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The amygdala is a heterogeneous group of nuclei that plays a role in emotional and social learning. As such, there has been increased interest in its development in adolescent animals, a period in which emotional/social learning increases dramatically. While many mechanisms of amygdala development have been studied, the role of cell proliferation during adolescence has received less attention. Using bromodeoxyuridine (BrdU) injections in adolescent and adult rats, we previously found an almost fivefold increase in BrdU-positive cells in the amygdala of adolescents compared to adults. Approximately one third of BrdU-labeled cells in the amygdala contained the putative neural marker doublecortin (DCX), suggesting a potential for neurogenesis. To further investigate this possibility in adolescents, we examined the proliferative dynamics of DCX/BrdU-labeled cells. Surprisingly, DCX/BrdU-positive cells were found to comprise a stable subpopulation of BrdU-containing cells across survivals up to 56 days, and there was no evidence of neural maturation by 28 days after BrdU injection. Additionally, we found that approximately 50% of BrdU+ cells within the adolescent amygdala contain neural-glial antigen (NG2) and are therefore presumptive oligodendrocyte precursors (OPCs). We next characterized the response to a short-lived stressor (3-day repeated variable stress, RVS). The total BrdU-labeled cell number decreased by ∼30% by 13 days following RVS (10 days post-BrdU injection) as assessed by stereologic counting methods, but the DCX/BrdU-labeled subpopulation was relatively resistant to RVS effects. In contrast, NG2/BrdU-labeled cells were strongly influenced by RVS. We conclude that typical neurogenesis is not a feature of the adolescent amygdala. These findings point to several possibilities, including the possibility that DCX/BrdU cells are late-developing neural precursors, or a unique subtype of NG2 cell that is relatively resistant to stress. In contrast, many proliferating OPCs are significantly impacted by a short-lived stressor, suggesting consequences for myelination in the developing amygdala., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

Adolescence is characterized by changes in both behavior and neural organization. During this period, the amygdala, a structure that mediates social and emotional behaviors, is changing in terms of neural and glia density. We examined cell proliferation within the amygdala of adolescent (post natal day (PND) 31) and adult (PND 70) male Sprague-Dawley rats using BrdU (bromodeoxyuridine) to label dividing cells. BrdU-labeled cells were distributed throughout the amygdala, often found in fibers surrounding major nuclei. Using two independent cell counting strategies under light and confocal microcopy, respectively, we found significantly more labeled cells in the amygdala in adolescent compared to adult animals (239.3 ± 87.18 vs. 44.75 ± 13.68; n=4/group; p<.05). BrdU/doublecortin (DCX) positive cells constitute approximately 30% of all dividing cells in the amygdala in both adolescents and adults. These data suggest that compared to young adulthood, adolescence is a relatively active period of cell proliferation in the amygdala. Moreover, the normal decline in dividing cells with age does not preferentially affect cells co-containing DCX-immunoreactivity., (Copyright © 2013 Wiley Periodicals, Inc.)

