L. Michael Romero

Seasonal changes in plasma glucocorticoid concentrations in free-living vertebrates

The vertebrate stress response helps animals respond to environmental dangers such as predators or storms. An important component of the stress response is glucocorticoid (GC) release, resulting from activation of the hypothalamic-pituitary-adrenal axis. After release, GCs induce a variety of behavioral and physiological changes that presumably help the animal respond appropriately to the situation. Consequently, GC secretion is often considered an obligatory response to stressful situations. Evidence now indicates, however, that free-living species from many taxa can seasonally modulate GC release. In other words, the magnitudes of both unstressed and stressed GC concentrations change depending upon the time of year. This review examines the growing evidence that GC concentrations in free-living reptiles, amphibians, and birds, but not mammals, are commonly elevated during the breeding season. This evidence is then used to test three hypotheses with different focuses on GCs energetic or behavioral effects, as well as on GCs role in preparing the animal for subsequent stressors. These hypotheses attempt to place annual GC rhythms into a physiological or behavioral context. Integrating seasonal differences in GC concentrations with either different physiological states or different life history stages provides clues to a new understanding of how GCs actually help in survival during stress. Consequently, understanding seasonal modulation of GC release has far-reaching importance for both the physiology of the stress response and the short-term survival of individual animals.

The Reactive Scope Model - a new model integrating homeostasis, allostasis, and stress.

Allostasis, the concept of maintaining stability through change, has been proposed as a term and a model to replace the ambiguous term of stress, the concept of adequately or inadequately coping with threatening or unpredictable environmental stimuli. However, both the term allostasis and its underlying model have generated criticism. Here we propose the Reactive Scope Model, an alternate graphical model that builds on the strengths of allostasis and traditional concepts of stress yet addresses many of the criticisms. The basic model proposes divergent effects in four ranges for the concentrations or levels of various physiological mediators involved in responding to stress. (1) Predictive Homeostasis is the range encompassing circadian and seasonal variation - the concentrations/levels needed to respond to predictable environmental changes. (2) Reactive Homeostasis is the range of the mediator needed to respond to unpredictable or threatening environmental changes. Together, Predictive and Reactive Homeostasis comprise the normal reactive scope of the mediator for that individual. Concentrations/levels above the Reactive Homeostasis range is (3) Homeostatic Overload, and concentrations/levels below the Predictive Homeostasis range is (4) Homeostatic Failure. These two ranges represent concentrations/levels with pathological effects and are not compatible with long-term (Homeostatic Overload) or short-term (Homeostatic Failure) health. Wear and tear is the concept that there is a cost to maintaining physiological systems in the Reactive Homeostasis range, so that over time these systems gradually lose their ability to counteract threatening and unpredictable stimuli. Wear and tear can be modeled by a decrease in the threshold between Reactive Homeostasis and Homeostatic Overload, i.e. a decrease in reactive scope. This basic model can then be modified by altering the threshold between Reactive Homeostasis and Homeostatic Overload to help understand how an individuals response to environmental stressors can differ depending upon factors such as prior stressors, dominance status, and early life experience. We illustrate the benefits of the Reactive Scope Model and contrast it with the traditional model and with allostasis in the context of chronic malnutrition, changes in social status, and changes in stress responses due to early life experiences. The Reactive Scope Model, as an extension of allostasis, should be useful to both biomedical researchers studying laboratory animals and humans, as well as ecologists studying stress in free-living animals.

