Free Corticosterone Research Articles

Distinct, Time-Dependent Effects of Voluntary Exercise on Circadian and Ultradian Rhythms and Stress Responses of Free Corticosterone in the Rat Hippocampus

Previous work has shown that allowing rats to voluntarily exercise in a running wheel for 4 wk modifies the hypothalamic-pituitary-adrenal axis and behavioral coping responses to stress. To investigate whether long-term voluntary exercise would also affect the free, biologically active fraction of corticosterone in the brain, we conducted an in vivo microdialysis study in the hippocampus of rats. We monitored both the baseline circadian and ultradian patterns of corticosterone in hippocampus dialysates over the diurnal cycle and the responses to forced swim and novelty stress at different stages of exercise. Exercise for 1 d, 2 d, or 1 wk did not affect baseline circadian and ultradian pulse parameters or stress-induced hippocampal free corticosterone concentrations suggesting that acute or short-term periods of exercise do not affect baseline and stress-induced hormone levels. Baseline hormone parameters in 4 wk exercised rats, however, showed significantly increased pulse amplitudes (+108%) and mean free corticosterone levels (+42%) between 1500 and 2100 h but not between 0900 and 1500 h. Surprisingly, although our previous work showed substantial changes in stress-evoked plasma (total) corticosterone responses in long-term exercised animals, no differences in stress-induced hippocampal free hormone responses could be observed between exercised and sedentary animals. This lack of differences was not caused by compensatory changes in plasma corticosteroid-binding-globulin binding levels in exercising rats. Thus, long-term exercising rats show anticipatory increases in glucocorticoid output before the start of the active phase. These rats also reveal the putative existence of a containment mechanism preventing overexposure of the brain to glucocorticoid hormones.

Regulation of free corticosterone and CBG capacity under different environmental conditions in altricial nestlings

The concentration of circulating glucocorticoids is regulated in response to environmental and endogenous conditions. Total circulating corticosterone, the main glucocorticoid in birds, consists of a fraction which is bound to corticosterone-binding globulins (CBG) and a free fraction. There is increasing evidence that the environment modulates free corticosterone levels through varying the concentration of CBG, but experimental evidence is lacking. To test the hypothesis that the regulation of chronic stress in response to endogenous and environmental conditions involves variation in both corticosterone release and CBG capacity, we performed an experiment with barn owl ( Tyto alba) nestlings in two different years with pronounced differences in environmental conditions and in nestlings experimentally fed ad libitum. In half of the individuals we implanted a corticosterone-releasing pellet to artificially increase corticosterone levels and in the other half we implanted a placebo pellet. We then repeatedly collected blood samples to measure the change in total and free corticosterone levels as well as CBG capacity. The increase in circulating total corticosterone after artificial corticosterone administration varied with environmental conditions and with the food regime of the nestlings. The highest total corticosterone levels were found in nestlings growing up in poor environmental conditions and the lowest in ad libitum fed nestlings. CBG was highest in the year with poor environmental conditions, so that, contrary to total corticosterone, free corticosterone levels were low under poor environmental conditions. When nestlings were fed ad libitum total corticosterone, CBG and free corticosterone did not increase when administering corticosterone. These results suggest that depending on the individual history an animal experienced during development the HPA-axis is regulated differently.

The Combination of a High-fat Diet and Chronic Stress Aggravates Insulin Resistance in Wistar Male Rats

We observed the effects of the combination of a high-fat diet and chronic stress on insulin resistance. Male Wistar rats were fed on either a control or a high-fat diet and given chronic stress with the electric foot shock or not for 10 weeks. After checking the glucose infusion rate (GIR) and the HOMA-IR index, the results showed that the three groups all revealed insulin resistance with increased free fatty acid (FFA), adrenocorticotropic hormone (ACTH) and corticosterone in the serum, in addition to increased tumour necrosis factor alpha (TNF-alpha) in the serum and adipose tissue, and decreased density of high affinity receptors (R1) and expression of peroxisome proliferator-activated receptor-alpha (PPARalpha) mRNA in the hepatocytes as compared with the control, but the highest alteration on aforementioned parameters revealed in the chronic stress fed with a high-fat diet. Significant interactions between high-fat diet and chronic stress were revealed on GIR, HOMA-IR index, FFA, ACTH, corticosterone, TNF-alpha (in adipose tissue) and R1. These observations strongly suggest that a combination of a high-fat diet and chronic stress can produce a synergic effect on aggravating insulin resistance associated with the abnormal hypothalamic-pituitary-adrenocortical axis, endocrine abnormality of the adipose tissue, and pathological changes of the liver.