Exposure to traumatic stressors typically causes lasting changes in emotionality and behavior. However, coping strategies have been shown to prevent and alleviate many stress consequences and the biological mechanisms that underlie coping are of great interest. Whereas the laboratory stressor inescapable tail-shock induces anxiety-like behaviors, here we demonstrate that permitting a rat to chew on a wooden dowel during administration of tail-shock prevented the development of anxiety like behaviors in the open field and juvenile social exploration tests. Uncontrollable stressors increase corticosterone and decrease thyroid hormone, and we hypothesized that coping would blunt these changes. While tail-shock did produce these effects, active coping did not alter hormone levels. The dissociation between behavioral resilience and circulating hormones is discussed with regard to the utility of these molecules as biomarkers for psychiatric disease., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Adolescence is a developmental time period marked by rapid changes in behavior and brain structure. Stress during adolescence has been shown to cause long-lasting behavioral changes, including increased anxiety- and depression-like behavior, in both rats and humans. These findings have led to the hypothesis that adolescence may be a particularly vulnerable or sensitive period for stress exposure. To investigate this hypothesis, we directly compared the effects of a 3-day stress exposure during either an adolescent (post natal day (PND) 27-29) or adult (PND 72-74) time window on exploratory behavior in male rats, measured 6 weeks post-stress (PND 72 or PND 117). Exploratory behavior was tested in both 'anxiogenic' and 'anxiolytic' environments, using the open field and novel object tests, respectively. We hypothesized that rats stressed as adolescents would show greater behavioral changes than rats stressed as adults. Contrary to our hypothesis, we found a decrease in exploratory behavior in the open-field test in both age groups. Furthermore, the magnitude of difference between stress and age-matched control animals was similar between age groups. In contrast, stress had no effect at either age in the novel object test, a more anxiolytic environment. Older adults showed decreased exploration in the novel object test compared to the younger adults, regardless of stress experience. These results suggest that adolescence is not a sensitive period for the effects of repeated variable stress on exploratory behavior in an anxiogenic environment., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Thyroid hormones are essential regulators of growth, development and normal bodily function and their release is coordinated by the hypothalamic-pituitary-thyroid (HPT) axis. While the HPT axis has been established as an acutely stress-responsive neuroendocrine system, relatively little is known about the mechanisms of its stress regulation. The present study examined acute stress-induced changes in peripheral hormone levels [triiodothyronine (T3); thyroxine (T4), thyroid-stimulating hormone (TSH), reverse triiodothyronine (rT3)] and central mRNA levels of regulators of the HPT axis [thyrotropin-releasing hormone (TRH), somatostatin (SST), type II deiodinase (D2)] in response to an inescapable tail-shock, a rodent model of stress. Additionally, we examined whether individual differences in spontaneous exploratory behavior in an open field test predicted basal levels of TH or differential susceptibility to the effects of stress. The stress condition was associated with decreases in peripheral T3, T4 and TSH, but not rT3, when compared with controls. No changes were observed in TRH or SST mRNA levels, but there was a trend suggesting stress-related increases in D2 mRNA. We also found that an animal's exploratory behavior in an unfamiliar open field arena was positively related to peripheral thyroid hormone levels and predicted the magnitude of stress-induced changes. In conclusion, we found suggestive evidence for stress-induced decrease in central drive HPT axis, but the central mechanisms of its stress regulation remain to be elucidated. Additionally, we found that individual differences in animals' exploratory behavior were correlated with peripheral TH levels., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Regulation of the production of hypothalamic-pituitary-adrenal (HPA) axis secretagogues, corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP), may be differentially sensitive to the negative feedback effects of glucocorticoids. We chose to study this phenomenon by examining the ability of dexamethasone to influence CRH and AVP heteronuclear RNA (hnRNA) levels in an escapable/inescapable (ES/IS) foot-shock stress paradigm. On Day 1, adult male rats were subjected to either ES or IS foot-shock; on Day 2, saline or dexamethasone (100 microg/kg) was administered 2 h prior to the stressor. We found that ES/IS foot-shock stimulated similar robust increases in plasma adrenocorticotrophic hormone (ACTH) and corticosterone concentrations, and medial parvocellular division of the paraventricular nucleus (mpPVN) AVP and CRH hnRNA and c-fos mRNA levels in saline-treated ES/IS rats. Dexamethasone pretreatment suppressed ACTH and corticosterone levels similarly in IS and ES animals. Dexamethasone pretreatment also suppressed mpPVN CRH and AVP hnRNA levels at 30 min. However, by 120 min, the mpPVN AVP hnRNA levels in dexamethasone-treated rats were similar to those measured in the saline group. We also found that rats that received the most shocks on Day 1 had greater HPA axis activation on Day 2. We conclude that the magnitude of the foot-shock stressor, determined by learned and immediate cues, is important in determining the magnitude of the HPA response.

Understanding environmental effects on mouse brain development would allow us to take advantage of powerful genetic tools to determine the interaction between genetic and epigenetic factors governing brain development in C57BL/6 mice. Experiment 1 examined whether time of day for neonatal manipulations affects adult stress-induced hormone secretion. Three rearing groups were examined: early handled (EH; dam removed 10 min/day); maternal separated (MS; dam removed 180 min/day); and an animal facility raised (AFR) control. Separations occurred during either the first or last 3 h of the light phase. Corticosterone (CORT) secretion in response to 100 dB white noise was assessed in adulthood. Both EH and MS males separated during the last 3 h of the light phase exhibited blunted stress-induced CORT compared to all other groups. Experiment 2 varied time of behavior testing. A fourth group was also added: maternal isolated (MI; separated from dam and littermates 180 min/day). Adult male behavior was assessed in three different tests. EH males tested in the elevated zero maze (EZM) during the light phase and MS males tested in the EZM during the dark phase exhibited diminished anxiety-like behavior compared to the other groups. We conclude that the EH protocol is marginally effective in blunting stress-induced CORT secretion and anxiety-like behavior in C57BL/6 mice, and these early handling effects are influenced by time of day. We also conclude that the 3 h MS or MI protocol is not effective in exacerbating future adult stress-induced CORT secretion or anxiety-like behavior in C57BL/6 mice.

