The effects of early-life and intergenerational stress on the brain.

Stress experienced during ontogeny can have profound effects on the adult phenotype. However, stress can also be experienced intergenerationally, where an offspring's phenotype can be moulded by stress experienced by the parents. Although early-life and intergenerational stress can alter anatomy, physiology, and behaviour, nothing is known about how these stress contexts interact to affect the neural phenotype. Here, we examined how early-life and intergenerational stress affect the brain in eastern fence lizards (Sceloporus undulatus). Some lizard populations co-occur with predatory fire ants, and stress from fire ant attacks exerts intergenerational physiological and behavioural changes in lizards. However, it is unclear if intergenerational stress, or the interaction between intergenerational and early-life stress, modulates the brain. To test this, we captured gravid females from fire ant invaded and uninvaded populations, and subjected offspring to three early-life stress treatments: (1) fire ant attack, (2) corticosterone, or (3) a control. Corticosterone and fire ant attack decreased some aspects of the neural phenotype while population of origin and the interaction of early-life stress and population had no effects on the brain. These results suggest that early-life stressors may better predict adult brain variation than intergenerational stress in this species.

Obtaining plasma to measure baseline corticosterone concentrations in reptiles: How quick is quick enough?

There is growing interest in the use of glucocorticoid (GC) hormones to understand how wild animals respond to environmental challenges. Blood is the best medium for obtaining information about recent GC levels; however, obtaining blood requires restraint and can therefore be stressful and affect GC levels. There is a delay in GCs entering blood, and it is assumed that blood obtained within 3 min of first disturbing an animal reflects a baseline level of GCs, based largely on studies of birds and mammals. Here we present data on the timing of changes in the principle reptile GC, corticosterone (CORT), in four reptile species for which blood was taken within a range of times 11 min or less after first disturbance. Changes in CORT were observed in cottonmouths (Agkistrodon piscivorus; 4 min after first disturbance), rattlesnakes (Crotalus oreganus; 2 min 30 s), and rock iguanas (Cyclura cychlura; 2 min 44 s), but fence lizards (Sceloporus undulatus) did not exhibit a change within their 10-min sampling period. In both snake species, samples taken up to 3-7 min after CORT began to increase still had lower CORT concentrations than after exposure to a standard restraint stressor. The "3-min rule" appears broadly applicable as a guide for avoiding increases in plasma CORT due to handling and sampling in reptiles, but the time period in which to obtain true baseline CORT may need to be shorter in some species (rattlesnakes, rock iguanas), and may be unnecessarily limiting for others (cottonmouths, fence lizards).

Population history with invasive predators predicts innate immune function response to early-life glucocorticoid exposure in lizards.

Early-life stress can suppress immune function, but it is unclear whether transgenerational stress exposure modulates the immune consequences of early stress. In populations where, historically, the immune system is frequently activated, e.g. persistent stressors that cause injury, it may be maladaptive to suppress immune function after early-life stress. Thus, the relationship between early-life stress and immune function may vary with population-level historical stressor exposure. We collected gravid fence lizards (Sceloporus undulatus) from populations that naturally differ in long-term exposure to invasive fire ants (Solenopsis invicta). We manipulated early-life stress in the resulting offspring via weekly exposure to fire ants, application of the stress-relevant hormone corticosterone or control treatment from 2 to 43 weeks of age. We quantified adult immune function in these offspring with baseline and antigen-induced hemagglutination and plasma bacterial killing ability. Early-life corticosterone exposure suppressed baseline hemagglutination in offspring of lizards from populations not exposed to fire ants but enhanced hemagglutination in those from populations that were exposed to fire ants. This enhancement may prepare lizards for high rates of wounding, toxin exposure and infection associated with fire ant attack. Adult bacterial killing ability and hemagglutination were not affected by early-life exposure to fire ants, but the latter was higher in offspring of lizards from invaded sites. A population's history of persistent stress may thus alter individual long-term immunological responses to early-life stressors. Further consideration of historical stressor exposure (type and duration) may be important to better understand how early-life stressors affect adult physiology.