The Ultradian and Circadian Rhythms of Free Corticosterone in the Brain are Not Affected by Gender: An In Vivo Microdialysis Study in Wistar Rats

Recently, we described that free corticosterone levels in the brain of male Wistar rats, as assessed by in vivo microdialysis, show an ultradian rhythm with a pulse frequency of 1.2 pulses/h. To establish whether gender influences brain free corticosterone rhythms, we studied free corticosterone levels in the female Wistar rat under baseline and stressful conditions using microdialysis in the hippocampus. Analysis of the data with the PULSAR algorithm revealed that hippocampal free corticosterone levels show a clear ultradian pattern in female rats with a pulse frequency of 1.16+/-0.05 pulses/h between 09.00 h and 21.00 h. Further analysis showed that the pulse amplitude is significantly higher during the late afternoon/early night (15.00-21.00 h) than during the morning/early afternoon (09.00-15.00 h) phase (0.13+/-0.03 versus 0.07+/-0.01 microg/dl, respectively, P < 0.05). Pulse characteristics were extremely reproducible as demonstrated by the almost identical pulse parameters derived from two consecutive 24-h periods [pulse frequency: 1.13+/-0.09 and 1.19+/-0.08 pulses/h; pulse amplitude: 0.11+/-0.05 and 0.10+/-0.02 microg/dl for day 1 and day 2 (09.00-21.00 h) respectively, P > 0.05]. Both exposure to a novel environment and forced swim stress increased hippocampal free corticosterone levels. However, the stress-induced rise reached higher levels and was more prolonged after forced swimming (area under the curve: 46.84+/-9.25 and 12.08+/-1.69 arbitrary units for forced swimming and novelty stress respectively, P = 0.01). Importantly, the ultradian rhythm was rapidly restored after termination of the stress response. This is the first demonstration that the female rat brain is exposed to free corticosterone levels that follow a circadian as well as an ultradian pattern and show almost identical pulse characteristics as recently reported in male animals. These observations are of significance for further investigations into the dynamics of glucocorticoid action in the brain of both genders.

Corticosterone and corticosteroid binding globulin in birds: Relation to urbanization in a desert city

As cities expand worldwide, understanding how species adapt to novel urban habitats will become increasingly important to conservation. The adrenocortical stress response enables vertebrates to cope with novel environmental challenges to homeostasis. We examined total and estimates of free baseline and stress-induced corticosterone (CORT) concentrations and CORT binding globulin (CBG) levels in five passerine species within and around Phoenix, Arizona. We tested whether baseline and stress-induced CORT patterns differed among species living at varying densities in Phoenix and tested the hypothesis that, for species capable of successfully colonizing cities, individuals living in urban areas have a decreased acute stress response compared to individuals living in native desert. Baseline total CORT levels were generally similar in urban and rural birds. Capture and handling stress typically produced greater total CORT responses in urban birds than in rural birds, although these responses differed as a function of the life history stage (non-breeding, breeding or molt). CBG binding capacity did not change with life history stage or locality. Estimated free CORT concentrations differed less between groups than total CORT concentrations. Urban birds showed less variability in stress responses across life history stages than rural birds. We propose that more predictable resources in the city than in rural areas may decrease the need to vary stress responsiveness across life history stages. The results highlight the species-specific effects of urbanization on stress physiology and the difficulty to predict how urbanization impacts organisms.