There has been little research on effects of chronic stressors on neuroendocrine function in adolescence despite increasing evidence of enduring effects of stressors during this period on behaviour in adulthood. We previously reported that social stress (SS: daily 1 h isolation and new cage partner for 16 days) in adolescence altered locomotor responses to psychostimulants in adulthood. Here, we investigated neuroendocrine responses over the duration of the procedure that may underlie the enduring effects of SS. SS rats were compared to rats undergoing daily isolation only (ISO) and controls (CTL) to determine responses to acute and repeated isolation with and without social instability. At 30 days of age (first isolation), higher plasma corticosterone and corticotrophin-releasing hormone (CRH) mRNA expression in the paraventricular nucleus (PVN) of the hypothalamus and in the central nucleus of the amygdala (CeA) were found in males caged with a new partner (SS) after isolation than those returned to their original partner (ISO). On day 45, SS males and females showed less habituation (higher bioactive levels of corticosterone based on plasma corticosterone and corticosteroid binding globulin levels) to the 16th episode of isolation than did ISO. SS and ISO had higher baseline expression of CRH mRNA in the PVN on day 45 than did CTL, and only CTL had increased levels after isolation. CRH mRNA expression in the CeA increased to a first isolation in CTL and to a 16th isolation in SS but not in ISO males. Modest differences in social interactions were observed between SS and ISO when returned to their cages after isolation. The results suggest that mild social stressors in adolescence impede neuroendocrine adaptation to homotypic stressors. The resultant increase in exposure to glucocorticoids over adolescence may alter ongoing brain development and increase vulnerability to psychopathology.

Changes in peripheral thyroid hormone levels are associated with changes in human affective disorders, particularly depression. In the current study we used an animal stress paradigm, proposed to be an animal model of depression, to examine peripheral T(3) levels during and after escapable or inescapable stress in adult male rats. In this model, one animal can control the termination of foot-shock stress by performing a lever press, and therefore experiences escapable stress. His lever press also terminates the shock for his yoked partner, who has no control over the stressor, and therefore experiences inescapable stress. In three separate experiments, blood samples were collected during and after one or two sessions of escapable/inescapable stress. We found that exposure to inescapable stress, but not escapable stress, caused a decrease in T(3) levels 120 min post-stress initiation. Peripheral T(3) levels were not significantly altered in animals exposed to escapable stress. In sum, these results add to a large body of previous data indicating that psychological coping can prevent the effects of physical stress on many diverse systems.

Previous work has indicated that acute and repeated stress can alter thyroid hormone secretion. Corticosterone, the end product of hypothalamic-pituitary-adrenal (HPA) axis activation and strongly regulated by stress, has been suggested to play a role in hypothalamic-pituitary-thyroid (HPT) axis regulation. In the current study, we sought to further characterize HPT axis activity after repeated exposure to inescapable foot-shock stress (FS), and to examine changes in proposed regulators of the HPT axis, including plasma corticosterone and hypothalamic arcuate nucleus agouti-related protein (AGRP) mRNA levels. Adult male Sprague-Dawley rats were subjected to one daily session of inescapable FS for 14 days. Plasma corticosterone levels were determined during and after the stress on days 1 and 14. Animals were killed on day 15, and trunk blood and brains were collected for measurement of hormone and mRNA levels. Repeated exposure to FS led to a significant decrease in serum levels of 3,5,3'-triiodothyronine (T3) and 3,5,3',5'-tetraiodothyronine (T4). Stress-induced plasma corticosterone levels were not altered by repeated exposure to the stress. Despite the decrease in peripheral hormone levels, thyrotropin-releasing hormone (TRH) mRNA levels within the paraventricular nucleus of the hypothalamus were not altered by the stress paradigm. Arcuate nucleus AGRP mRNA levels were significantly increased in the animals exposed to repeated FS. Additionally, we noted significant correlations between stress-induced plasma corticosterone levels and components of the HPT axis, including TRH mRNA levels and free T4 levels. Additionally, there was a significant correlation between AGRP mRNA levels and total T3 levels. Changes in body weight were also correlated with peripheral corticosterone and TRH mRNA levels. These results suggest that repeated exposure to mild-electric foot-shock causes a decrease in peripheral thyroid hormone levels, and that components of the HPA axis and hypothalamic AGRP may be involved in stress regulation of the HPT., ((c) 2005 S. Karger AG, Basel)

This study investigated the effects of neonatal handling and maternal separation on the development of anxiety behavior and the hypothalamic-pituitary-adrenal axis of C57BL/6 mice. We hypothesized short periods of neonatal handling would diminish anxiety and secretion of corticosterone, while longer periods of maternal separation would elevate anxiety and plasma corticosterone compared to a nonhandled group. Mice were bred and reared as follows. After birth, each litter was assigned to one of four groups: mother and pups removed from the home cage for 10 min (group 1) or 180 min a day (group 2); mother only removed from home cage 180 min a day (group 3); and no handling until weaning (group 4). All separation occurred on the first 10 days of life. Juvenile males that experienced 10 min of separation/day exhibited decreased anxiety behavior compared to all other mice. A second group of litters were bred and reared according to groups 1, 2, and 4 as described above. Upon adulthood, anxiety behavior was assessed in males, and the corticosterone response to an acoustic stressor was quantified. No effect of differential rearing was observed on behavior, but there was a marked effect on plasma corticosterone secretion between the groups. Adult male mice neonatally handled for 10 min/day exhibited a blunted corticosterone response, and mice that experienced 180 min of maternal separation exhibited a prolonged corticosterone response to the acoustic stimulus compared to the nonhandled group. These results demonstrate the development of the mouse's hypothalamic-pituitary-adrenal axis can be modified by neonatal rearing conditions, and suggest that the mouse could be a viable animal model to determine the genetic-environmental interactions governing brain development.

