Corticotropin-releasing Hormone Receptor mRNA Research Articles

Effects of continuous infusion of interleukin 1 beta on corticotropin-releasing hormone (CRH), CRH receptors, proopiomelanocortin gene expression and secretion of corticotropin, beta-endorphin and corticosterone.

A number of recent studies suggest that interleukin 1 beta (IL-1 beta) is a major mediator of hypothalamo-pituitary-adrenal (HPA) responses following infectious aggression. We investigated whether IL-1 beta mediates long-term changes in HPA activity and studied the cellular regulation of the anterior pituitary. To mimic chronically elevated IL-1 beta production thought to occur during infectious diseases, osmotic pumps (Alzet type) were implanted in the peritoneal cavity of male rats and hIL-1 beta was infused continuously at rates of 1 or 3 micrograms/day. Effects of hIL-1 beta action on plasma ACTH, beta-endorphin (beta-EP) and corticosterone (CORT) secretion and on anterior pituitary (AP), ACTH and beta-EP content were followed. In addition, hypothalamic (HT) CRH mRNA and in AP, CRH receptor (CRH-Rc) mRNA, POMC nuclear primary transcript RNA, POMC nuclear intermediate processing RNA and POMC nuclear and cytoplasmic mRNA were quantified using a highly sensitive solution hybridization nuclease protection assay. Continuous infusion of hIL-1 beta stimulated the HPA axis at varying degrees. Increased HT CRH gene expression, AP POMC gene transcription, ACTH and beta-EP release occurred only during the first 3 days of the treatment. A long-lasting enhancement of ACTH and beta-EP synthesis and of POMC gene expression resulted from activated POMC gene transcription followed by an increased POMC mRNA stability and decreased POMC mRNA turnover. In the AP, stimulation of ACTH and beta-EP secretion and POMC gene transcription disappeared after continuous IL-1 beta treatment, possibly in part due to a refractory process mediated by decreased CRH-Rc gene expression in corticotropes.

Regulation of messenger ribonucleic acid for corticotropin releasing hormone receptor in the pituitary during stress.

The mechanism regulating pituitary CRH receptors during stress was studied by analysis of the changes in CRH receptor messenger RNA (mRNA) and CRH binding after acute and repeated stress and CRH and vasopressin (VP) administration in intact and adrenalectomized rats. Acute stress caused time- and stress type-dependent changes in pituitary CRH receptor expression. In situ hybridization studies showed biphasic changes in CRH receptor mRNA after immobilization stress for 1 h and decreases by 2 h (P < 0.01). Increases (P < 0.01) were seen 4 and 8 h after the initiation of the stress, and a return to near basal levels by 12 and 18 h. A different pattern, with a decrease by 4 h (P < 0.01) and levels similar to controls after 12 and 18 h, was observed after a single ip injection of hypertonic saline (1.5 M NaCl). Binding autoradiography showed significant increases in pituitary CRH binding 4, 10, and 12 h after immobilization stress, but significant decreases 4, 12, and 18 h after ip hypertonic saline. In contrast, repeated immobilization or ip hypertonic saline for 8 or 14 days increased pituitary CRH receptor mRNA, and CRH binding was decreased. To determine the role of hypothalamic CRH and VP on these stress-induced changes, rats were injected for 14 days with CRH, VP, or their combination at doses mimicking stress levels in pituitary portal circulation (1 microgram/day sc). Repeated injection of CRH or VP increased CRH receptor mRNA and CRH binding (P < 0.05). CRH receptor mRNA levels further increased after combined administration of CRH and VP (P < 0.01), but CRH binding showed a tendency to decrease. The role of glucocorticoids on CRH receptor regulation was studied by analysis of the effects of stress on CRH receptor mRNA and CRH binding in adrenalectomized (ADX) rats with and without corticosterone replacement in the drinking water. Although in 6-day ADX rats pituitary CRH receptor mRNA levels were markedly reduced after acute immobilization, glucocorticoid replacement restored the stimulatory effect of stress to levels observed in intact rats. Similarly, a single sc injection of CRH (1 microgram) decreased CRH receptor mRNA in ADX rats but not in glucocorticoid-replaced ADX rats. CRH binding showed the expected decrease after ADX and was unchanged after stress or CRH injection. The increased pituitary CRH receptor mRNA after stress suggests that stress-induced CRH receptor down-regulation is due to increased receptor occupancy and internalization rather than to a decrease in receptor synthesis. The data suggest that increased hypothalamic secretion of CRH and VP mediates the delayed up-regulatory effect of stress on CRH receptor mRNA, and that resting levels of glucocorticoids are required for this effect. In addition, increased VP levels are permissive for the down-regulation of CRH binding induced by chronic pituitary exposure to stress levels of CRH.

