PACAP centrally mediates emotional stress-induced corticosterone responses in mice

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic neuropeptide widely distributed in the nervous system. Recently, PACAP was shown to be involved in restraint stress-induced corticosterone release and concomitant expression of the genes involved in hypothalamic-pituitary-adrenal (HPA) axis activation. Therefore, in this study, we have addressed the types of stressors and the levels of the HPA axis in which PACAP signaling is involved using mice lacking PACAP (PACAP?/?). Among four different types of stressors, open-field exposure, cold exposure, ether inhalation, and restraint, the corticosterone response to open-field exposure and restraint, which are categorized as emotional stressors, but not the other two, was markedly attenuated in PACAP?/? mice. Peripheral administration of corticotropin releasing factor (CRF) or adrenocorticotropic hormone induced corticosterone increase similarly in PACAP?/? and wild-type mice. In addition, the restraint stress-induced c-Fos expression was significantly decreased in the paraventricular nucleus (PVN) and medial amygdala (MeA), but not the medial prefrontal cortex, in PACAP?/? mice. In the PVN of PACAP?/? mice, the stress-induced c-Fos expression was blunted in the CRF neurons. These results suggest that PACAP is critically involved in activation of the MeA and PVN CRF neurons to centrally regulate the HPA axis response to emotional stressors.     

The role of corticotrophin-releasing hormone receptors in the calcitonin gene-related peptide-induced suppression of pulsatile luteinising hormone secretion in the female rat

Corticotrophin-releasing hormone (CRH) plays a pivotal role in the suppression of the gonadotrophin-releasing hormone (GRH) pulse generator in response to stress and intracerebroventricular (i.c.v.) administration of calcitonin gene-related peptide (CGRP). We have previously shown both CRH receptor subtypes, CRH-R1 and CRH-R2, are involved in the stress-induced suppression of LH pulses. The aims of the present study were to examine the role of CRH-R1 and CRH-R2 in CGRP-induced suppression of LH pulses, and to investigate the effects of CGRP on CRH expression in the paraventricular nucleus (PVN) and central nucleus of the amygdala (CeA), which have prominent CRH neurone populations that receive dense CGRP innervations. The suppression of LH pulses by CGRP (1.5 microg i.c.v.) was completely prevented by intravenous administration of the CRH-R1 antagonist SSR125543Q (7.5 mg/rat i.v., 30 min before CGRP), but was not affected by the CRH-R2 antagonist, astressin(2)-B (100 microg i.c.v., 10 min before CGRP). CGRP increased the CRH mRNA expression in PVN and CeA. These results provide evidence of a role for CRH-R1 in mediating the suppressive effects of CGRP on pulsatile LH secretion in the female rat, and additionally raise the possibility of an involvement of PVN and CeA CRH neuronal populations in this suppression.     

Correlation between Changes in Stress-Induced Corticosterone Secretion and GR mRNA Levels

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 effect of adrenalectomy on Fos expression in vasopressinergic and oxytocinergic neurons in response to stress in the rat

This study evaluated the responses of vasopressin (AVP) and oxytocin (OT) neurons to alterations in hypothalamo-pituitary axis activity by adrenalectomy (ADX) or after restraint stress compared with basal conditions. Wistar male rats were perfuse-fixed by cardiac perfusion under anesthesia 3 h, 1, 3 and 14 days after ADX or Sham surgery. Coronal hypothalamic sections were used for evaluation of Fos, AVP and OT expression by immunohistochemistry. Under basal conditions and after stress, Fos-AVP double labeling showed no difference in the magnocellular subdivisions of the paraventricular nuclei (PVN) or in the supraoptic nuclei (SON), suggesting that the magnocellular AVP system is unlikely to contribute to ACTH secretion after restraint in both Sham and ADX rats. Fos-AVP double labeling in the parvocellular medial paraventricular nucleus (PaMP) in ADX groups was increased after 3 h in basal conditions, and in all periods after restraint stress. There were no differences between Sham and ADX groups in Fos-OT double labeling in any subdivision of the PVN; however, in the SON, the number of Fos-OT double labeled cells was increased at all time-points after stress in the ADX group. Fos expression was increased in the PaMP after 3 h and after restraint stress in the Sham and ADX groups, especially in the ADX group. In conclusion, Fos expression in different cell populations of the PVN can be differentially regulated by short- and long-term absence of glucocorticoid negative feedback and also by stress-related excitatory and/or inhibitory neural inputs. The Fos-AVP double labeling findings in the PaMP also indicate a minor participation of these vasopressinergic neurons in the regulation of the HPA axis after ADX.     