Stress decreases sexual activity. However, emerging research suggests that the psychological aspect of control prevents the detrimental effects of stress on male mating behaviour. The present study examined the effects of chronic escapable/inescapable stress on mating behaviour in the male Syrian hamster. Additionally, the ability of the antidepressant clomipramine to prevent the adverse effects of stress on mating behaviour was explored. In this paradigm, two groups received the same electric footshock stress, but differed in the psychological aspect of control. Cohorts were divided into two groups. One group received clomipramine via a sugar water solution while the other received plain sugar water. Mating behaviour was quantified before and after 12 consecutive days of stress. The morning following the final stress and behaviour session, trunk blood and brains were collected to assess: (i) plasma concentrations of testosterone and glucocorticoids and (ii) corticotropin-releasing hormone (CRH) mRNA expression within the paraventricular nucleus of the hypothalamus (PVN). In the drug-free groups, several aspects of mating behaviour were disrupted by inescapable but not escapable stress, including anogenital investigation before the first ejaculation and time of first ejaculation. Additionally, both escapable and inescapable stress caused a decrease in total hit rate compared to the no-stress control group. Unlike the sugar-water treated animals, hamsters in either stress condition receiving clomipramine showed no differences in anogenital investigation, time of first ejaculation, hit rate, or any other aspect of mating behaviour measured, compared to the clomipramine no-stress control males. The stress-induced inhibition of mating behaviour could not be explained by changes in baseline plasma concentrations of testosterone or total glucocorticoids; these values did not vary between any of the six treatment groups. It was found that clomipramine lowers CRH mRNA expression in the PVN by 74%, regardless of stressor conditions. The results of the present study have broad implications for understanding the relationships between stress, depression and reproduction, and for the treatment of people and animals suffering from the adverse effects of stress.

Stress decreases sexual activity, but it is uncertain which aspects of stress are detrimental to reproduction. This study used an escapable/inescapable stress paradigm to attempt to dissociate physical from psychological components of stress, and assess each component's impact on reproductive behavior in the male Syrian hamster (Mesocricetus auratus). Two experiments were completed using this protocol where two animals receive the same physical stressor (an electric footshock) but differ in the psychological aspect of control. One group (executive) could terminate the shock for themselves as well as a second group (yoked) by pressing a bar. Experiment 1 demonstrated a significant increase in plasma glucocorticoids at the end of a single 90-min stress session with no difference in glucocorticoid levels between the executive and yoked groups at any time point. Experiment 2 quantified male reproductive behavior prior to and immediately following 12 days of escapable or inescapable stress in executive, yoked, and no-stress control hamsters (n = 12/group). Repeated-measures analysis of variance revealed a number of significant changes in reproductive behavior before and after stress in the three treatment groups. The most striking difference was a decrease in hit rate observed only in the animals that could not control their stress (yoked group). Hit rate in the executive males that received the exact same physical stressor but could terminate the shock by pressing a bar was nearly identical to control animals that never received any foot shock. Therefore, we conclude that coping or control can ameliorate the negative effects of stress on male reproductive behavior.

Cross-sectional and short-term prospective studies in humans support the concept that low energy availability, and not other factors associated with exercise, causes the development of exercise-induced reproductive dysfunction. To rigorously test this hypothesis, we performed a longitudinal study, examining the role of low energy availability on both the development and the reversal of exercise-induced amenorrhea, using a monkey model (Macaca fascicularis). Eight adult female monkeys developed amenorrhea (defined as absence of menses for at least 100 d, with low and unchanging concentrations of LH, FSH, E2, and P4) after gradually increasing their daily exercise to 12.3 +/- 0.9 km/d of running over a 7- to 24-month period. Food intake remained constant during exercise training. To test whether amenorrhea is caused by low energy availability, four of the eight amenorrheic monkeys were provided with supplemental calories (138-181% of calorie intake during amenorrhea) while they maintained their daily training. All four monkeys exhibited increased reproductive hormone levels and reestablished ovulatory cycles, with recovery times for circulating gonadotropin levels ranging from 12-57 d from the initiation of supplemental feeding. The rapidity of recovery within the reproductive axis in a given monkey was directly related to the amount of energy that was consumed during the period of supplemental feeding (r = -0.97; P < 0.05). Repeated measurements of plasma T3 concentrations, a marker of cellular energy availability, revealed a tight correlation between the changes in reproductive function and T3 levels, such that T3 significantly decreased (27%) with the induction and significantly increased (18%) with the reversal of amenorrhea (P < 0.05). These data provide strong evidence that low energy availability plays a causal role in the development of exercise-induced amenorrhea.

