Effects Of Corticotropin-releasing Hormone Research Articles

The renal effects of corticotropin releasing hormone: implication of vasopressin and atrial natriuretic peptide

To evaluate the hypothesized renal actions of corticotropin releasing hormone (CRH), adult male conscious normally hydrated or volume-expanded rats (6 ml saline) were injected iv with various doses of CRH (1–5 μg). Some groups of rats were injected with arginine vasopressin (AVP) antagonist (2 or 10 μg) iv prior to CRH (5 μg). Urine was collected for volume, Na, K, and cyclic guanosine monophosphate (cGMP) determination. AVP and atrial natriuretic pepride (ANP) were determined in blood by radioimmunoassay. Blood pressure (BP) was measured by telemetry. In both normally hydrated and volume expanded conscious rats, the biphasic effects of CRH injected iv were observed. Initially, within the first hour, CRH caused dose-dependent antidiuresis, decreased sodium, potassium and urinary cGMP excretion. Plasma AVP was already increased significantly (p < 0.01) within 5 min after injection from 3.9 ± 0.5 pg/ml to 15.5 ± 3.3 pg/ml (n = 15) at 15 min, then declined by 30 min to the concentration reached at 5 min and was maintained for 180 min. Changes in plasma ANP followed a mirror image of AVP. Plasma ANP correlated with the decrease of ANP synthesis in the left ventricle. Four hours after the injection, a marked diuresis, natriuresis, kaliuresis and an increase in urinary cGMP occurred. This was accompanied by increased ANP mRNA in the right atrium at 180 min. All urinary changes were reversed by a V2 receptor antagonist (2 or 10 μg) given iv. Control injection of saline evoked an immediate (3–5 min) increase in BP, and this elevation was markedly inhibited by CRH (5 μg). Heart rate was further increased. We hypothesize that the renal effects are mediated by vasodilation induced by CRH that decreases the baroreceptor input to the brain, leading to the rapid release of AVP which acts on the heart and inhibits ANP release and synthesis. These effects could be of physiological significance during the stress.

Immunocytochemical distribution of corticotropin-releasing hormone receptor type-1 (CRF(1))-like immunoreactivity in the mouse brain: light microscopy analysis using an antibody directed against the C-terminus.

Corticotropin-releasing hormone (CRH) receptor type 1 (CRF(1)) is a member of the receptor family mediating the effects of CRH, a critical neuromediator of stress-related endocrine, autonomic, and behavioral responses. The detailed organization and fine localization of CRF(1)-like immunoreactivity (CRF(1)-LI) containing neurons in the rodent have not been described, and is important to better define the functions of this receptor. Here we characterize in detail the neuroanatomical distribution of CRF(1)-immunoreactive (CRF(1)-ir) neurons in the mouse brain, using an antiserum directed against the C-terminus of the receptor. We show that CRF(1)-LI is abundantly yet selectively expressed, and its localization generally overlaps the target regions of CRH-expressing projections and the established distribution of CRF(1) mRNA, with several intriguing exceptions. The most intensely CRF(1)-LI-labeled neurons are found in discrete neuronal systems, i.e., hypothalamic nuclei (paraventricular, supraoptic, and arcuate), major cholinergic and monoaminergic cell groups, and specific sensory relay and association thalamic nuclei. Pyramidal neurons in neocortex and magnocellular cells in basal amygdaloid nucleus are also intensely CRF(1)-ir. Finally, intense CRF(1)-LI is evident in brainstem auditory associated nuclei and several cranial nerves nuclei, as well as in cerebellar Purkinje cells. In addition to their regional specificity, CRF(1)-LI-labeled neurons are characterized by discrete patterns of the intracellular distribution of the immunoreaction product. While generally membrane associated, CRF(1)-LI may be classified as granular, punctate, or homogenous deposits, consistent with differential membrane localization. The selective distribution and morphological diversity of CRF(1)-ir neurons suggest that CRF(1) may mediate distinct functions in different regions of the mouse brain.