Effects of corticosterone pellets on baseline and stress-induced corticosterone and corticosteroid-binding-globulin

Exogenous administration of glucocorticoids is a widely used and efficient tool to investigate the effects of elevated concentrations of these hormones in field studies. Because the effects of corticosterone are dose and duration-dependent, the exact course of plasma corticosterone levels after exogenous administration needs to be known. We tested the performance of self-degradable corticosterone pellets (implanted under the skin) in elevating plasma corticosterone levels. We monitored baseline (sampled within 3 min after capture) total corticosterone levels and investigated potential interactions with corticosteroid-binding-globulin (CBG) capacity and the endogenous corticosterone response to handling in Eurasian kestrel Falco tinnunculus and barn owl Tyto alba nestlings. Corticosterone pellets designed for a 7-day-release in rodents elevated circulating baseline total corticosterone during only 2–3 days compared to placebo-nestlings. Highest levels occurred 1–2 days after implantation and levels decreased strongly thereafter. CBG capacity was also increased, resulting in a smaller, but still significant, increase in baseline free corticosterone levels. The release of endogenous corticosterone as a response to handling was strong in placebo-nestlings, but absent 2 and 8 days after corticosterone pellet implantation. This indicates a potential shut-down of the hypothalamo–pituitary–adrenal axis after the 2–3 days of elevated baseline corticosterone levels. 20 days after pellet implantation, the endogenous corticosterone response to handling of nestlings implanted with corticosterone pellets attained similar levels as in placebo-nestlings. Self-degradable pellets proved to be an efficient tool to artificially elevate circulating baseline corticosterone especially in field studies, requiring only one intervention. The resulting peak-like elevation of circulating corticosterone, the concomitant elevation of CBG capacity, and the absence of an endogenous corticosterone response to an acute stressor have to be taken into account.

Reproductive experience alters corticosterone and CBG levels in the rat dam

Reproductive experience has significant effects on the brain, behavior and hormone profiles of the mother. Recent work has demonstrated that primiparous rats exhibit decreased dendritic arborizations in the hippocampus, and enhanced hippocampus-dependent spatial memory performance at the time of weaning compared to nulliparous and, to a lesser degree, multiparous rats. Interestingly, enhanced spatial learning and reduced dendritic arbors are seen in nulliparous female rats exposed to chronic stress or repeated corticosterone administration. Based on these observations, we hypothesized that corticosterone may be altered in primiparous rats compared to multiparous and nulliparous rats. The present study investigated whether the levels of circulating corticosterone and its binding protein, corticosteroid binding globulin (CBG), are altered with reproductive experience and pup-exposure during late pregnancy and the postpartum. Total serum corticosterone and CBG were assayed from five groups; multiparous, primiparous, nulliparous, primip-no-pups, and sensitized rats during gestation (days 14 and 19) and the postpartum period (days 1, 5, 14, 21, and 35). Results show that primiparous rats had significantly elevated total corticosterone on postpartum day 1. In addition, primiparous and multiparous rats had significantly lower CBG throughout the postpartum period than all other groups, with primiparous rats exhibiting lower levels than multiparous rats during mid-lactation. These data suggest that free corticosterone is elevated in both primiparous and multiparous dams and is elevated to a greater degree in primiparous compared to multiparous dams during lactation. Corticosterone and CBG levels were positively correlated with specific maternal behaviors during the first week postpartum in parturient rats, but not in sensitized rats, suggesting a role for corticosterone in the modulation of maternal behavior in parturient rats alone.

Corticosterone levels of tufted puffins vary with breeding stage, body condition index, and reproductive performance