Age-related changes in the expression of corticotropin-releasing hormone receptor mRNA in the rat pituitary.

Hypothalamic corticotropin releasing hormone (CRH) controls the release of adrenocorticotrophic hormone (ACTH) from the pituitary and, and therefore has a major role in the activation of the hypothalamic-pituitary-adrenal (HPA) axis. The HPA axis function has been shown to be impaired in the neonatal period as well as in aging. Since corticotropin-releasing hormone receptor (CRHr) plays a crucial role in the regulation of HPA axis, using in situ hybridization histochemistry we have analyzed the rat pituitary for the presence of CRHr mRNA in the neonatal period and during aging. The results show an increase in CRHr mRNA in 3-day-old rats, with a progressive increase within the first month. In the aging rat, we observed a down-regulation of the CRHr mRNA localized in the anterior pituitary, gland viceversa, an increased signal in the intermediate lobe. Our findings demonstrate age-related changes in the expression of the CRHr mRNA in the pituitary, with a differential regulation in the anterior and intermediate lobes of aging rats.

Neural regulation of corticotropin releasing hormone (CRH) and CRH receptor mRNA in the hypothalamic paraventricular nucleus in the rat.

The role of afferent innervation to the hypothalamic paraventricular nucleus (PVN) on CRH mRNA and CRH receptor mRNA levels was studied in control and stressed rats. Groups of rats were subjected to unilateral transection of the stria terminalis (ST), the medial forebrain bundle at the rostral hypothalamic level (RMFB), or the lower brainstem through the medulla oblongata between the obex and the locus coeruleus (CBs). Twelve days after surgery, each group of rats was further divided into controls (basal conditions) and stressed (1 h immobilization), before collecting brains for mRNA analysis by in situ hybridization histochemistry. While ST and RMFB cuts had no effect on basal CRH mRNA levels in the PVN, CBs cut decreased CRH mRNA in the PVN ipsilaterally to the knife cut but it was without effect on the contralateral side (-40% and -37% vs contralateral and sham-operated, respectively, P < 0.01). Acute stress (rats were killed 3 h after immobilization) increased CRH mRNA levels by about 30% bilaterally, an effect which was unchanged by any of the three hemisections. Under basal conditions, CRH receptor mRNA levels in the PVN were indistinguishable from the surrounding areas in sham-operated controls, ST and RMFB operated rats. However, brainstem hemisection resulted in clear expression of CRH receptor mRNA in areas consistent with the dorsal, medial-ventral and lateral parvicellular subdivisions of the PVN, ipsilateral to the transection. CRH neurons in these subdivisions project to the lower brainstem and the spinal cord. Expression of CRH receptor mRNA in the medial-dorsal and anterior parvicellular divisions (CRH neurons with median eminence projections) was not affected by CBs cut. In these subdivisions, immobilization stress markedly increased CRH receptor mRNA levels but it did not influence CBs cut-induced CRH receptor expression. ST and RMFB hemisections were without effect on PVN CRH receptor mRNA levels under basal or stress conditions. Oxytocin (OT) and vasopressin (VP) mRNA levels in the magnocellular subdivision of the PVN were unchanged after immobilization, or following ST, RMFB or CBs cuts, whereas OT mRNA in the medial-ventral and caudal parvicellular subdivisions was decreased by 52% after CBs cut. The data demonstrate that: 1) basal CRH mRNA levels in the PVN are under tonic stimulatory influence of the lower brainstem (and/or spinal cord) afferents; 2) CRH receptor mRNA expression in PVN subdivisions (pituitary vs lower brainstem/spinal cord projecting neurons) is under different control mechanisms, and 3) immobilization-induced changes in CRH mRNA and CRH receptor mRNA levels are mediated either by neural inputs from brain areas other than those investigated here, or by humoral factors.