Synergistic role of cAMP and IP 3 in corticotropin-releasing hormone-induced cell shape changes in invertebrate immunocytes

Immunocytes from the mollusc Mytilus galloprovincialis express corticotropin-releasing hormone (CRH) receptor subtype (CRH-R1 and CRH-R2)-like mRNAs. Using computer-assisted microscopic image analysis, we have found that exogenous CRH provokes changes in the cellular shape of immunocytes, and that this response is extracellular Ca 2+-dependent. The various inhibitors of transduction signaling pathways, i.e. suramin sodium, 2′,5′-dideoxyadenosine, neomycin sulfate, calphostin C, H-89, and wortmannin, completely or partially inhibit these changes. The present findings demonstrate that PKA, PKC, and PKB/Akt are involved in CRH-induced cell shape changes in immunocytes, and that the cellular effect of CRH needs the synergistic action of the two second messengers, cAMP and IP 3.

Corticotropin-releasing hormone increases the expression of the prostaglandin E(2) receptor subtype EP1 in amnion WISH cells.

The purpose of this study was to investigate the effect of corticotropin-releasing hormone (CRH) on the expression of the prostaglandin (PG) E(2) EP1 receptor subtype and PGE(2) production in amnion WISH cells (AWC). AWC cultures were incubated with CRH. Culture fluid was collected for PGE(2) measurement, and the cells were collected and analyzed for EP1 protein and mRNA. Immunohistochemical localization of the EP1 receptor was also performed. Incubation of AWC with CRH resulted in a dose-dependent increase (r = 0.97) in the level of EP1 receptor protein (P < 0.001). Coincubation of AWC with CRH and indomethacin resulted in the decreased production of PGE(2) while having no effect on EP1 receptor expression. A significant but not dose-dependent increase in EP1 mRNA expression was also observed (P < 0.01). Immunohistochemical evaluation verified cell membrane localization of the receptor in both stimulated and unstimulated cells and confirmed the increased expression of EP1 receptor in response to CRH. Incubation of AWC with CRH also resulted in increased culture fluid PGE(2) levels (P < 0.01). These results suggest that the role CRH plays in the initiation of labor may also involve the promotion of elevated PGE(2) levels and increased expression of the EP1 receptor in amnion.

Corticotropin-releasing hormone-binding protein in brain and pituitary of food-deprived obese (fa/fa) Zucker rats.

The present study was conducted to verify whether experimental conditions such as obesity and food deprivation, which promote food intake and reduce thermogenesis, could modify the expression of the corticotropin-releasing hormone (CRH)-binding protein (BP) in the rat brain. In situ hybridization, histochemistry, and immunohistochemistry were used to assess the expression of CRH-BP in lean (Fa/?) and obese (fa/fa) Zucker rats that were fed ad libitum, food deprived for 24 h, or food deprived for 24 h and refed for 6 h. In both lean and obese rats, food deprivation led to a reduction in body weight that was accompanied by a reversible increase in plasma corticosterone levels. Food deprivation and, to a lesser degree, obesity induced the expression of CRH-BP mRNA in the dorsal part of the medial preoptic area (MPOA). This induction of the CRH-BP gene led to by food deprivation was confirmed by the appearance in the dorsal part of the MPOA of neurons immunoreactive to CRH-BP. Food deprivation (in particular) and obesity also increased the levels of CRH-BP mRNA in the basolateral amygdala (BLA). The enhanced CRH-BP expression in the MPOA and BLA in response to food deprivation was reversed by refeeding. In lean Fa/? rats, the CRH-BP mRNA level in the pituitary cells was significantly decreased after food deprivation and restored after refeeding. When food was provided ad libitum, the number of cells expressing CRH-BP in the anterior pituitary was significantly higher in lean rats than in obese animals. Food deprivation for 24 h decreased dramatically the number of pituitary cells expressing CRH-BP in lean rats. Altogether, the present results demonstrate that food deprivation and, to a lesser extent, obesity can selectively affect the expression of CRH-BP. Given both the inactivating effect of CRH-BP on the CRH system and the potential roles played by the MPOA and BLA in the thermogenic and anorectic effects of CRH, it can be argued that the induction of the CRH-BP gene in obesity and after food deprivation occurs as a mechanism to reduce energy expenditure and to stimulate food intake.