Substance P is involved in terminating the hypothalamo- pituitary-adrenal axis response to acute stress through centrally located neurokinin-1 receptors

The neurokinin substance P (SP) has been previously shown to inhibit basal hypothalamo-pituitary-adrenal (HPA) axis activity. This study was designed to investigate the effects of central injection of the specific neurokinin-1 receptor antagonist RP67580 on the HPA axis response to acute restraint stress. In non-restrained rats injected with RP67580, plasma ACTH and corticosterone levels were elevated at 30 and 60 min compared to rats injected with vehicle, but there were no differences between vehicle and RP67580 groups at 4h. In restrained rats injected with vehicle, plasma ACTH and corticosterone levels were significantly elevated at 30 min and 60 min following initiation of the stress but had returned to basal levels at 4h. In restrained rats injected icv with RP67580, plasma corticosterone and ACTH levels were significantly elevated at 30 min and 60 min, with no significant differences compared to the restraint stressed vehicle-injected group. However, in the RP67580-injected group, corticosterone and ACTH levels remained significantly elevated at 4h following onset of restraint compared to those in the restraint stressed vehicle-injected group. Corticotrophin-releasing factor mRNA levels in the parvocellular subdivision of the paraventricular nucleus of the hypothalamus and POMC mRNA levels in the anterior pituitary were significantly increased in the stressed group 4h following injection with RP67580 compared to the stressed group injected with vehicle alone. These data show that endogenous SP does not inhibit the initial magnitude of the HPA axis response to restraint stress, but does act through neurokinin-1 receptors at a central level to reduce the duration of the response to stress. This suggests that SP may be an important central agent controlling the transition between acute and chronic stress.     

Dexamethasone and stressor-magnitude regulation of stress-induced transcription of HPA axis secretagogues in the rat

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.     

The effect of hypothalamic lesions on hypothalamo-pituitary-adrenal axis activity and inflammation in adjuvant-induced arthritis

Adjuvant-induced arthritis (AA) was induced in control and in hypothalamic lesioned Piebald-Viral-Glaxo (PVG) rats. Following discrete paraventricular nucleus (PVN) lesions plasma corticosterone was increased 14 days after adjuvant injection as in controls, when hind paw inflammation was apparent. PVN lesion did not affect the severity of inflammation.In contrast, following medial basal hypothalamus (MBH) lesions adjuvant did not increase corticosterone levels and the increase in paw volume at day 14 was potentiated. Basal proopiomelanocortin(POMC) mRNA expression in the anterior lobe was unchanged by PVN lesions and decreased by MBH lesions. AA increased POMC mRNA in controls and in both PVN and MBH lesioned rats. After complete MBH lesion, surviving anterior pituitary tissue maintained morning levels of corticosterone.Thus, AA may activate the hypothalamo-pituitary-adrenal axis without the mediation of PVN neurones projecting to the median eminence. However, the loss of the corticosterone response to AA and the increase in severity of inflammation in the MBH lesioned rats suggests a central (non-PVN) component mediates effects of inflammation. Furthermore, the increase in POMC mRNA in the MBH lesioned AA rats suggests that part of this process is not mediated by releasing factors in the hypothalamo-hypophysial portal system, and that extrahypothalamic(peripheral) mediators act on the pituitary during chronic inflammation.     