Presence of an invasive species reverses latitudinal clines of multiple traits in a native species.

Understanding the processes driving formation and maintenance of latitudinal clines has become increasingly important in light of accelerating global change. Many studies have focused on the role of abiotic factors, especially temperature, in generating clines, but biotic factors, including the introduction of non-native species, may also drive clinal variation. We assessed the impact of invasion by predatory fire ants on latitudinal clines in multiple fitness-relevant traits-morphology, physiological stress responsiveness, and antipredator behavior-in a native fence lizard. In areas invaded by fire ants, a latitudinal cline in morphology is opposite both the cline found in museum specimens from historical populations across the species' full latitudinal range and that found in current populations uninvaded by fire ants. Similarly, clines in stress-relevant hormone response to a stressor and in antipredator behavior differ significantly between the portions of the fence lizard range invaded and uninvaded by fire ants. Changes in these traits within fire ant-invaded areas are adaptive and together support increased and more effective antipredator behavior that allows escape from attacks by this invasive predator. However, these changes may mismatch lizards to the environments under which they historically evolved. This research shows that novel biotic pressures can alter latitudinal clines in multiple traits within a single species on ecological timescales. As global change intensifies, a greater understanding of novel abiotic and biotic pressures and how affected organisms adapt to them across space and time will be central to predicting and managing our changing environment.

Ancestry trumps experience: Transgenerational but not early life stress affects the adult physiological stress response.

Exposure to stressors can affect an organism's physiology and behavior as well as that of its descendants (e.g. through maternal effects, epigenetics, and/or selection). We examined the relative influence of early life vs. transgenerational stress exposure on adult stress physiology in a species that has populations with and without ancestral exposure to an invasive predator. We raised offspring of eastern fence lizards (Sceloporus undulatus) from sites historically invaded (high stress) or uninvaded (low stress) by predatory fire ants (Solenopsis invicta) and determined how this different transgenerational exposure to stress interacted with the effects of early life stress exposure to influence the physiological stress response in adulthood. Offspring from these high- and low-stress populations were exposed weekly to either sub-lethal attack by fire ants (an ecologically relevant stressor), topical treatment with a physiologically-appropriate dose of the stress-relevant hormone, corticosterone (CORT), or a control treatment from 2 to 43weeks of age. Several months after treatments ended, we quantified plasma CORT concentrations at baseline and following restraint, exposure to fire ants, and adrenocorticotropic hormone (ACTH) injection. Exposure to fire ants or CORT during early life did not affect lizard stress physiology in adulthood. However, offspring of lizards from populations that had experienced multiple generations of fire ant-invasion exhibited more robust adult CORT responses to restraint and ACTH-injection compared to offspring from uninvaded populations. Together, these results indicate that transgenerational stress history may be at least as important, if not more important, than early life stress in affecting adult physiological stress responses.

How do duration, frequency, and intensity of exogenous CORT elevation affect immune outcomes of stress?

Stress is typically characterized as "acute" (lasting from minutes to hours) or "chronic" (lasting from days to months). These terms are of limited use as they are inconsistently used and only encompass one aspect of the stressor (duration). Short and long duration stress are generally thought to produce specific outcomes (e.g. acute stress enhances while chronic stress suppresses immune function). We propose that aspects of stress other than duration, such as frequency and intensity, are important in determining its outcome. We experimentally manipulated duration, frequency, and intensity of application of exogenous corticosterone, CORT, in Sceloporus undulatus (Eastern fence lizards) and measured the immune outcomes. Our findings reveal that immune outcomes of stress are not easily predicted from the average amount or duration of CORT elevation, but that intensity plays an important role. Although three of our treatments received the same average amount of CORT, they produced different effects on immune outcomes (hemagglutination). As predicted by the literature, short-duration exposure to low-dose CORT enhanced hemagglutination; however, short-duration exposure to high-dose CORT suppressed hemagglutination, suggesting that stressor intensity affects immune outcomes of stress. While both are traditionally termed "acute" based on duration, these treatments produced different immune outcomes. Long-duration ("chronic") exposure to CORT did not produce the expected suppression of hemagglutination. Frequency of CORT application did not alter immune outcomes at low intensities. These results highlight the need to quantify more than just the duration of a stressor if we are to understand and manage the ecological consequences of stress. Specifically, we should consider stressor frequency and intensity, as well as duration, for a more complete characterization and understanding of stress.