Pituitary CRH Receptor Blockade Reduces Waking in the Rat

We have previously hypothesized that corticotropin-releasing hormone (CRH) is involved in the regulation of physiological waking. Central administration of CRH receptor antagonists reduces spontaneous waking in the rat. Some of the responses to central administration of CRH receptor antagonists may be mediated by mechanisms involving the hypothalamic–pituitary–adrenal axis, either by direct actions on the hypothalamus or by actions at the level of the pituitary due to leakage of the antagonists from the cerebrospinal fluid to blood. To further clarify the role of the hypothalamic–pituitary–adrenal axis as a mediator of responses to CRH receptor blockade, we administered intravenously into freely behaving rats in their home recording cages two specific CRH receptor antagonists, astressin or α-helical CRH 9–41, and determined subsequent changes in waking and sleep. Our results indicate that both antagonists reduce spontaneous waking, but with different time courses. Astressin, a potent antagonist of pituitary CRH receptors, reduces waking during postinjection hours 9–10, whereas high doses of α-helical CRH 9–41 reduce waking only during the first postinjection hour. These results indicate that some effects of CRH on sleep–wake behavior may be meditated by pituitary CRH receptors.

Infantile Spasms: Hypothesis-Driven Therapy and Pilot Human Infant Experiments Using Corticotropin-Releasing Hormone Receptor Antagonists

Background and Rationale: Infantile spasms (IS) are an age-specific seizure disorder occurring in 1:2,000 infants and associated with mental retardation in ∼90% of affected individuals. The costs of IS in terms of loss of lifetime productivity and emotional and financial burdens on families are enormous. It is generally agreed that the seizures associated with IS respond poorly to most conventional anticonvulsants. In addition, in the majority of patients, a treatment course with high-dose corticotropin (ACTH) arrests the seizures completely within days, often without recurrence on discontinuation of the hormone. However, the severe side effects of ACTH require development of better treatments for IS. Based on the rapid, all-or-none and irreversible effects of ACTH and on the established physiological actions of this hormone, it was hypothesized that ACTH eliminated IS via an established neuroendocrine feedback mechanism involving suppression of the age-specific endogenous convulsant neuropeptide corticotropin-releasing hormone (CRH). Indeed, IS typically occur in the setting of injury or insult that activate the CNS stress system, of which CRH is a major component. CRH levels may be elevated in the IS brain, and the neuropeptide is known to cause seizures in infant rats, as well as neuronal death in brain regions involved in learning and memory. If ‘excess’ CRH is involved in the pathogenesis of IS, then blocking CRH receptors should eliminate both seizures and the excitotoxicity of CRH-receptor-rich neurons subserving learning and memory. Patients and Methods: With FDA approval, α-helical CRH, a competitive antagonist of the peptide, was given as a phase I trial to 6 infants with IS who have either failed conventional treatment or who have suffered a recurrence. The study was performed at the Clinical Research Center of the Childrens Hospital, Los Angeles. The effects of α-helical CRH on autonomic parameters (blood pressure, pulse, temperature, respiration) were determined. In addition, immediate and short-term effects on ACTH and cortisol and on electrolytes and glucose were examined. The potential efficacy of α-helical CRH for IS was studied, using clinical diaries and video EEG. Results: α-Helical CRH, a peptide, did not alter autonomic or biochemical parameters. Blocking peripheral CRH receptors was evident from a transient reduction in plasma ACTH and cortisol. No evidence for the compound’s penetration of the blood-brain barrier was found, since no central effects on arousal, activity or seizures and EEG patterns were observed. In addition, a striking resistance of the patients’ plasma ACTH to the second infusion of α-helical CRH was noted. Conclusions: Peptide analogs of CRH do not cross the blood-brain barrier, and their effects on peripheral stress hormones are transient and benign. Nonpeptide compouds that reach CNS receptors are required to test the hypothesis that blocking CRH receptors may ameliorate IS and its cognitive consequencs.