Corticosterone (CORT) levels in free-living animals are seasonally modulated and vary with environmental conditions. Although most studies measure total CORT concentrations, levels of corticosteroid binding globulin (CBG) may also be modulated, thus altering the concentration of CORT available for diffusion into tissues (free CORT). We investigated the seasonal dynamics of CBG, total CORT, and free CORT in breeding tufted puffins ( Fratercula cirrhata) during 2 years characterized by high rates of nestling growth and survival. We then compared concentrations of total CORT in this population to levels in chick-rearing puffins at another colony during 2 years with low productivity. At the high productivity colony, levels of CBG, total baseline CORT, free baseline CORT, and total maximum CORT were all higher prior to egg-laying than during late incubation and late chick-rearing. Levels of CBG were positively correlated with body condition index (BCI) and free baseline CORT was negatively correlated with BCI. Total baseline levels of CORT during chick-rearing were two to four times higher at the colony with low rates of nestling growth and survival. Our results demonstrate the need for long-term datasets to disentangle seasonal trends in CORT levels from trends driven by changes in environmental conditions. Given the negative effects associated with chronic elevation of CORT, our results indicate the cost of reproduction may be higher during years characterized by low productivity.

Food restricted Tufted Puffin (Fratercula cirrhata) nestlings increase vocal activity during handling without modulating total or free corticosterone

In some species, corticosterone (CORT) appears to play a role in the control of begging behavior. Because of the potentially high costs associated with chronic elevation of CORT, it has also been proposed as a mechanism to ensure begging is an honest signal. We determined the effects of moderate food restriction (50% of high calorie treatment) on vocal behavior during handling, and on baseline levels of both total and ‘free’ unbound CORT in Tufted Puffin (Fratercula cirrhata) nestlings. Chick vocalizations during handling were similar to begging calls, and we assumed they were representative of begging behavior. We also measured total and free CORT in free-living Tufted Puffin chicks to determine if hormone levels in our experiment were comparable to natural levels. We found no effect of caloric restriction on either total or free baseline CORT, yet food-restricted nestlings vocalized more intensely during handling than chicks in the high calorie group. Mean plasma concentrations of total and free CORT in experimentally manipulated birds did not differ from levels in free-living nestlings. These results suggest that CORT does not play a role in modulating begging behavior in this species.

Circadian pattern of total and free corticosterone concentrations, corticosteroid-binding globulin, and physical activity in mice selectively bred for high voluntary wheel-running behavior

In vertebrates, baseline glucocorticoid concentrations vary predictably on a diel basis, usually peaking shortly before the onset of activity. Presumably, circadian patterns in glucocorticoid secretion have evolved to match predictable rises in energetic need. In mice from lines selectively bred for high voluntary wheel-running, previous studies have reported that baseline plasma corticosterone concentrations at two different times during the photophase are elevated twofold above those of non-selected control lines. Here, we tested the hypothesis that the elevated daytime corticosterone levels could be explained by a shift in the circadian pattern of corticosterone levels. We measured baseline total plasma corticosterone levels, corticosteroid-binding globulin (CBG) capacity, and calculated free corticosterone levels (corticosterone not bound to corticosteroid-binding globulin and potentially biologically active) at six points during the 24-hour cycle in males on a 12:12 photoperiod. We also examined the daily pattern of both wheel-running and home-cage activity. Based on combined analysis of all six points, the circadian pattern of total corticosterone, corticosteroid-binding globulin, and free corticosterone levels did not significantly differ between high-runner and control mice (linetype∗time interaction P=0.56, 0.45, and 0.55, respectively); however, all varied with time (all P<0.0001) and mice from the selected lines had significantly elevated total (P=0.0125) and free (P=0.0140) corticosterone, with no difference in CBG binding capacity (P=0.77). All mice were active primarily during the dark phase, and the factorial increase in activity of selected relative to controls lines was 2.33 for total daily wheel revolutions and 2.76 for total daily home-cage activity. The onset of the active period for both measures of locomotor activity coincided with peak total and free corticosterone levels in both selected and control lines. These findings lend support to our hypothesis that elevated circulating corticosterone levels have evolved as an adaptation to support increased locomotor activity in the selected lines.