Cross-sectional studies of exercise-induced reproductive dysfunction have documented a high proportion of menstrual cycle disturbances in women involved in strenuous exercise training. However, longitudinal studies have been needed to examine individual susceptibility to exercise-induced reproductive dysfunction and to elucidate the progression of changes in reproductive function that occur with strenuous exercise training. Using the female cynomolgus monkey (Macaca fascicularis), we documented changes in menstrual cyclicity and patterns of LH, FSH, estradiol, and progesterone secretion as the animals developed exercise-induced amenorrhea. As monkeys gradually increased running to 12.3 +/- 0.9 km/day, body weight did not change significantly although food intake remained constant. The time spent training until amenorrhea developed varied widely among animals (7-24 months; mean = 14.3 +/- 2.2 months) and was not correlated with initial body weight, training distance, or food intake. Consistent changes in function of the reproductive axis occurred abruptly, one to two menstrual cycles before the development of amenorrhea. These included significant declines in plasma reproductive hormone concentrations, an increase in follicular phase length, and a decrease in luteal phase progesterone secretion. These data document a high level of interindividual variability in the development of exercise-induced reproductive dysfunction, delineate the progression of changes in reproductive hormone secretion that occur with exercise training, and illustrate an abrupt transition from normal cyclicity to an amenorrheic state in exercising individuals, that is not necessarily associated with weight loss.

We have previously demonstrated that microinjection of norepinephrine (NE) into the paraventricular nucleus of the hypothalamus (PVN) of conscious rats elicits a marked increase in CRH gene transcription, indicated by CRH hnRNA levels, without changing AVP hnRNA levels. We hypothesized that this differential response is due to differential sensitivity of AVP and CRH gene transcription to the inhibitory effects of the NE-induced rise in corticosterone. In the current study, we used animals that had been adrenalectomized and implanted with a subcutaneous corticosterone pellet (ADX/B) which prevented the NE-induced rise in corticosterone levels. NE (50 nmol) or artificial CSF was injected into the PVN of conscious rats, which had undergone either sham-operation (SHAM) or ADX/B 1 week earlier. CRH and AVP hnRNA levels were semi-quantitated by in situ hybridization using intron-specific riboprobes. In both SHAM and ADX/B animals, CRH hnRNA levels were significantly elevated at the 15 min time-point and returned to basal levels by 120 min. At 15 min, the magnitude of the CRH hnRNA response was only slightly greater in the ADX/B group than SHAM. In contrast, changes in medial parvocellular PVN AVP hnRNA levels in the ADX/B group were significantly greater than the changes observed in the SHAM group, at both the 15 and 120 min time-points. These results suggest that corticosterone has a greater impact on the transcriptional regulation of AVP than CRH, suggesting important differences and distinct roles of these secretagogues in the regulation of the hypothalamic-pituitary-adrenal axis.

All stress-related inputs are conveyed to the hypothalamus via several brain areas and integrated in the parvocellular division of the paraventricular nucleus (PVN) where corticotropin-releasing hormone (CRH) is synthesized. Arginine vasopressin (AVP) is present in both magnocellular and parvocellular divisions of the PVN, and the latter population of AVP is colocalized with CRH. CRH and AVP are co-secreted in the face of certain stressful stimuli, and synthesis of both peptides is suppressed by glucocorticoid. CRH and AVP stimulate corticotropin (ACTH) secretion synergistically, but the physiological relevance of the dual corticotroph regulation is not understood. Norepinephrine (NE) is a well known neurotransmitter that regulates CRH neurons in the PVN. We explored the mode of action of NE on CRH and AVP gene transcription in the PVN to examine the effect of the neurotransmitter on multiple genes that are responsible for a common physiological function. After NE injection into the PVN of conscious rats, CRH heteronuclear (hn) RNA increased rapidly and markedly in the parvocellular division of the PVN. AVP hnRNA did not change significantly in either the parvocellular or magnocellular division of the PVN after NE injection. The present results show that the transcription of CRH and AVP genes is differentially regulated by NE, indicating the complexity of neurotransmitter regulation of multiple releasing hormone genes in a discrete hypothalamic neuronal population.

Many stressors elicit changes in corticotrophin (CRH), enkephalin (ENK), and neurotensin (NT) mRNA levels within the medial parvocellular region of the paraventricular nucleus of the hypothalamus (mpPVN), and the pattern of changes in mRNA levels appears to depend on the physical characteristics of the stressor. We questioned whether psychologically distinct stressors would cause different patterns of neuropeptide mRNA expression within the PVN. Psychologically distinct stressors were created by employing a paradigm of escapable (controllable) vs. non-escapable (yoked) tail shock. An adult male rats could terminate the stress stimulus by performing wheel-turning behaviour; his behaviour also terminated the stress for his yoked partner, who had no control over the termination of the shock. Four h post-stress, brains were collected and processed for in-situ hybridization histochemistry. Tail-shock stress stimulated a significant increase in CRH, ENK, and NT mRNA levels within the mpPVN. The number of CRH identified neurones coexpressing AVP mRNA was also significantly elevated in both stress groups. Moreover, the pattern and magnitude of the stress-induced increases in mRNA was similar in both stress groups. Additionally, no stress-induced changes in CRH mRNA levels were observed in the central nucleus of the amygdala. In sum, two psychologically distinct stressors, escapable vs. yoked tail shock stress, stimulated similar increases in CRH, NT, ENK, and AVP mRNA levels within the mpPVN. These results suggest that physical attributes of a stress, rather than psychological, may be the more important factors in determining the PVN mRNA response.