Stress-induced progesterone secretion and progesterone receptor immunoreactivity in the paraventricular nucleus are modulated by pubertal development in male rats

Male rats show a differential adrenocortical response to stress before and after pubertal development, such that prepubertal animals have a more prolonged stress-induced corticosterone response compared to adults. Whether pubertal maturation affects other adrenocortical responses to stress is currently unknown. To address this question, we assessed stress-induced progesterone secretion in both intact and gonadectomized prepubertal (28 days of age) and adult (77 days of age) male rats either before or after exposure to a 30 min session of restraint stress. We found that prepubertal males show a greater and more prolonged stress-induced progesterone response compared to adults. We also found a similar effect in castrated prepubertal and adult males, indicating the differential stress-induced progesterone response is not gonadal in origin. We also examined progesterone receptor (PR) levels by immunohistochemistry in the paraventricular nucleus (PVN) of the hypothalamus, a key regulatory nucleus of the hypothalamic-pituitary-adrenal (HPA) axis, and found lower PR protein expression in the PVN of prepubertal compared to adult males. These data indicate that in addition to corticosterone, stress-induced adrenocortical progesterone levels are differentially affected by pubertal maturation. Furthermore, these data raise the possibility of different progesterone sensitivity of the PVN before and after puberty. The significance of this differential response is presently unknown. However, given the pleiotropic effects of progesterone on male physiology and behaviour, it is likely that the disparate post-stress exposure to progesterone affects the prepubertal and adult male differently.     

A single exposure to severe stressors causes long-term desensitisation of the physiological response to the homotypic stressor

Although some laboratories have reported that a single session of stress is able to induce a long-lasting sensitisation of the hypothalamic-pituitary-adrenal (HPA) response to further exposures to stress, we have found that a single exposure to severe emotional (immobilisation, restraint or shock) or systemic (endotoxin) stressors reduces the responsiveness of the HPA to the same, but not to a novel (heterotypic), stressor, in which case a slight sensitisation was observed. Long-term desensitisation has been found to reduce not only secretion of peripheral HPA hormones (ACTH and corticosterone), but also to reduce responses of central components of the HPA axis (c-fos and CRF gene expression at the level of the paraventricular nucleus of the hypothalamus, PVN). In addition, desensitisation also applies to the impact of the stressor on food intake and, probably, to stress-induced hyperglycaemia. The development of long-term desensitisation of the HPA axis does not appear to be a universal consequence of exposure to severe stressors as it was not observed in response to insulin-induced hypoglycaemia. Whether or not the development of long-term effects of stress depend on the specific pathways activated by particular stressors remains to be tested. The observed desensitisation of the HPA axis in response to the homotypic stressor shows two special features which makes it difficult to be interpreted in terms of an habituation-like process: (a) the effect increased with time (days to weeks) elapsed between the first and second exposure to the stressor, suggesting a progressive maturational process; and (b) the stronger the stressor the greater the long-term desensitisation. Therefore, it is possible that desensitisation of the HPA axis is the sum of two different phenomena: long-term effects and habituation-like processes. The contribution of the former may be more relevant with severe stressors and longer inter-stress intervals, and that of the latter with mild stressors and repeated exposures. Long-term stress-induced changes may not take place at the level of the PVN itself, but in brain nuclei showing synaptic plasticity and putatively involved in the control of the HPA axis and other physiological responses. As for the precise areas involved, these remain to be characterized.     

Hypothalamo-pituitary-adrenocortical axis function and hedonic behavior in adult male and female rats prenatally stressed by maternal food restriction