Silicone Elastomer Uptake Method for Determination of Free 1‐Alkyl‐2‐Pyrrolidone Concentration in Micelle and Hydroxypropyl‐β‐Cyclodextrin Systems Used in Skin Transport Studies

Previous investigations in our laboratory demonstrated how the polar head group and alkyl chain of amphiphilic chemical skin permeation enhancers contribute to enhancer potency. In those studies enhancers with n-alkyl chain lengths of eight or less were investigated. In order to investigate enhancers with longer n-alkyl chain lengths, enhancer-solubilizing agents should be considered. Corticosterone (CS) flux enhancement along the lipoidal pathway of hairless mouse skin (HMS) was determined with the enhancers 1-hexyl- (HP), 1-octyl- (OP), 1-decyl- (DP), and 1-dodecyl-2-pyrrolidone (DoP) solubilized in 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(polyethylene glycol-2000] (DSPE) micelles or in hydroxypropyl-beta-cyclodextrin (HPbetaCD). The free CS, HP, OP, DP, and DoP aqueous concentrations in the DSPE micelle and HPbetaCD systems were determined using a partitioning method. Comparisons of the enhancer potencies based on the free concentration of the enhancers revealed a nearly semi-logarithmic linear relationship between enhancer potency and the carbon number of the alkyl chain length with a slope of approximately 0.55. The observed n-alkyl chain length dependency in the aqueous phase is consistent with the hydrophobic effect. This study shows that longer chain enhancers may be studied by employing a solubilizing system, and free enhancer concentration in these systems can be determined with the aid of the silicone elastomer uptake method.

The effect of chronic psychological stress on corticosterone, plasma metabolites, and immune responsiveness in European starlings

Although increases in glucocorticoid concentrations during acute stress are believed to help animals survive stressful events, chronic changes in glucocorticoid concentrations can alter metabolism and lead to disease. We studied the effect of chronic psychological stress on corticosterone (CORT), corticosterone binding globulin (CBG), glucose, and triglyceride concentrations as well as immune responsiveness to a T-cell mitogen challenge in European starlings, Sturnus vulgaris. To induce chronic stress we used a chronic stress protocol consisting of five stressors (loud radio, cage tapping, cage rolling, human voice, and bag restraint) administered in random order for 30 min for 4 times/day over 18 days. Total CORT decreased throughout the chronic stress period, which parallels a previous study with starlings. CBG capacity did not significantly change with chronic stress, thus free CORT followed the same pattern of attenuation as total CORT during chronic stress. Despite the change in regulation of CORT release, daytime glucose and triglyceride concentrations did not change with chronic stress. In addition, immune responsiveness in chronically stressed and unstressed birds was similar. Our results, together with a previous study using a similar CSP in European starlings, suggest that starlings physiologically dampen the HPA axis during chronic psychological stress to avoid pathology associated with chronically augmented CORT concentrations such as hyperglycemia and impaired immune function.

The temporal dynamics of intrahippocampal corticosterone in response to stress-related stimuli with different emotional and physical load: An in vivo microdialysis study in C57BL/6 and DBA/2 inbred mice

There is strong evidence for a pivotal interaction of corticosteroid signalling and behavioral adaptation to stress. To further elucidate this relation, we monitored the dynamics of free corticosterone in the murine hippocampus of two inbred mouse strains using in vivo microdialysis. C57BL/6JOlaHsd (C57BL/6) and DBA/2OlaHsd (DBA/2) inbred mouse strains have been shown to differ in their anxiety-related and depression-like behavior and provide, thus, an interesting animal model to study the stimulus-response profile of the hypothalamus-pituitary-adrenocortical (HPA) system as a function of emotional and physical load. We, first, compared peripheral and intracerebral concentration patterns of corticosterone by simultaneous microdialysis of the jugular vein and the hippocampus in anesthetized mice and found strain differences in blood versus intracerebral free corticosterone concentrations. C57BL/6 showed almost the same steroid levels in either compartment, whereas DBA/2 mice displayed higher glucocorticoid levels in the circulation than in the hippocampus. This data suggest a strain difference in the tissue environment influencing the amount of biological active corticosterone at the receptor site. Measurements of intrahippocampal corticosterone in freely moving mice revealed that DBA/2 display a prolonged glucocorticoid increase in response to a single forced swimming stress (FST), as compared to C57BL/6 mice indicating a reduced inhibitory HPA axis feedback. Exposure to a novel environment (NE) induced a desensitization of the HPA system in DBA/2 animals as they show an attenuated intracerebral corticosterone dynamics after a subsequent FST. Testing animals in an elevated plus-maze (EPM), however, did not significantly stimulate coriticosterone release in either strain. The analysis of the area under the curve revealed a high amount of corticosterone released through FST and a low glucocorticoid release after NE or EPM exposure that are independent of the strain. This data indicate a strong stimulus dependency of corticosterone secretion that is strain independent, whereas the dynamics and feedback of the HPA axis is different between both inbred strains. Behavioral phenotyping of animals revealed a strong impact of microdialysis procedure on FST and EPM performance. Innate emotionality differences of both strains, however, were not affected. Though descriptive in nature, the present results suggest an altered corticosteroid signalling in the DBA/2 strain compared to C57BL/6 mice. Whether this observation causally underlies the differences in anxiety-related and depression-like behavior has to be further experimentally validated. In addition, our study highlights the use of in vivo microdialysis to assess the neuroendocrine endophenotype of animal models via profiling of stimulus-response patterns of stress hormones.