Developmental profile of messenger RNA for the corticotropin-releasing hormone receptor in the rat limbic system

The ontogeny of corticotropin-releasing hormone (CRH) receptor messenger ribonucleic acid (mRNA) in rat brain, using in situ hybridization, is the focus of this study. The developmental profile of CRH receptor using binding assays and receptor autoradiography has been reported, but may be confounded by the presence of a binding protein. The recent cloning of the rat CRH receptor gene has permitted the use of in situ hybridization histochemistry to map the distribution of cells expressing CRH receptor mRNA in the developing brain. We used antisense 35S-labeled oligodeoxynucleotide probes for the two reported splice-variants of the CRH receptor mRNA, which yielded essentially identical localization patterns. CRH receptor mRNA was clearly detectable in infant brain starting on the second postnatal day. Signal in hippocampal CA1, CA2 and CA3a increased to 300–600% of adult levels by postnatal day 6 with a subsequent gradual decline. In the amygdala, in contrast, CRH receptor mRNA abundance increased steadily between the second and the ninth postnatal days, to levels twice higher than those in the adult. In the cortex, CRH receptor mRNA levels were high on postnatal day 2 and decreased to adult levels by day 12. Transient signal over the hypothalamic paraventricular nucleus, observed on the second postnatal day, was not evident at older ages. These results demonstrate robust synthesis of CRH receptor as early as on the second postnatal day and unique region-specific developmental profiles for CRH receptor gene expression.

Hypothalamic-pituitary corticotroph function after shunting of magnocellular vasopressin and oxytocin to the hypophyseal portal circulation.

Surgical pituitary stalk compression (PSC) causes neural lobe denervation and development of ectopic magnocellular terminals in the external zone of the median eminence, resulting in shunting of magnocellular vasopressin (VP) and oxytocin (OT) to the pituitary portal circulation. To determine the effects of PSC on hypothalamic and pituitary function, VP and CRH receptors and POMC messenger RNA (mRNA) in the pituitary, and CRH, VP, and OT mRNA levels in the PVN were examined in 7-day PSC and sham-operated rats. Immunohistochemical staining revealed marked accumulation of immunoreactive VP in the swollen pituitary stalk and the external zone of the distal median eminence of PSC rats. Plasma ACTH and corticosterone levels were significantly increased by PSC, an effect that was attenuated by minipump infusion of desamino-[D-Arg8]-vasopressin (DDAVP) given to correct the diabetes insipidus. [3H]VP binding to anterior pituitary membranes from PSC rats was reduced by 50% compared to that in sham-operated controls, but VP V1b receptor mRNA levels measured by in situ hybridization were unchanged. Pituitary CRH receptors measured by binding autoradiography and CRH receptor mRNA levels did not change after PSC. POMC mRNA was unchanged in the anterior pituitary lobe, but markedly increased in the intermediate lobe of PSC rats, with or without DDAVP infusion. In the hypothalamic supraoptic and paraventricular nuclei, PSC resulted in reduction of VP mRNA (-83%) and OT mRNA (-38%) levels, probably reflecting retrograde degeneration of magnocellular neurons. This decrease was more pronounced for VP mRNA than for OT mRNA CRH mRNA levels in parvicellular cells of the paraventricular nuclei of PSC rats were reduced by 55%. Correction of the diabetes insipidus by DDAVP treatment further decreased hypothalamic VP mRNA levels, but restored CRH mRNA to control levels. The data show that continuous exposure of the pituitary to high VP levels from ectopic magnocellular fibers in the median eminence results in VP, but not CRH, receptor loss. Pituitary VP receptor down-regulation and decreased hypothalamic CRH expression are likely to contribute to the reduced corticotroph responsiveness to VP reported in this experimental model.

Regulation of corticotropin-releasing hormone receptor messenger ribonucleic acid in the rat brain and pituitary by glucocorticoids and stress.

