plasticity (similar rank-related changes occur in growth, neuroendocrinology, and behavior [7-10]), and such plastic deferment of aging may reveal novel targets for preventing aging-related diseases [11]. Here, we investigate whether breeding dominants exhibit extended longevity and delayed age-related physiological declines in wild cooperatively breeding meerkats. We show that dominants outlive subordinates but exhibit faster telomere attrition (a marker of cellular senescence and hallmark of aging [12]) and that in dominants (but not subordinates), rapid telomere attrition is associated with mortality. Our findings further suggest that, rather than resulting from specialized aging profiles, differences in longevity between dominants and subordinates are driven by subordinate dispersal forays, which become exponentially more frequent with age and increase subordinate mortality. These results highlight the need to critically examine the causes of rank-related longevity contrasts in other cooperatively breeding vertebrates, including social mole-rats, where they are currently attributed to specialized aging profiles in dominants [4].

Patterns in stress hormone (glucocorticoid: GC) levels and their relationship to reproductive condition in natural populations are rarely investigated. In this study, we (1) validate an enzyme-immunoassay to measure fecal cortisol metabolite (FCM) levels in North American red squirrels (Tamiasciurus hudsonicus), and (2) examine relationships between FCM levels and reproductive condition in a free-ranging red squirrel population. Injected radiolabeled cortisol was entirely metabolized and excreted in both the urine (mean ± SE; 70.3 ± 0.02%) and feces (29.7 ± 0.02%), with a lag time to peak excretion in the feces of 10.9 ± 2.3 h. Our antibody reacted with several cortisol metabolites, and an adrenocorticotropic injection significantly increased FCM levels above baseline levels at 8 h post-injection. Relative to baseline levels, manipulation by handling also tended to increase FCM levels at 8 h post-manipulation, but this difference was not significant. FCM levels did not differ significantly between samples frozen immediately and 5 h after collection. Reproductive condition significantly affected FCM levels in free-ranging females (pregnant > lactating > post-lactating > non-breeding) but not males (scrotal testes vs. abdominal testes). Among females with known parturition dates, FCM levels increased during gestation, peaked at parturition, and declined during lactation. The difference between pregnant and lactating females was therefore dependent upon when the fecal samples were obtained during these periods, suggesting caution in categorizing reproductive stages. This study demonstrates the utility of fecal hormone metabolite assays to document patterns of glucocorticoid levels in free-ranging animals.