Corticotropin-releasing-hormone lacks analgesic properties: an experimental study in humans, using non-inflammatory pain

The antinociceptive potency of corticotropin-releasing-hormone (CRH) has been established in several animal studies in which both central and peripheral sites of action were considered. However, there have not yet been any experimental trials, besides one attempt using clinical dental pain demonstrating the potential analgesic properties of CRH in humans. For this reason, we studied the effect of CRH on experimental heat pain sensitivity in 18 healthy men, using a double-blind, cross-over and placebo-controlled design. A dose of 100 μg (i.v.) was chosen because of its well-known neuroendocrine effects in humans. The pain parameters assessed were, visual analog scale (VAS) ratings for pain intensity and pain unpleasantness, pain thresholds and scores for discrimination ability. To differentiate between a direct analgesic effect of CRH and indirect effects via evoked hormonal responses in the hypothalamic-pituitary-adrenocortical (HPA) system ( β-endorphin, ACTH, cortisol), CRH was applied with and without a pre-treatment with dexamethasone. In neither of the two conditions was there any systematic change in our pain parameters. This failure to find any evidence suggesting an analgesic action of CRH or of the subsequent hormones of the HPA system was obtained despite the fact that CRH produced clear neuroendocrine responses such as increases in the plasma concentration of β-endorphin and cortisol. It is unclear whether the lack of analgesic action of CRH is due to its non-existence in humans, due to the use of a pain model which does not assess minute changes in pain sensitivity and does not trigger substantial inflammatory responses, or due to an insufficient dose of CRH.

Corticotropin-releasing hormone stimulates steroidogenesis in cultured human adrenal cells

The effects of corticotropin releasing hormone (CRH) on steroid production by cultures of human fetal adrenal cells was investigated. We found that CRH, at concentrations that have been reported to exist in human fetal serum, stimulated dehydroepiandrosterone sulfate (DS) and cortisol production by cultured fetal zone and neocortical zone cells. A dose-dependent increase in secretion of both steroids was noted, with the cortisol pathway being preferentially enhanced by CRH at high concentrations. Pretreatment of adrenal cells for 3 days made them more responsive to ACTH stimulation and such effects were dose-dependent also. Inclusion of the antagonist, α-helical CRH (9–41) blocked CRH-induced stimulation of DS and cortisol over a broad dose range and also interfered with the augmentation of cortisol secretion noted after ACTH in CRH treated cells. CRH had no effects on adrenal cell proliferation or total cell protein. These studies are suggestive that CRH, either of systemic origin or else produced within the adrenal itself, has the potential to be a modulator of adrenal steroid production in the human.

Decreased Type 2 Corticotropin-Releasing Hormone Receptor mRNA Expression in the Ventromedial Hypothalamus during Repeated Immobilization Stress

Chronic or repeated stress results in reduction of food intake and body weight in rats. Stress-induced anorexia has been attributed to increased corticotropin-releasing hormone (CRH) function in the central nervous system. To explore possible roles of other neuropeptides and peripheral hormones involved in food intake and energy utilization during continuing stress, we examined the impact of repeated immobilization stress on expression of mRNAs coding for CRH, neuropeptide Y (NPY), galanin and pro-opiomelanocortin (POMC) mRNAs in such hypothalamic nuclei as the paraventricular nucleus (PVN), arcuate nucleus (ARC) and dorsomedial hypothalamus (DMH), as well as plasma insulin and leptin concentrations. Changes in type 2 CRH receptor (CRHR-2) mRNA in the ventromedial hypothalamus (VMH), a possible target of anorectic CRH effect, were also examined. Rats were immobilized for 2 h daily for 6 days and sacrificed 24 h after the last immobilization. Immobilized rats had lower food intake and body weight and higher levels of PVN CRH mRNA than controls. Repeated immobiliza tion also lowered plasma insulin and leptin concentrations and VMH CRHR-2 mRNA levels. These results provide additional evidence linking VMH CRHR-2 mRNA levels to plasma leptin concentration. ARC NPY and DMH galanin mRNAs increased following repeated immobilization, while ARC POMC mRNA decreased. DMH NPY mRNA and ARC galanin mRNA were unaltered by immobilization. Since NPY and galanin are considered orexigenic, while the POMC-melanocortin-4 receptor system is apparently anorexigenic, the changes in neuropeptide mRNAs and VMH CRHR-2 mRNA may play counterregulatory roles against anorectic CRH effects.