Characterization and quantification of corticosteroid-binding globulin in a southern toad, Bufo terrestris, exposed to coal-combustion-waste

Corticosteroid-binding globulin (CBG) is a plasma protein that binds corticosterone and may regulate access of hormone to tissues. The role of CBG during a stress response is not clear. At least two hypotheses have been proposed: 1) CBG levels may increase in response to a stressor, thereby decreasing the amount of circulating free corticosterone, or 2) CBG levels may decline, making corticosterone available for its role in increased metabolic needs during stress. In this study, southern toads, Bufo terrestris, were exposed to a chronic pollutant (coal-combustion-waste), to determine changes in CBG and free corticosterone levels. Since toads exposed to chronic pollutants in previous studies did not exhibit the predicted changes in metabolic rate and mass, but did experience a significant elevation in total corticosterone, we hypothesized that CBG would likewise increase and thus, mitigate the effects of a chronic (i.e. 2 months) pollutant stressor. To conduct this study, we first characterized the properties of CBG in southern toads. Toad CBG has a K d = 20.6 ± 1.0 nM and a B max = 332.2 ± 5.1 nmol/L plasma. The rank order potencies for steroid inhibition of tritiated corticosterone are: dihydrotestosterone > corticosterone ≫ progesterone = testosterone ⋙ estrogen = dexamethasone. After characterization, we monitored the changes in CBG, total corticosterone, and free corticosterone in male toads that were exposed to either coal-combustion-waste or control conditions. CBG increased in all groups throughout the experiment. Total corticosterone, on the other hand, was only significantly elevated at four weeks of exposure to coal-combustion-waste. The increase in CBG did not parallel the increase in total corticosterone; as a result, free corticosterone levels were not buffered by CBG, but showed a peak at four weeks similar to total corticosterone. This finding indicates that, in this species, CBG may not provide a protective mechanism during long-term pollution exposure.

Development of stress reactivity in white-crowned sparrow nestlings: Total corticosterone response increases with age, while free corticosterone response remains low

Activation of the adrenocortical response to stress during development can have fundamental consequences over the lifetime of the organism; as such, many organisms are less responsive to stress during critical developmental periods. In this study, we evaluated stress reactivity in nestling white-crowned sparrows, examining corticosterone and binding globulin levels in response to both restraint stress and ACTH challenge. Restraint stress induced a significant corticosterone response in both 4 to 6- and 7 to 9-day-old nestlings, but not in the youngest group (1–3 days); this non-significant increase in corticosterone in the youngest birds resembles the mammalian hyporesponsive period, wherein young animals are resistant to most stressors. Binding globulin levels appear to extend this period of low reactivity: when free corticosterone levels were calculated, only the oldest age group (7–9 day) showed a significant response to restraint. ACTH challenge data revealed that all ages of white-crowned sparrow nestlings responded to exogenous ACTH treatment with significant elevation of corticosterone, although early-stage nestlings did not reach adult levels of response.