The goal of this article is to describe some of the central nervous system circuits involved in the regulation of the hypothalamopituitary-adrenocortical (HPA) axis, with an emphasis on animal models believed to mimic the human experience of emotional stress. First, the basic constitutive elements of the HPA axis that control glucocorticoid secretion are reviewed. A description of the neural systems assumed to regulate the activity of the HPA axis, both anatomically and functionally, follows. It is argued that hypothalamic, septal and bed nucleus of the stria terminalis neurons are involved in the regulation of the HPA axis by situations eliciting emotional responses.

The current study was conducted to determine the potential relationship between stress-induced corticosterone secretion and corticosteroid receptor mRNA levels after 5 days of intermittent stress. In particular, we were interested in the rate at which animals terminate a stress response, and how this termination may be altered by repeated stress. Adult male Sprague-Dawley rats were subjected to either 5 days of restraint stress or 5 days of an unpredictable stress paradigm. Restraint-stress induced corticosterone secretion was measured on Days 1 and 5 in both groups, and animals were killed on Day 6. Glucocorticoid receptor (GR), and mineralocorticoid (MR) mRNA levels were determined using in-situ hybridization techniques. Five days of restraint stress caused an habituation of the plasma corticosterone response to stress measured 60 and 90 min post-stress initiation; this pattern of corticosterone secretion was not observed in the animals subjected to unpredictable stress. Five days of either stress paradigm did not alter MR mRNA levels measured within the hippocampus or GR mRNA levels within the hippocampus or the medial parvocellular division of the paraventricular nucleus of the hypothalamus (mpPVN). However, an individual's GR mRNA levels measured within the CA1/2 region of the hippocampus and the mpPVN were significantly correlated with the degree of habituation of the corticosterone response to stress measured on Day 5. This suggests that an increase in the rate of termination of the stress response and levels of GR within the hippocampus and mpPVN may be functionally related.

The elicitation and reduction of fear were indexed with fear-potentiated startle and corticosterone release and induction of the immediate-early gene c-fos as a marker of neural activity in male Sprague-Dawley rats. Conditioning consisted of pairing one stimulus with footshock, which was withheld when the conditioned stimulus was preceded by a different modality stimulus, the conditioned inhibitor. On the test day, approximately 60% of the rats were used for c-fos in situ hybridization, and were presented with either the conditioned stimulus alone, the conditioned inhibitor alone, a compound of the two stimuli, or no stimuli, and killed 30 min following the presentation of 10 such stimuli. The remaining rats were tested with the fear-potentiated startle paradigm. Rats displayed reliable fear-potentiated startle and corticosterone release to the conditioned stimulus, and both measures were reduced when the conditioned stimulus was preceded by the conditioned inhibitor. The ventral bed nucleus of the stria terminalis, septohypothalamic nucleus, some tegmental nuclei, and the locus coeruleus had particularly high c-fos induction in rats that received the conditioned inhibitor, providing one of the first functional indication that these nuclei might be important in behavioural or endocrine inhibition. Conditioning specific c-fos induction in the three groups that received a stimulus on the test day was observed in many hypothalamic areas, the medial geniculate body and the central gray, structures previously involved in fear and anxiety. The cingulate, infralimbic and perirhinal cortex, nucleus accumbens, lateral septum, dorsal endopiriform nucleus, and ventral tegmental area had higher c-fos induction in rats presented with the fearful conditioned stimulus, confirming previous studies. The amygdala and hippocampus of conditioned rats did not show higher c-fos induction than in rats repeatedly exposed to the context. Many regions displayed c-fos messenger RNA induction in the control condition, suggesting that processes other than fear and anxiety participate in c-fos induction.