Central administration of corticotrophin releasing hormone but not arginine vasopressin stimulates the secretion of luteinizing hormone in rams in the presence and absence of testosterone.

This study tested the hypothesis that central administration of corticotrophin-releasing hormone (CRH) and/or arginine vasopressin (AVP) will affect the secretion of LH in rams and that testosterone is necessary for these actions to occur. Plasma LH levels were measured in castrated rams during 1 h infusion of either 100 microliter vehicle/mock cerebrospinal fluid (CSF) or mock CSF containing 25 microgram CRH, 25 microgram AVP or 25 microgram of each peptide through guide cannulae into the third cerebral ventricle. These intracerebroventricular (i.c.v.) infusions were given to the castrated rams following injections (i.m.) each 12 h of oil or 8 mg testosterone propionate for 7 days. Blood samples were collected every 10 min for 4 h before i.c.v. infusion, during infusion and for 4 h following the infusion. Infusion of vehicle did not affect any endocrine parameters. In contrast, the plasma concentrations of LH and the amplitude of LH pulses were increased significantly during and following infusion of CRH, and this effect was not influenced by whether the castrated rams were treated with testosterone propionate or whether the CRH was administered in combination with AVP. Infusion of AVP alone did not affect LH secretion. The frequency of LH pulses and the plasma concentrations of FSH did not change with any of the i.c.v. treatments. The plasma concentrations of cortisol were significantly increased by CRH and AVP infusions. The plasma concentrations of cortisol achieved during and following i.c.v. infusion of CRH and AVP combined were greater than the concentrations achieved as a result of treatment with AVP alone but were similar to those with CRH. There was no effect of testosterone propionate on cortisol levels. These results show that CRH, but not AVP, is capable of acting either centrally or at the pituitary level to increase the secretion of LH in rams and these actions are not affected by testosterone. The stimulatory effects of CRH on LH secretion are to increase the amplitude of GnRH pulses and/or the responsiveness of the pituitary to the actions of GnRH with no effect on the frequency of GnRH pulses. The secretion of FSH in rams is not influenced by either CRH or AVP. The effect of CRH to increase LH pulse amplitude occurs in the face of increased cortisol levels, further reinforcing our belief that this adrenal steroid does not affect the reproductive axis in this species.

Corticotropin-releasing hormone acts on guinea pig ileal smooth muscle via protein kinase A.

In contraction studies corticotropin-releasing hormone (CRH) was found to relax ileal but not gastric and jejunal smooth muscles of the guinea-pig, precontracted with BaCl2. Under whole-cell patch-clamp conditions, CRH concentration-dependently activated Ca2+-sensitive K+ currents (IK) with ED50=20 pM at 100 nM and ED50=0. 13 pM at 500 nM intracellular Ca2+ respectively. This increase was accompanied by significant hyperpolarization of the cell membranes. CRH 9-41 peptide fragment did not affect IK amplitude, membrane potential or contraction. The CRH-induced increase of IK densities was accelerated in the presence of high intracellular Ca2+ concentrations (500 nM) and was abolished by pretreatment of cells with either ryanodine or thapsigargin, which cause depletion of intracellular Ca2+ stores, as well as in cells treated under conditions prohibiting intracellular Ca2+ store refilling. The effect of CRH on IK was not affected by bath application of various selective inhibitors of membrane-bound phospholipases, protein kinase C, cGMP-dependent protein kinase or Ca2+/calmodulin-dependent protein kinase II, but was effectively antagonized by blockers of protein kinase A (PKA) or adenylyl cyclase. Neither forskolin nor the catalytic subunit of PKA could mimic the effect of CRH on IK. Thus, it was suggested that CRH exerts its relaxing activity on ileal smooth muscle cells via PKA-dependent phosphorylation of some intracellular target coupled to sarcoplasmic reticulum Ca2+ storage machinery.