Neuroendocrine activation during stress is affected by many factors contributing to the variability of the stress response. The present study was aimed at evaluating long-term changes in hypothalamo-pituitary-adrenocortical (HPA) axis function and in hedonic behavior in adult offspring prenatally stressed by maternal food restriction, with attention on possible gender differences. Adult offspring were blood sampled via a tail artery cannula. Prenatally stressed females had significantly higher adrenal weights compared to males. Plasma ACTH levels, which rose in response to acute stress induced by handling, were significantly higher in females compared to those in males. A similar pattern was found in plasma corticosterone. The rise in ACTH levels was more pronounced in prenatally stressed rats though the rise in corticosterone failed to be modified. Corticotropin releasing hormone (CRH) and proopiomelanocortin mRNA levels in the hypothalamic paraventricular nucleus and anterior pituitary, respectively, were found to be unchanged. The present experiments failed to reveal a decrease in hedonic behavior in prenatally stressed rats. In contrast, in male offspring a tendency to a higher sucrose preference was observed. These data together with observed changes in hormone and CRH mRNA levels indicate that the gestational stress used did not result in a depression-like state in adult offspring.     

The central nucleus of the amygdala; a conduit for modulation of HPA axis responses to an immune challenge?

Physical stressors such as infection, inflammation and tissue injury elicit activation of the hypothalamic-pituitary-adrenal (HPA) axis. This response has significant implications for both immune and central nervous system function. Investigations in rats into the neural substrates responsible for HPA axis activation to an immune challenge have predominantly utilized an experimental paradigm involving the acute administration of the pro-inflammatory cytokine interleukin- 1β (IL-1β). It is well recognized that medial parvocellular corticotrophin-releasing factor cells of the paraventricular nucleus (mPVN CRF) are critical in generating HPA axis responses to an immune challenge but little is known about how peripheral immune signals can activate and/or modulate the mPVN CRF cells. Studies that have examined the afferent control of the mPVN CRF cell response to systemic IL-1β have centred largely on the inputs from brainstem catecholamine cells. However, other regulatory neuronal populations also merit attention and one such region is a component of the limbic system, the central nucleus of the amygdala (CeA). A large number of CeA cells are recruited following systemic IL-lβ administration and there is a significant body of work indicating that the CeA can influence HPA axis function. However, the contribution of the CeA to HPA axis responses to an immune challenge is only just beginning to be addressed. This review examines three aspects of HPA axis control by systemic IL-1β: (i) whether the CeA has a role in generating HPA axis responses to systemic IL-1β, (ii) the identity of the neural connections between the CeA and mPVN CRF cells that might be important to HPA axis responses and(iii) the mechanisms by which systemic IL-Iβ triggers the recruitment of CeA cells.     

Hypothalamic circuit regulating colonic transit following chronic stress in rats

Although acute stress accelerates colonic transit, the effect of chronic stress on colonic transit remains unclear. In this study, rats received repeated restraint stress (chronic homotypic stress) or various types of stress (chronic heterotypic stress) for 5 and 7 days, respectively. Vehicle saline, oxytocin (OXT), OXT receptor antagonist or corticotropin-releasing factor (CRF) receptor antagonists were administered by intracerebroventricular (ICV) injection prior to restraint stress for 90 min. Immediately after the stress exposure, the entire colon was removed and the geometric center (GC) of Na51CrO4 (a nonabsorbable radioactive marker; 0.5 μCi) distribution was calculated to measure the transit. Gene expression of OXT and CRF in the paraventricular nucleus (PVN) was evaluated by in situ hybridization. Accelerated colonic transit with the acute stressor was no longer observed following chronic homotypic stress. This restored colonic transit was reversed by ICV injection of an OXT antagonist. In contrast, chronic heterotypic stress significantly accelerated colonic transit, which was attenuated by ICV injection of OXT and by a CRF receptor 1 antagonist. OXT mRNA expression in the PVN was significantly increased following chronic homotypic stress, but not chronic heterotypic stress. However, CRF mRNA expression in the PVN was significantly increased following acute and chronic heterotypic stress, but not chronic homotypic stress. These results indicate that central OXT and CRF play a pivotal role in mediating the colonic dysmotility following chronic stress in rats.     