CORTICOSTERONE RESPONSES IN WILD BIRDS: THE IMPORTANCE OF RAPID INITIAL SAMPLING

Abstract Corticosterone concentrations in birds usually rise in response to capture and handling, and it is often assumed that this change is predictable. We tested this assumption by leaving Gambels White-crowned Sparrows (Zonotrichia leucophrys gambelii), House Sparrows (Passer domesticus), and Lapland Longspurs (Calcarius lapponicus) in nets or traps for 15 min following capture and comparing their corticosterone response over the next 60 min with birds removed immediately. White-crowned Sparrows and House Sparrows left in mist nets for 15 min and then bled had significantly elevated corticosterone concentrations compared to controls that were immediately removed from the net and bled. Corticosterone concentrations over the next 45 min of handling and restraint were similar between groups. In another experiment, White-crowned Sparrows and Lapland Longspurs were captured using seed-baited Potter traps. The corticosterone response of White-crowned Sparrows left in the trap for 15 min did not differ from White-crowned Sparrows removed immediately. Leaving Lapland Longspurs in the trap had no effect in the initial 10 min of handling and restraint, but at 30 and 60 min these birds had significantly lower corticosterone concentrations than longspurs removed immediately from the trap. These data indicate that failing to immediately remove birds from nets or traps can alter the corticosterone response to subsequent stressful stimuli in unpredictable ways. This result emphasizes that the elapsed time from capture is a critical variable in assessing stress responses in free-living birds. Respuestas de los Niveles de Corticosterona en Aves Silvestres: La Importancia de un Muestreo Inicial Inmediato Resumen. Las concentraciones de corticosterona en las aves usualmente aumentan en respuesta a la captura y manipulación, y muchas veces se supone que estos cambios son predecibles. Pusimos a prueba esta suposición reteniendo individuos de las especies Zonotrichia leucophrys gambelii, Passer domesticus y Calcarius lapponicus en redes o trampas durante los 15 minutos subsecuentes a la captura y comparamos sus respuestas en los niveles de corticosterona durante los siguientes 60 minutos con las de individuos removidos inmediatamente de las trampas y redes. Las muestras de sangre de Z. l. gambelii y P. domesticus que fueron obtenidas después de 15 minutos de retención en las redes tuvieron niveles de corticosterona significativamente más altos que las de los individuos control obtenidas inmediatamente después de la captura. Durante los 45 minutos siguientes de manipulación y captura, las concentraciones de corticosterona fueron similares entre los dos grupos. En otro experimento, Z. l. gambelii y C. lapponicus fueron capturados mediante trampas “Potter” cebadas con semillas. La respuesta en los niveles de corticosterona de Z. l. gambelii no fue diferente entre individuos retenidos en las trampas por 15 minutos e individuos removidos inmediatamente. Para individuos de C. lapponicus retenidos en las trampas no hubo un efecto durante los 10 minutos iniciales de manipulación y captura, pero a los 30 y 60 minutos estas aves tuvieron concentraciones significativamente menores que los individuos removidos inmediatamente. Estos resultados indican que al no remover inmediatamente a las aves de las redes o trampas, las respuestas en los niveles de corticosterona a estímulos estresantes pueden verse alteradas de una manera impredecible. Estos resultados enfatizan que en aves silvestres, el lapso de tiempo desde la captura es una variable crítica en la determinación de las respuestas al estrés.

Season and Migration Alters the Corticosterone Response to Capture and Handling in an Arctic Migrant, the White-Crowned Sparrow (Zonotrichia leucophrys gambelii)

The Gambels race of white-crowned sparrow (Zonotrichia leucophrys gambelii) migrates each year from their wintering grounds in the Southwestern United States to the Arctic to breed. Associated with this migration is a change in both the nonstressed and the stress-induced levels of circulating plasma corticosterone. Birds were captured at two sites on their wintering grounds (New Mexico and Arizona). Although stress-induced corticosterone levels were elevated at each site compared to nonstressed levels, there were no differences between wintering sites. These were also similar to levels in birds caught in Washington state during fall migration. In contrast, nonstressed corticosterone values were greatly elevated in birds on their breeding grounds in Alaska and similar to stress-induced levels in wintering birds. Corticosterone levels rose even further in response to the stress of capture and handling in breeding birds. These augmented corticosterone levels during breeding were not associated with weather. Both nonstressed and stress-induced corticosterone levels were similar in birds caught on their breeding grounds on two different years, one with temperatures during mid-May of approximately 0 degree C during a snow storm and the other with temperatures in the mid-20s. These results suggest that seasonal physiological changes, and not local conditions, underlie seasonal changes in corticosterone levels. Furthermore, birds caught in Washington state during spring migration had intermediate levels of both nonstressed and stress-induced corticosterone. Corticosterone release may function differently during spring and fall migrations.