Stress, corticotropin-releasing hormone, glucocorticoids, and the immune/inflammatory response: acute and chronic effects.

Corticotropin-releasing hormone (CRH) influences the immune system indirectly, through activation of the hypothalamic-pituitary-adrenal axis and sympathetic system, and directly, through local modulatory actions of peripheral (immune) CRH. We recently demonstrated that catecholamines and histamine potently inhibited interleukin (IL)-12 and stimulated IL-10, whereas glucocorticoids suppressed IL-12, but did not affect IL-10 production ex vivo. Thus, both glucocorticoids and catecholamines, the end products of the stress system, and histamine, a product of activated mast cells, may selectively suppress cellular immunity and favor humoral immune responses. We localized immunoreactive CRH in experimental carrageenin-induced aseptic inflammation and, in humans, in inflamed tissues from patients with several autoimmune disease. In addition, we demonstrated that CRH activated mast cells via a CRH receptor type 1-dependent mechanism, leading to release of histamine and hence vasodilatation and increased vascular permeability. Thus, activation of the stress system, through direct and indirect effects of CRH, may influence the susceptibility of an individual to certain autoimmune, allergic, infectious or neoplastic diseases. Antalarmin, a novel nonpeptide CRH antagonist, prevented several proinflammatory effects of CRH, thus revealing its therapeutic potential in some forms of inflammation.

Effects of corticotrophin releasing hormone on contractile activity of myometrium from pregnant women.

To determine whether corticotrophin releasing hormone plays a role in the regulation of tone in term nonlabouring human myometrium. A teaching hospital research laboratory. Thirty-seven women undergoing elective nonlabour caesarean section under regional anaesthesia. Human corticotrophin releasing hormone (1, 10, 100 nmol/L) was added to strips of term, nonlabouring myometrium mounted in an organ bath, and the effect on spontaneous, oxytocin (1 nmol/L) or prostaglandin F2alpha (100 nmol/L) stimulated contractions determined. Cyclic adenosine monophosphate (cAMP) content of the tissue was also determined by enzyme immunoassay. Corticotrophin releasing hormone did not affect myometrial tension development in any of the experimental protocols. cAMP increased transiently after addition of corticotrophin releasing hormone (166.7 +/- 12.7% at 1 minute) but this was not reflected by any change in tension. This study suggests that despite high maternal plasma concentrations of corticotrophin releasing hormone in pregnancy at term, this peptide is unlikely to play a direct role in the control of myometrial contractility in nonlabouring myometrium.

Evidence that corticotropin-releasing hormone acts as a growth hormone-releasing factor in a primitive teleost, the European eel (Anguilla anguilla).

The inhibitory control of growth hormone (GH) release by somatostatin (SRIH) has been conserved throughout vertebrate evolution. In contrast, the neuropeptides involved in the stimulatory control of GH vary according to species and/or physiological situations. We investigated the direct pituitary regulation of GH release in a primitive teleost, the European eel (Anguilla anguilla L.) at the juvenile stage. Short-term serum-free primary cultures of dispersed pituitary cells were used, and GH release was measured by an homologous radioimmunoassay. Whereas growth hormone-releasing hormone (GHRH), gonadotropin-releasing hormone (GnRH), thyrotropin-releasing hormone (TRH), neuropeptide Y (NPY) and cholecystokinin (CCK) failed to induce any change in GH release, corticotropin-releasing hormone (CRH) dose-dependently stimulated GH release with a significant effect at 1 nM and a maximal effect (> or =400% of controls at 24 h) at 100 nM. In agreement with our previous studies, PACAP also stimulated GH release but its maximal effect was lower than that of CRH. Proopiomelanocortin (POMC)-peptides, corticotropin (ACTH), melanotropin (alpha-MSH), beta-endorphin) had no effect on GH release, at any dose tested (0.1-1000 nM), indicating that the stimulatory effect of CRH on GH release by somatotrophs was not mediated by CRH-induced release of POMC-peptides from corticotrophs and melanotrophs. The CRH antagonist, alpha-helical CRH(9-41), significantly inhibited the stimulatory effect of CRH on GH release, suggesting the implication of specific CRH receptors related to mammalian ones. The stimulatory effect of CRH on GH release was reduced after 24 h of incubation, indicating a desensitization. In contrast, no desensitization to the inhibitory effect of SRIH was observed. SRIH inhibited CRH action in a dose-dependent manner. The effect of SRIH was overriding, 1 nM SRIH being able to abolish the effect of 1000 nM CRH. In conclusion, in the eel, CRH stimulates GH release directly at the pituitary cell level. GH and cortisol secretions could interact in controlling several physiological functions such as metabolism and ion exchange. This study suggests that CRH may have played an important early role in vertebrates co-ordinating the activation of various endocrine axes involved in metamorphosis, osmoregulation, stress and fasting. The stimulatory role of CRH on GH release may have been partially conserved during evolution, as it is found in some human physio-pathological situations such as stress, fasting and depression.