The importance of physiological ecology in conservation biology

Many of the threats to the persistence of populations of sensitive species have physiological or pathological mechanisms, and those mechanisms are best understood through the inherently integrative discipline of physiological ecology. The desert tortoise was listed under the Endangered Species Act largely due to a newly recognized upper respiratory disease thought to cause mortality in individuals and severe declines in populations. Numerous hypotheses about the threats to the persistence of desert tortoise populations involve acquisition of nutrients, and its connection to stress and disease. The nutritional wisdom hypothesis posits that animals should forage not for particular food items, but instead, for particular nutrients such as calcium and phosphorus used in building bones. The optimal foraging hypothesis suggests that, in circumstances of resource abundance, tortoises should forage as dietary specialists as a means of maximizing intake of resources. The optimal digestion hypothesis suggests that tortoises should process ingesta in ways that regulate assimilation rate. Finally, the cost-of-switching hypothesis suggests that herbivores, like the desert tortoise, should avoid switching food types to avoid negatively affecting the microbe community responsible for fermenting plants into energy and nutrients. Combining hypotheses into a resource acquisition theory leads to novel predictions that are generally supported by data presented here. Testing hypotheses, and synthesizing test results into a theory, provides a robust scientific alternative to the popular use of untested hypotheses and unanalyzed data to assert the needs of species. The scientific approach should focus on hypotheses concerning anthropogenic modifications of the environment that impact physiological processes ultimately important to population phenomena. We show how measurements of such impacts as nutrient starvation, can cause physiological stress, and that the endocrine mechanisms involved with stress can result in disease. Finally, our new syntheses evince a new hypothesis. Free molecules of the stress hormone corticosterone can inhibit immunity, and the abundance of "free corticosterone" in the blood (thought to be the active form of the hormone) is regulated when the corticosterone molecules combine with binding globulins. The sex hormone, testosterone, combines with the same binding globulin. High levels of testosterone, naturally occurring in the breeding season, may be further enhanced in populations at high densities, and the resulting excess testosterone may compete with binding globulins, thereby releasing corticosterone and reducing immunity to disease. This sequence could result in physiological and pathological phenomena leading to population cycles with a period that would be essentially impossible to observe in desert tortoise. Such cycles could obscure population fluctuations of anthropogenic origin.

Hyporesponsiveness to Glucocorticoids in Mice Genetically Deficient for the Corticosteroid Binding Globulin

Corticosteroid binding globulin (CBG) is the carrier for glucocorticoids in plasma. The protein is believed to keep the steroids inactive and to regulate the amount of free hormone acting on target tissues (free hormone hypothesis). Here, we generated a mouse model genetically deficient for CBG to test the contribution of the carrier to glucocorticoid action and adrenocortical stress response. The absence of CBG resulted in a lack of corticosterone binding activity in serum and in an approximately 10-fold increase in free corticosterone levels in CBG-null mice, consistent with its role in regulation of circulating free hormone levels. Surprisingly, cbg(-/-) animals did not exhibit features seen in organisms with enhanced glucocorticoid signaling. Rather, the mice exhibited increased activity of the pituitary axis of hormonal control, normal levels of gluconeogenetic enzymes, and fatigue, as well as an aggravated response to septic shock, indicating an inability to appropriately respond to the excess free corticosterone in the absence of CBG. Thus, our data suggest an active role for CBG in bioavailability, local delivery, and/or cellular signal transduction of glucocorticoids that extends beyond a function as a mere cargo transporter.