Hypothalamic-pituitary-adrenal axis changes allow seasonal modulation of corticosterone in a bird

We examined possible mechanisms underlying seasonal stress modulation in Lapland longspurs ( Calcarius lapponicus), a species that breeds and molts (the energetically costly replacement of feathers) in the Alaskan Arctic. Free-living Lapland longspurs show dramatically reduced maximal corticosterone release during molt compared with the breeding season, an effect lost in captive birds. Neither changes in corticosterone binding proteins nor the overall condition of the bird (assessed by weight and fat storage) can explain different seasonal corticosterone responses. Adrenal insensitivity also does not fully explain reduced maximal output because exogenous ACTH enhanced corticosterone release during molt. Exogenous ACTH in molting birds, however, cannot stimulate corticosterone to stress-induced levels during breeding, implying reduced adrenal capacity. Lapland longspur pituitaries appeared to respond to exogenous corticotropin-releasing factor, arginine vasotocin, and mesotocin (the avian equivalents of arginine vasopressin and oxytocin) during molt, suggesting that a mechanism upstream of the pituitary blunts corticosterone release. Taken together, these results indicate that seasonal modulation of corticosterone release in this species is controlled at multiple sites in the hypothalamic-pituitary-adrenal axis.

Exposure to tourism reduces stress-induced corticosterone levels in Galápagos marine iguanas

Unstressed and stress-induced plasma corticosterone levels in Galapagos marine iguanas (Amblyrhynchus cristatus) were compared in animals from a site heavily exposed to tourism to animals from a site undisturbed by humans. Initial corticosterone levels not only did not differ between the two groups, but they were 50% of levels in iguanas known to be chronically stressed. These data suggest that iguanas in tourist areas are not chronically stressed. Both groups of iguanas exhibited elevated corticosterone levels after 30 min of capture and restraint, indicating that they can physiologically respond to stressful stimuli. The stress response was lower, however, at the tourist site. This result indicates that iguanas are physiologically affected by tourism, although it is presently unknown whether these changes are ultimately beneficial or harmful.

Impacts of varying habitat quality on the physiological stress of spotted salamanders (Ambystoma maculatum)

We sampled blood from free-living spotted salamanders (Ambystoma maculatum) to test whether differences in the concentrations of a stress hormone (corticosterone) were associated with different qualities of breeding and migration habitat. Spotted salamanders are forest specialists that migrate to vernal pools to breed, and upland habitat degradation may have sub-lethal effects on animals that lead to population declines. An individual’s level of physiological stress may function as a biomonitor for sub-lethal effects, and thus as a biomonitor for ecosystem quality. We compared unstressed (baseline) and stress-induced corticosterone concentrations in spotted salamanders: (1) at sites that differed in amount of forest loss; (2) during breeding migration across forest habitat versus pavement; (3) in microhabitats that varied in soil drainage and canopy cover. Removal of large amounts of terrestrial habitat surrounding a breeding pond was correlated with lower baseline (in males) and stress-induced corticosterone concentrations, which may indicate healthy individuals with a reduced ability to respond to additional stress or individuals experiencing chronic stress. Male salamanders migrating across pavement had elevated baseline corticosterone concentrations compared to animals migrating through a forest, consistent with an acute stress response. However, concentrations of corticosterone did not differ between individuals in microhabitats with canopy cover and well-drained soil versus those in microhabitats with no canopy cover and/or swampy soil. This endocrinological technique may be one useful measure of a population’s health, helping to identify populations where further ecological study is recommended to evaluate conservation concerns.