Leptin effects on the expression of type-2 CRH receptor mRNA in the ventromedial hypothalamus in the rat.

The product of the ob gene, leptin, is thought to act in the hypothalamus to reduce food intake and body weight (b.w.) in rats and mice; however, the mechanisms of leptin action in the brain have not been fully elucidated. Corticotropin-releasing hormone (CRH) is a potent anorectic neuropeptide, and its type-2 receptor (CRHR-2) in the ventromedial hypothalamus (VMH) appears to play an important role in the expression of this anorectic effect. We explored here the impact of systemic leptin administration on CRH mRNA expression in the hypothalamic paraventricular nucleus (PVN) and CRHR-2 mRNA expression in the VMH in male rats, using in-situ hybridization histochemistry. The expression of CRH mRNA in the PVN and CRHR-2 mRNA in the VMH were increased at 2 h and 6 h, respectively, after a single intraperitoneal injection of leptin (1.0 mg/kg). Continuous subcutaneous infusion of leptin (1.2 mg/kg/day) via an osmotic minipump for 5 days increased the expression of CRHR-2 mRNA in the VMH, but not the expression of CRH mRNA in the PVN, compared with vehicle treatment. The rats that received the single or continuous administration of leptin showed reductions of food intake and b.w. compared with vehicle-treated rats. These results are consistent with our previous findings that the expression of CRHR-2 mRNA in the VMH is positively correlated with plasma leptin concentrations under various conditions, and highlight the importance of circulating leptin for the regulation of VMH CRHR-2 mRNA. The present results also raise the possibility that leptin reduces food intake and b.w. at least partially due to the enhancement of the anorectic effect of CRH via increased PVN CRH expression and/or VMH CRHR-2 expression.

Changes in hippocampal theta following intrahippocampal corticotropin-releasing hormone (CRH) infusions in the rat

Hippocampal theta activity is a large amplitude, sinusoidal wave that occurs during attentive immobility and exploratory behaviour in the rat, and it is thought to be involved in memory formation. Recent reports suggest that corticotropin-releasing hormone (CRH) has pro-mnemonic effects in rodents. Because memory-enhancing substances/manipulations generally alter either theta frequencies or amplitudes, these variables were monitored in urethane-anaesthetised rats following intra-hippocampal infusions of CRH. Adult male, Lister hooded rats were implanted with a hippocampal recording electrode and a guide cannula, both aimed at the dentate gyrus. When CRH was infused into the hippocampus, the main change in the hippocampal EEG was a slow onset increase in the amplitude of spontaneous theta and, paradoxically, a significant decrease in the amount of time spent displaying theta. These data suggest that CRH has the ability to modulate ongoing hippocampal theta, but, considering the slow effect, the involvement of hippocampal CRH receptors is suspect. Regardless of locus, the described electrophysiological changes suggest that hippocampal cholinergic systems may play a role in the memory-enhancing effects of CRH.