Attenuating corticosterone levels on the day of memory assessment prevents chronic stress‐induced impairments in spatial memory

This study investigated whether chronic stress-induced spatial memory deficits were caused by changes in the hypothalamic-pituitary-adrenal axis, such as corticosterone (CORT) elevations on the day of memory assessment, rather than the consequence of structural changes in the hippocampus. Male Sprague-Dawley rats were restrained for 6 h/day/21 days, and spatial memory was assessed on the Y-maze on day 22. Ninety minutes before training, rats received a subcutaneous injection of vehicle or metyrapone, a CORT synthesis inhibitor, and then spatial memory was determined 4-h later. The highest dose of metyrapone (75 mg/kg, s.c.) was most effective at preventing stress-induced spatial memory deficits. Chronic stress increased total CORT levels following Y-maze exposure, while acute metyrapone treatment dose-dependently attenuated total and free (unbound) CORT levels in both stress and control conditions. Blood samples taken from a separate subset of chronically stressed rats showed that baseline CORT levels were similar across the restraint period. Finally, chronic stress down-regulated glucocorticoid, but not mineralocorticoid, receptor mRNA expression within the hippocampus (dentate gyrus, CA1, CA2, CA3). These findings suggest that chronic stress-induced spatial memory deficits may be mediated by hypothalamic-pituitary-adrenal axis dysregulation. Specifically, CORT elevations and reductions in hippocampal glucocorticoid receptor expression, at the time of behavioural assessment may be involved, as opposed to a direct effect that is solely dependent upon hippocampal structural changes. These results have significance for treating cognitive decline in conditions associated with elevated glucocorticoids that include subpopulations in ageing, depression, Cushing's disease and Alzheimer's disease.

Corticosterone responses change seasonally in free-living house sparrows ( Passer domesticus)

Avian corticosterone (CORT) concentrations vary seasonally for many species, but most studies examined species from extreme environments or from only a few times of the year. It is unclear how general a phenomenon this is. We examined both baseline and stress-induced CORT from house sparrows ( Passer domesticus) in two habitats—Massachusetts, similar to their ancestral habitat; and New Mexico, a semi-arid desert where the house sparrow only thrives as an obligate human commensal. We captured both males and females during four times of the year—Spring (when the male cloacal protuberance indicated birds were in breeding condition), Fall, Winter, and in the late summer when birds were undergoing a prebasic molt. Birds were heaviest and had the longest wing chord lengths in the Fall at both sites and Massachusetts birds were approximately 10% heavier than New Mexico birds. House sparrows also showed a seasonal variation in the amount of fat stores, but the seasonal pattern differed. Massachusetts birds were fatter overall and showed the most fat during Fall and Winter, whereas New Mexico birds showed the most fat in the Spring. Both baseline and stress-induced total CORT did not differ between sexes or sites, with the exceptions of baseline CORT in the Fall and stress-induced CORT in the Spring being elevated in Massachusetts males. There was a distinct seasonal pattern at both sites, however, with total CORT being highest in the Spring and Winter and lowest during Fall and molt. This seasonal pattern was mirrored in corticosterone binding globulin (CBG) capacities, and when free CORT was estimated, the seasonal pattern disappeared. Stress-induced free CORT, however, was higher in Massachusetts males and females during the Fall and Winter, suggesting a potential differential response to stress associated with commensalism.

Behavioral and physiological responses to experimentally elevated testosterone in female dark-eyed juncos (Junco hyemalis carolinensis)

Testosterone mediates the expression of many fitness-related traits in male vertebrates and is thought to account for numerous sex differences in trait expression. Testosterone is also secreted by females; however, far less is known regarding its effects on female physiology and behavior. Using a bird species in which the effects of testosterone on males are well characterized, the dark-eyed junco (Junco hyemalis), we tested whether an increase in exogenous testosterone in females would alter the phenotypic expression of a suite of behavioral and physiological traits. We found that increased testosterone levels in female dark-eyed juncos led to decreased cell-mediated immune function and increased intrasexual aggression, hypothalamo-pituitary–adrenal (HPA) axis responsiveness, baseline corticosterone and corticosterone-binding globulin (CBG) levels. Furthermore, immunosuppression following testosterone implantation was negatively correlated with total and free testosterone but did not appear to be related to either total or free corticosterone. These results demonstrate that the phenotypic impact of elevated testosterone is not confined to males in dark-eyed juncos, and that the impact in adults can be similar in males and females. We discuss these results in the context of potential endocrine–immune interactions and the evolution of sexual dimorphism.