Identifying hormonal habituation in field studies of stress

Habituation is a term commonly used to explain a decrement in response intensity to a repeated stimulus or set of stimuli. In the stress literature, hormonal habituation is often used to describe a situation where an individual has learned to perceive a repeated stressor as innocuous, and thus the intensity of the release of hormonal stress mediators reduces over time. Consequently, a habituated individual is not considered stressed. There are, however, situations where an individual may be chronically stressed despite a reduction in the response intensity of hormonal stress mediators to a repeated stimulus. These alternative explanations are rarely considered in field studies even though a false conclusion that an individual has habituated (i.e., is not stressed) may lead to false conclusions regarding the animals overall physiology and health. The present paper provides four alternative explanations for an observed attenuation in the response of hormonal stress mediators to a repeated stimulus or set of stimuli which lead to six criteria that define habituation in a field context. Furthermore, we propose four diagnostic tests to help distinguish hormonal habituation from these alternative explanations in field studies. These tests will help identify hormonal habituation in free-living animals and prevent potential problems of falsely describing an individual or population of individuals as habituated.

Alterations in hypothalamic–pituitary–adrenal function associated with captivity in Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii)

Gambels white-crowned sparrows were captured and brought into captivity in order to study seasonal changes in the function of the hypothalamic-pituitary-adrenal (HPA) axis in captive birds. 30 min of restraint elicited a rise in corticosterone titers that varied depending upon the season and physiological state of the birds. Restraint elevated corticosterone titers significantly more during the fall (within 2 weeks of capture from the wild) than during either the winter or during a prealternate or prebasic molt. We also examined what changes in the HPA axis could account for altered corticosterone levels. Exogenous ACTH significantly elevated corticosterone levels beyond the response to restraint during the fall, indicating a dramatic enhancement of the adrenals ability to secrete corticosterone. Exogenous ACTH was ineffective at other times, suggesting that the adrenals ability to release corticosterone often limits circulating levels. We further inferred the pituitarys ACTH secretory ability by injecting exogenous corticotrophin-releasing factor, arginine vasotocin, and mesotocin and measuring corticosterone release. Pituitaries failed to respond to any exogenous releasing factor during the fall, suggesting that the pituitary may be the site in the HPA axis regulating corticosterone release at this time. When compared to wild-caught birds, these results suggest that captivity alters both adrenal and pituitary function during restraint in white-crowned sparrows, and that this change depends upon the season and/or physiological state of the animal. Captivity thus appears to have a profound affect on the function of the HPA axis, and these results reiterate the caution that must be used to extrapolate laboratory data to field conditions.

Stress physiology as a predictor of survival in Galapagos marine iguanas

Although glucocorticoid hormones are considered important physiological regulators for surviving adverse environmental stimuli (stressors), evidence for such a role is sparse and usually extrapolated from glucocorticoid effects under laboratory, short-term and/or non-emergency conditions. Galápagos marine iguanas (Amblyrhynchus cristatus) provide an excellent model for determining the ultimate function of a glucocorticoid response because susceptibility to starvation induced by El Niño conditions is essentially their only major natural stressor. In a prospective study, we captured 98 adult male marine iguanas and assessed four major components of their glucocorticoid response: baseline corticosterone titres; corticosterone responses to acute stressors (capture and handling); the maximal capacity to secrete corticosterone (via adrenocorticotropin injection); and the ability to terminate corticosterone responses (negative feedback). Several months after collecting initial measurements, weak El Niño conditions affected the Galápagos and 23 iguanas died. The dead iguanas were typified by a reduced efficacy of negative feedback (i.e. poorer post-stress suppression of corticosterone release) compared with surviving iguanas. We found no prior differences between dead and alive iguanas in baseline corticosterone concentrations, responses to acute stressors, nor in capacity to respond. These data suggest that a greater ability to terminate a stress response conferred a survival advantage during starvation.