Immune responses of eastern fence lizards (Sceloporus undulatus) to repeated acute elevation of corticosterone.

Prolonged elevations of glucocorticoids due to long-duration (chronic) stress can suppress immune function. It is unclear, however, how natural stressors that result in repeated short-duration (acute) stress, such as frequent agonistic social encounters or predator attacks, fit into our current understanding of the immune consequences of stress. Since these types of stressors may activate the immune system due to increased risk of injury, immune suppression may be reduced at sites where individuals are repeatedly exposed to potentially damaging stressors. We tested whether repeated acute elevation of corticosterone (CORT, a glucocorticoid) suppresses immune function in eastern fence lizards (Sceloporus undulatus), and whether this effect varies between lizards from high-stress (high baseline CORT, invaded by predatory fire ants) and low-stress (low baseline CORT, uninvaded) sites. Lizards treated daily with exogenous CORT showed higher hemagglutination of novel proteins by their plasma (a test of constitutive humoral immunity) than control lizards, a pattern that was consistent across sites. There was no significant effect of CORT treatment on bacterial killing ability of plasma. These results suggest that repeated elevations of CORT, which are common in nature, produce immune effects more typical of those expected at the acute end of the acute-chronic spectrum and provide no evidence of modulated consequences of elevated CORT in animals from high-stress sites.

Tradeoffs, competition, and coexistence in eastern deciduous forest ant communities.

Ecologists have long sought to explain the coexistence of multiple potentially competing species in local assemblages. This is especially challenging in species-rich assemblages in which interspecific competition is intense, as it often is in ant assemblages. As a result, a suite of mechanisms has been proposed to explain coexistence among potentially competing ant species: the dominance-discovery tradeoff, the dominance-thermal tolerance tradeoff, spatial segregation, temperature-based niche partitioning, and temporal niche partitioning. Through a series of observations and experiments, we examined a deciduous forest ant assemblage in eastern North America for the signature of each of these coexistence mechanisms. We failed to detect evidence for any of the commonly suggested mechanisms of coexistence, with one notable exception: ant species appear to temporally partition foraging times such that behaviourally dominant species foraged more intensely at night, while foraging by subdominant species peaked during the day. Our work, though focused on a single assemblage, indicates that many of the commonly cited mechanisms of coexistence may not be general to all ant assemblages. However, temporal segregation may play a role in promoting coexistence among ant species in at least some ecosystems, as it does in many other organisms.

The impacts of invaders: basal and acute stress glucocorticoid profiles and immune function in native lizards threatened by invasive ants.

As anthropogenic stressors increase exponentially in the coming decades, native vertebrates will likely face increasing threats from these novel challenges. The success or failure of the primary physiological mediator of these stressors--the HPA axis--will likely involve numerous and chaotic outcomes. Among the most challenging of these new threats are invasive species. These have the capacity to simultaneously challenge the HPA axis and the immune system as they are often associated with, or the cause of, emerging infectious diseases, and energetic tradeoffs with the HPA response can have immunosuppressive effects. To determine the effects of invasive species on the vertebrate GC response to a novel stressor, and on immunity, we examined the effects of invasive fire ants on native lizards, comparing lizards from sites with long histories with fire ants to those outside the invasion zone. We demonstrated higher baseline and acute stress (captive restraint) CORT levels in lizards from within fire ant invaded areas; females are more strongly affected than males, suggesting context-specific effects of invasion. We found no effect of fire ant invasion on the immune parameters we measured (complement bacterial lysis and antibody hemagglutination) with the exception of ectoparasite infestation. Mites were far less prevalent on lizards within fire ant invaded sites, suggesting fire ants may actually benefit lizards in this regard. This study suggests that invasive species may impose physiological stress on native vertebrates, but that the consequences of this stress may be complicated and unpredictable.