Glucocorticoids and stress are known to influence the synthesis of corticotropin-releasing hormone (CRH) at a variety of sites in brain, including the hypothalamus and amygdala. The recent cloning of the CRH receptor (CRH-R) enabled us to determine whether glucocorticoids or stress influenced CRH action via regulation of CRH-R. We, therefore, used in situ hybridization to measure CRH-R messenger RNA (mRNA) levels in the hypothalamic paraventricular nucleus (PVN), anterior pituitary (AP), amygdala, and bed nucleus of the stria terminalis (BNST) under several conditions. Systemic corticosterone (CORT) treatment, both daily injection (5 mg/rat.day) up to 14 days and pellet implant (200 mg) for 14 days, decreased CRH-R mRNA in the PVN and lateral and basolateral nucleus of the amygdala (BLA). Corticosterone injection (10 mg/rat.day, for 7 days) decreased CRH-R mRNA in the AP. Adrenalectomy also decreased CRH-R mRNA in the PVN and AP, but did not alter it in the BLA. In both sham and adrenalectomized rats with CORT pellet replacement (39 mg; ADX+CORT rats), acute (2-h) and repeated (2 h daily for 14 days) immobilization stress (which produced a large increase in plasma CORT in sham rats) increased CRH-R mRNA in the PVN and decreased it in the AP, but did not affect CRH-R mRNA in the BLA. However, ADX+CORT rats consistently had higher levels of CRH-R mRNA in both the PVN and AP than sham rats after stress. Brain stem hemisection, which damaged all ascending catecholaminergic fibers with the exception of the locus ceruleus, attenuated immobilization stress-induced up-regulation of CRH-R mRNA ipsilaterally in the PVN. None of the treatments affected CRH-R mRNA levels in the central and medial nucleus of the amygdala or the BNST. These results suggest that high concentrations of CORT or CRH synergistically decrease CRH-R mRNA levels in the AP, and that at least high CORT has an inhibitory effect on PVN CRH-R mRNA levels. However, stress input can override such inhibitory effects and thus up-regulate CRH-R mRNA in the PVN. The extrahypothalamic regions, such as amygdala and BNST may have different sensitivities to CORT or CRH for the regulation of CRH-R mRNA.

Regulation of corticotropin-releasing hormone receptor messenger ribonucleic acid in the rat brain and pituitary by glucocorticoids and stress

Glucocorticoids and stress are known to influence the synthesis of corticotropin-releasing hormone (CRH) at a variety of sites in brain, including the hypothalamus and amygdala. The recent cloning of the CRH receptor (CRH-R) enabled us to determine whether glucocorticoids or stress influenced CRH action via regulation of CRH-R. We, therefore, used in situ hybridization to measure CRH-R messenger RNA (mRNA) levels in the hypothalamic paraventricular nucleus (PVN), anterior pituitary (AP), amygdala, and bed nucleus of the stria terminalis (BNST) under several conditions. Systemic corticosterone (CORT) treatment, both daily injection (5 mg/rat.day) up to 14 days and pellet implant (200 mg) for 14 days, decreased CRH-R mRNA in the PVN and lateral and basolateral nucleus of the amygdala (BLA). Corticosterone injection (10 mg/rat.day, for 7 days) decreased CRH-R mRNA in the AP. Adrenalectomy also decreased CRH-R mRNA in the PVN and AP, but did not alter it in the BLA. In both sham and adrenalectomized rats with CORT pellet replacement (39 mg; ADX+CORT rats), acute (2-h) and repeated (2 h daily for 14 days) immobilization stress (which produced a large increase in plasma CORT in sham rats) increased CRH-R mRNA in the PVN and decreased it in the AP, but did not affect CRH-R mRNA in the BLA. However, ADX+CORT rats consistently had higher levels of CRH-R mRNA in both the PVN and AP than sham rats after stress. Brain stem hemisection, which damaged all ascending catecholaminergic fibers with the exception of the locus ceruleus, attenuated immobilization stress-induced up-regulation of CRH-R mRNA ipsilaterally in the PVN. None of the treatments affected CRH-R mRNA levels in the central and medial nucleus of the amygdala or the BNST. These results suggest that high concentrations of CORT or CRH synergistically decrease CRH-R mRNA levels in the AP, and that at least high CORT has an inhibitory effect on PVN CRH-R mRNA levels. However, stress input can override such inhibitory effects and thus up-regulate CRH-R mRNA in the PVN. The extrahypothalamic regions, such as amygdala and BNST may have different sensitivities to CORT or CRH for the regulation of CRH-R mRNA.