Differential behavioural effects of chronic infusion of CRH 1 and CRH 2 receptor antisense oligonucleotides into the rat brain

The purpose of this study was to investigate the functional role the two corticotropin-releasing hormone (CRH) receptor subtypes play in regulating the behavioural performance of rats in various well-defined test situations. Antisense oligodeoxynucleotides (ODNs) corresponding to either the rat CRH1 or CRH2 receptor mRNA were infused chronically into the lateral ventricle of male rats via osmotic minipumps (5 μg/0.5 μl/h over 6 days). Control groups received infusions of either a scrambled sequence ODN or mixed bases ODN or vehicle. On day 4 after surgery, the rats were subjected to 10 min of social defeat and immediately afterwards tested on the elevated plus-maze. Compared to a scrambled sequence control ODN, CRH1 receptor antisense ODN infusion was found to exert an anxiolytic-like effect whereas CRH2 receptor antisense ODN infusion had no effect on defeat-induced anxiety-related behaviour. In contrast, the CRH2 receptor antisense ODN increased immobility in a forced swim test whereas CRH1 receptor ODN-treated rats did not differ from controls. No influence of either ODN was found on general locomotor activity in an open field or on short-term memory performance in a social discrimination test. Furthermore, the CRH2 receptor antisense ODN did not affect spatial learning in a Morris water maze task. An additional experiment comparing a mixture of both missense ODNs and a vehicle control group confirmed that the former failed to induce non-specific (toxic) side effects, further substantiating the specificity of the respective antisense effects measured in this study. The results support the hypothesis that the two CRH receptor subtypes selectively mediate differential effects of endogenous CRH or CRH-related peptides at the brain level with the CRH1 receptor contributing predominantly to emotional behaviour and the CRH2 receptor being involved in the regulation of stress coping behaviour.

Effects of acute and chronic fluoxetine treatment of CRH-induced anxiety.

The effects of acute and chronic fluoxetine treatment on anxiogenic effect of corticotropin-releasing hormone (CRH) were studied in the social interaction test in male Sprague-Dawley rats. Injection of CRH (100 ng, i.c.v.), fluoxetine (5 mg/kg, i.p.) or their combination had significant anxiogenic effects (decrease in total interaction time and increase in self-grooming) in the social interaction test. The effects of CRH and fluoxetine in combination were not additive. Fluoxetine, but not CRH significantly inhibited locomotor activity. After chronic fluoxetine treatment (5 mg/kg/day, 21 days) the acute anxiogenic effects of either CRH or fluoxetine were abolished. Our studies provide evidence for a 5-HT/CRH interaction in the regulation of anxiety.

Differential Behavioral and Endocrinological Effects of Corticotropin-Releasing Hormone (CRH) in the Syracuse High- and Low-Avoidance Rats

The Syracuse high- and low-avoidance rats, which have been selectively bred for good (SHA/Bru) or poor (SLA/Bru) avoidance learning in a two-way shuttle box, differ in emotionality. This experiment investigated the effect of corticotropin-releasing hormone (CRH), administered centrally (0, 0.1, 0.5, and 1.0 μg), on conditioned suppression and on the hypothalamic–pituitary–adrenocortical system. Three groups of animals were used: SHA/Bru rats conditioned at 0.21 or 0.43 mA and SLA/Bru rats conditioned at 0.21 mA. The results confirm those of previous studies which found that SLA/Bru rats show greater conditioned suppression than the SHA/Bru rats at the low shock intensity and that at 0.43 mA, the SHA/Bru animals acquire a level of conditioning comparable to that of the SLA/Bru animals at 0.21 mA. The results show that the nonlinear behavioral effect of CRH is independent of strain and produces comparable effects in animals of both strains, but only when level of conditioning is equated. Adrenal and plasma concentrations of corticosterone increased in all three groups of animals as a direct linear function of dose of CRH. Both greater levels of conditioning and larger amounts of CRH increase the synthesis of corticosterone more in SHA/Bru animals than in the SLA/Bru animals. Thus, genetic variation, which differentiates the behavioral and endocrinological characteristics of these animals, shows that these effects of CRH can be independent of each other and suggests that some minimal level of conditioned fear is necessary for CRH to exert its anxiogenic effect.