The paraventricular nucleus of the hypothalamus (PVN) serves as the origin of the final common pathway in the secretion of glucocorticoid hormones in response to stress. Various stress-related inputs converge upon the cells of the medial parvocellular division of the PVN. These neurons, which synthesize and release corticotropin-releasing hormone, arginine vasopressin, and other secretagogues, are responsible for a cascade of events which culminates in the adrenocorticotropin-induced release of corticosteroids from the adrenal cortex. Previous data have suggested complex afferent regulation of PVN neurons, although the neuronal pathways by which the effects of stress are mediated remain to be fully disclosed. The present experiment sought to identify forebrain areas potentially involved in afferent regulation of the PVN in response to an acute stressor. Discrete injections of the retrograde tracer Fluoro-gold were delivered to the PVN, and rats were subsequently subjected to an acute swim stress. Brains were processed immunocytochemically for the simultaneous detection of the tracer and Fos, the protein product of the immediate early gene c-fos, utilized as a marker for neuronal activation. The majority of Fluoro-gold/Fos labeled neurons were detected in the parastrial nucleus, the medial preoptic area, the anterior hypothalamic area, the dorsomedial hypothalamic nucleus and adjacent posterior hypothalamic area, and, to a lesser extent, the supramammillary nucleus. These findings are discussed in relation to neural pathways mediating activation and inhibition of the hypothalamic-pituitary-adrenocortical axis.

Previously, we determined the pattern of stress-induced c-fos mRNA expression throughout the brain in order to gain further insight into the identification of the neural circuits mediating stress-induced regulation of the hypothalamic-pituitary-adrenal axis. In the present study, we determined if rapid effects of increased glucocorticoid levels after stress contribute to changes in c-fos mRNA expression. To this end, stress-induced c-fos expression was characterized in adrenalectomized (ADX) or adrenalectomized and corticosterone replaced (ADX/B) male rats. Animals were sacrificed 30 min post-onset of a 10 min swim stress, and in situ hybridization histochemistry was used to detect c-fos mRNA throughout the brain. The pattern of c-fos induction in the ADX and ADX/B animals was similar to that observed in the sham operated animals. Additionally, densitometric measurements were made to quantify the c-fos response in the paraventricular nucleus of the hypothalamus and the CA1/2 region of the hippocampus. We found that ADX did not alter the magnitude of the c-fos response to stress in these areas, but there was a slight dampening of the response in ADX/B animals. In sum, these results suggest that the pattern of c-fos expression observed 30 min post-stress is independent of stress-induced increases in circulating glucocorticoid concentrations.

States of chronic undernutrition can cause a profound suppression of reproductive function. To begin to determine the time course and the nature of the mechanism by which undernutrition suppresses the activity of the reproductive axis we have examined the effects of brief periods of fasting on reproductive hormone secretion in men and male rhesus monkeys. In monkeys there is a significant suppression of pulsatile luteinizing hormone (LH) and testosterone secretion after a single day of fasting, that is apparent within the first 4 h after a meal is missed. The suppression of pulsatile LH secretion on a day of fasting does not appear to be caused by the psychological stress experienced when monkeys are deprived of their daily meal in that monkeys who are maintained in a metabolically fed state (by feeding a large excess of food on the day prior to fasting), but are deprived of a meal and displayed behavioural agitation associated with fasting, have no suppression of LH secretion. The suppression of LH secretion on a day of fasting cannot be reversed by naloxone infusion, indicating that increased secretion of opioid peptides is not the primary mechanism causing the decrease in the central drive to the reproductive axis during fasting.(ABSTRACT TRUNCATED AT 250 WORDS)