Fos expression in CRF-containing neurons in the rat paraventricular nucleus after central administration of neuromedin U

We examined the effects of centrally administered neuromedin U (NMU) on corticotrophin-releasing factor (CRF)-containing neurons in the hypothalamic paraventricular nucleus (PVN) of rats, using double immunohistochemistry for CRF and Fos. Almost all CRF-containing neurons in the parvocellular divisions of the PVN expressed Fos-like immunoreactivity 90 min after intracerebroventricular administration of NMU (3 nmol/rat). This results suggest the possibility that central NMU may be involved in stress-induced activation of CRF-containing neurons in the PVN.     

Patterns of neuronal activation in the rat brain and spinal cord in response to increasing durations of restraint stress

By most accounts the psychological stressor restraint produces a distinct pattern of neuronal activation in the brain. However, some evidence is incongruous with this pattern, leading us to propose that the restraint-induced pattern in the central nervous system might depend on the duration of restraint used. We therefore determined the pattern of neuronal activation (as indicated by the presence of Fos protein) seen in the paraventricular nucleus (PVN), bed nucleus of the stria terminalis, amygdala, locus coeruleus, nucleus tractus solitarius (NTS), ventrolateral medulla (VLM) and thoracic spinal cord of the rat in response to 0, 15, 30 or 60 min periods of restraint. We found that although a number of cell groups displayed a linear increase in activity with increasing durations of restraint (e.g. hypothalamic corticotrophin-releasing factor (CRF) cells, medial amygdala neurons and sympathetic preganglionic neurons of the thoracic spinal cord), a number of cell groups did not. For example, in the central amygdala restraint produced both a decrease in CRF cell activity and an increase in non-CRF cell activity. In the locus coeruleus, noradrenergic neurons did not display Fos in response to 15 min of restraint, but were significantly activated by 30 or 60 min restraint. After 30 or 60 min restraint a greater degree of activation of more rostral A1 noradrenergic neurons was observed compared with the pattern of A1 noradrenergic neurons in response to 15 min restraint. The results of this study demonstrate that restraint stress duration determines the amount and the pattern of neuronal activation seen in response to this psychological stressor.     

Acute exposure to a novel stressor further reduces the habituated corticosterone response to restraint in rats

The present study sought to identify dishabituation of the hypothalamic-pituitary-adrenal(HPA) axis response to different psychological stressors. Young adult male Sprague Dawley rats were exposed to five, 1 h sessions of restraint stress on five consecutive days. On the sixth day, and 2 h before additional exposure to restraint, animals were subjected to 30 min of a small (27cm square), elevated open field stressor (pedestal), which served as the dishabituating stimulus. We predicted HPA axis response dishabituation in chronically restrained rats exposed to the novel pedestal. Rats which underwent five days of restraint stress showed significantly blunted plasma corticosterone levels to restraint (habituation) as compared to restraint-nai've rats. However, rats which underwent five sessions of restraint responded with an enhanced habituation response when confronted with restraint shortly after exposure to the novel pedestal. Instead of HPA axis response dishabituation, we observed enhanced habituation. Subsequent experiments determined that a 1.25 mgkg corticosterone injection could substitute for pedestal exposure to produce enhanced restraint habituation.Combined treatment with both the glucocorticoid receptor antagonist RU40555 (30 mgkg)and the mineralocorticoid receptor antagonist RU283 18 (50 mgkg) blocked the expression of enhanced habituation after pedestal exposure. Thus, the delayed corticosterone negative feedback produced by novel stress led to enhanced expression of corticosterone response habituation.     

Maternal separation in early life impairs tumor immunity in adulthood in the F344 rat