One day of fasting suppresses pulsatile LH secretion in adult male rhesus monkeys. In the present study, we determined whether the hypothalamic-pituitary-adrenal (HPA) axis is activated by 1 day of fasting, and if there is any evidence that an increase in the activity of the HPA axis causes the fasting-induced suppression of LH secretion. In the first experiment, blood samples were collected at 15-min intervals from nine monkeys for 24 h (0800-0800 h) on a day of normal feeding (animals fed a single daily meal at 1100 h) and again on a day of fasting (animals not fed their daily meal) to assess changes in LH and cortisol secretion caused by fasting. LH secretion on the day of fasting (4.44 +/- 0.85 LH pulses/24 h) was significantly suppressed compared to that on a day of normal feeding (7.67 +/- 0.74 LH pulses/24 h; P < or = 0.01), and there was a slight, but significant, rise in plasma cortisol levels from 17.86 +/- 2.29 to 25.14 +/- 2.74 micrograms/dl (measured at 1400 h) that was evident soon after the meal was missed on the day of fasting. However, within individual animals, there was no correlation between the rise in plasma cortisol levels and the decrease in LH secretion observed on a day of fasting (r = 0.37). A second experiment was performed to determine whether the small rise in plasma cortisol levels that occurred on a day of fasting was capable of suppressing LH secretion. Hydrocortisone acetate (HCA) was administered to six normally fed monkeys, creating a rise in plasma cortisol levels somewhat greater than the levels occurring during fasting (HCA treatment, 44.51 +/- 1.48 micrograms/dl; fasting, 18.42 +/- 1.62 micrograms/dl; measured between 1100-2400 h). However, HCA treatment had no effect on LH secretion, suggesting that the rise in cortisol that occurs during fasting does not cause LH suppression. To determine whether increased secretion of the neuropeptides that provide the central drive to the adrenal axis causes the suppression of LH secretion during fasting, six animals were treated with dexamethasone (0.25 mg/kg, im) on a day of fasting to inhibit the activity of the glucocorticoid-sensitive CRH neurons of the adrenal axis via increased glucocorticoid negative feedback. Dexamethasone treatment caused a marked suppression of circulating cortisol levels (to approximately 1.0 micrograms/dl), but it did not prevent the fasting-induced suppression of pulsatile LH secretion.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult male rhesus monkeys have a diurnal pattern of reproductive hormone secretion that is characterized by significantly elevated LH and testosterone secretion in the evening hours and a nadir in secretion of these hormones in the morning. To test the hypothesis that the daily pattern of food intake may play a role in regulating the diurnal pattern of reproductive hormone secretion we performed three studies. First, to determine the relationship between the timing of the diurnal rise in LH secretion and meal consumption, blood samples were collected from 13 adult male rhesus monkeys via chronically indwelling venous catheters (samples every 15-20 min from 0800-0800 h) while monkeys were maintained on the standard feeding regimen in our colony (one meal of Purina monkey chow fed between 1100 and 1200 h). On a day of normal feeding there was a significant diurnal rhythm in mean LH concentrations with elevated levels at night (nadir: 13.41 +/- 0.82 ng/ml from 0800-1100 h; peak: 21.34 +/- 1.56 ng/ml from 2000-2300 h, P < or = 0.05). The rising phase of the diurnal rhythm in LH secretion was apparent starting in the early afternoon, shortly after the daily meal, at 1400 h (5 h before lights went off at 1900 h), and the diurnal rise in LH secretion was no longer apparent by 0500 h (several hours before the lights went on at 0700 h). Second, we examined the influence of missing the daily meal on the diurnal pattern of LH and testosterone secretion. Blood samples were collected for a 24-h period on a day of fasting from 9 monkeys. On a day of fasting there was no diurnal rise in plasma LH or testosterone concentrations; plasma concentrations of these hormones remained at the low morning levels throughout the day. Third, we examined the diurnal pattern of LH and testosterone secretion after adapting 5 monkeys (for 6-8 weeks) to a new meal time that was 6 h later in the day than the standard meal time (i.e. at 1700 h). After adaptation to this later feeding time monkeys showed a 6-h delay in the time of the first significant rise in LH secretion (which occurred at 2000 h).(ABSTRACT TRUNCATED AT 400 WORDS)

Short periods of fasting have been shown to cause a significant slowing of pulsatile LH secretion in men and male rhesus monkeys, which appears to result from a slowing of GnRH drive to the reproductive axis. To determine whether an increased activity of endogenous opioid peptides causes this slowing of pulsatile LH secretion, the ability of naloxone administration to reverse the fasting-induced suppression of LH secretion was tested. For this study, 6 adult male rhesus monkeys, with indwelling femoral and jugular venous catheters, were maintained on tether/swivel systems. Naloxone was administered to monkeys as a continuous infusion (0.25 mg/kg/h, with an initial loading dose of approximately 1.0 mg) for 8 h (16.00 to 24.00 h) on a day of normal feeding and again on a day of fasting. The LH response to naloxone was determined by collecting blood samples every 15 min from 12.00 to 24.00 h. LH pulse frequency on a day of normal feeding was 4.0 +/- 0.52 pulses/8 h, and naloxone administration on a day of feeding increased LH pulse frequency to 6.8 +/- 0.86 pulses/8 h. On a day of fasting, LH pulse frequency was 1.67 +/- 0.67 pulses/8 h, and naloxone administration on a day of fasting slightly, but not significantly, increased LH pulse frequency to 2.5 +/- 0.51 pulses/8 h. Similar studies were performed with a higher dose of naloxone (0.625 mg/kg/h, with an initial loading dose of approximately 2.0 mg) and again naloxone administration did not reverse the effects of fasting on pulsatile LH secretion. These results suggest that the slowing of pulsatile LH release that occurs with short periods of food restriction does not result from increased secretion of endogenous opioid peptides.

Recent reports indicate that in male rats dehydration, LiCl, and cholecystokinin (CCK) each stimulate pituitary oxytocin (OT) secretion and also decrease gastric emptying and motility. In contrast, the present experiments demonstrate that nipple attachment and sucking by pups, a well-known stimulus for neurohypophysial secretion of OT, did not decrease gastric motility in lactating rats. Moreover, systemic injection of naloxone, which is known to potentiate the inhibitory effects of LiCl and CCK on gastric motility in male rats, had no effect on gastric motility of lactating rats while nursing. These data indicate that pituitary OT secretion from magnocellular neurons is not invariably linked to decreased gastric motility in rats. As such, our results support previous findings that inhibition of gastric motility is not secondary to the pituitary secretion of OT but allow a possible role for parvocellular oxytocinergic neurons that project from the hypothalamic paraventricular nucleus to the brain stem in the control of gastric function.