Neonatal stress alters the hypothalamic-pituitary-adrenal (HPA) axis in rodents, such that, when these animals are exposed to stress as adults they hypersecrete corticosterone. Given that glucocorticoids are immunosuppressive, we examined the impact of maternal separation on HPA axis reactivity, natural killer (NK) cytotoxicity, and tumor growth in Fischer 344 rats following chronic restraint stress in adulthood. Pups underwent a chronic stress protocol whereby they were separated from their dams for 3 h on postnatal days 1-21. In adulthood, corticosterone responses were assessed following exposure to chronic (6 days for 10 h) restraint stress. Rats allocated to the chronic stress condition were inoculated with MADB106 tumor cells on day 4 of the restraint protocol. Blood was assessed for NK cytotoxicity on the final day of the chronic restraint protocol, and tumor colonization was assessed 3 weeks thereafter. Maternal separation impaired developmental weight gain (P < 0.05), depressed NK cytotoxicity (P < 0.05), and increased tumor colonization in the presence of chronic restraint stress in adulthood (P < 0.00 l). These findings occurred independently of circulating plasma corticosterone as only adult stress exposure potentiated corticosterone responses (P < 0.05). Our findings indicate that maternal separation and chronic stress can impair NK cytotoxicity and hence tumor immunity, but these effects are not directly mediated by perturbations in HPA axis function.     

Clamped Corticosterone (B) Reveals the Effect of Endogenous B on Both Facilitated Responsivity to Acute Restraint and Metabolic Responses to Chronic Stress

To determine the effects of both corticosterone (B) and chronic stressors on acute ACTH responses to restraint, young male rats were exposed to streptozotocin-induced diabetes, cold (5-7 degreesC) or intracerebroventricular (icv) neuropeptide Y (NPY) for 5 d and then exposed to restraint within 2 h after lights on. Two groups of rats were studied: intact and adrenalectomized replaced with B pellets that maintained plasma B in the normal mean 24-h range of intact rats. In addition to ACTH and B responses to restraint on d 5, body weight, food intake, fat depots, glucose and other hormones were measured to determine the role of stress-induced elevations in B on energy balance. ACTH responses to restraint were normal in intact rats subjected to diabetes or cold. By contrast, there was no ACTH or B response to restraint in NPY-infused intact rats. All 3 groups of chronically stimulated adrenalectomized rats with clamped B had facilitated ACTH responses to restraint compared to their treatment controls. Overall food intake increased in all groups of stressed rats; however, augmented intake occurred only during the light in intact rats and equally in the light and dark in B-clamped rats. White adipose depot weights were decreased by both diabetes and cold and increased by NPY in intact rats; the decreases with cold and increases with NPY were both blunted and changes in fat stores were not significant in adrenalectomized, B-clamped rats. We conclude that: 1. diabetes- and cold-induced facilitation of restraint-induced afferent input to hypothalamic control of the hypothalamo-pituitary-adrenal (HPA) axis is opposed in intact rats by the elevated feedback signal of B secretion; 2. NPY does not induce facilitation of afferent stress pathways; 3. chronic stimulation of the HPA axis induces acute hyperresponsiveness of hypothalamic neurons to restraint provided that the afferent input of this acute stimulus is not prevented by B feedback; 4. stimulus-induced elevations in B secretion result in day-time feeding; 5. insensitivity of both caloric efficiency and white fat stores to chronic stress in adrenalectomized, B-clamped rats results from loss of normally variable B levels.     

Restraint stress and corticotropin releasing hormone modulation of murine cutaneous POMC mRNA

The skin is a unique immunological defense barrier that protects the organism from occupational and environmental exposures and provides a model system in which to evaluate the interaction of the central nervous system with the peripheral immune response. In the studies presented here, we tested mild, acute restraint stress activation of the cutaneous corticotropin releasing hormone-pro-opiomelanocortin (CRH-POMC) axis. We verified that 2 h restraint stress increased the serum concentration of corticosterone and alpha-melanocyte stimulating hormone. We report for the first time that CRH upregulates POMC mRNA expression in mouse skin in vitro. We also demonstrated, by RT-PCR, that 2,4 di-nitrofluorobenzene (DNFB) upregulates cutaneous POMC mRNA expression, the production of which is suppressed by restraint stress. These data confirm the presence and functionality of two hormones of the hypothalamo-pituitary axis in the skin and suggest that activation of the central hypothalamo-pituitary-adrenal axis may over ride activation of the cutaneous CRH-POMC mechanism in the development of DNFB-stimulated allergic contact dermatitis.