Corticotropin-releasing hormone and its receptor distribution in fetal membranes and placenta of the rhesus monkey in late gestation and labor

Maternal plasma corticotropin-Releasing Hormone (CRH) rises from midgestation to term and increases further during labor in pregnant women. The primate placenta contains both the CRH peptide and its gene and is the likely source of circulating CRH. In the present study, we examined CRH messenger RNA (mRNA) and peptide expression in fetal membranes and the placenta of 14 pregnant rhesus monkeys (140-161 days gestational age) using Northern blot analysis, in situ hybridization, and immunocytochemistry to define the cellular distribution of CRH during the last third of pregnancy and in relation to onset of both spontaneous term and androstenedione-induced preterm labor. To localize the target tissues for placental CRH, CRH receptor gene expression was also studied in the fetal membranes and placenta. CRH mRNA in the placenta was of similar molecular size to hypothalamic CRH. Placental CRH mRNA increased significantly during both spontaneous term and androstenedione-induced preterm labor (P < 0.05). Placental CRH peptide detected by CRH immunostaining also increased, matching the changes of CRH mRNA. In situ hybridization and immunocytochemistry demonstrated that syncytiotrophoblast cells are the major cell type expressing CRH mRNA and producing CRH protein. CRH mRNA was not detected in either amnion or chorion. CRH receptor complementary DNA and oligo probes that successfully hybridized CRH receptor mRNA in the fetal rhesus monkey hypothalamus failed to reveal the existence of CRH receptor mRNA in amnion, chorion, and placenta by Northern blot hybridization. In conclusion, placental but not fetal membrane syncytiotrophoblasts are the source of CRH production in the pregnant rhesus monkey. The significant increase in CRH peptide and mRNA content in both spontaneous term and androstenedione-induced preterm labor indicates a role for CRH in the process of parturition. The lack of CRH receptor mRNA in either the fetal membranes or the placenta suggests that placental CRH exerts its action at sites other than these tissues.

Corticotropin-releasing hormone and its receptor distribution in fetal membranes and placenta of the rhesus monkey in late gestation and labor.

Maternal plasma corticotropin-Releasing Hormone (CRH) rises from midgestation to term and increases further during labor in pregnant women. The primate placenta contains both the CRH peptide and its gene and is the likely source of circulating CRH. In the present study, we examined CRH messenger RNA (mRNA) and peptide expression in fetal membranes and the placenta of 14 pregnant rhesus monkeys (140-161 days gestational age) using Northern blot analysis, in situ hybridization, and immunocytochemistry to define the cellular distribution of CRH during the last third of pregnancy and in relation to onset of both spontaneous term and androstenedione-induced preterm labor. To localize the target tissues for placental CRH, CRH receptor gene expression was also studied in the fetal membranes and placenta. CRH mRNA in the placenta was of similar molecular size to hypothalamic CRH. Placental CRH mRNA increased significantly during both spontaneous term and androstenedione-induced preterm labor (P < 0.05). Placental CRH peptide detected by CRH immunostaining also increased, matching the changes of CRH mRNA. In situ hybridization and immunocytochemistry demonstrated that syncytiotrophoblast cells are the major cell type expressing CRH mRNA and producing CRH protein. CRH mRNA was not detected in either amnion or chorion. CRH receptor complementary DNA and oligo probes that successfully hybridized CRH receptor mRNA in the fetal rhesus monkey hypothalamus failed to reveal the existence of CRH receptor mRNA in amnion, chorion, and placenta by Northern blot hybridization. In conclusion, placental but not fetal membrane syncytiotrophoblasts are the source of CRH production in the pregnant rhesus monkey. The significant increase in CRH peptide and mRNA content in both spontaneous term and androstenedione-induced preterm labor indicates a role for CRH in the process of parturition. The lack of CRH receptor mRNA in either the fetal membranes or the placenta suggests that placental CRH exerts its action at sites other than these tissues.

Regulation of hypothalamic and pituitary corticotropin-releasing hormone receptor messenger ribonucleic acid by adrenalectomy and glucocorticoids.

The effects of adrenalectomy and glucocorticoids on the regulation of corticotropin-releasing hormone (CRH) receptor expression in the hypothalamic paraventricular nucleus (PVN) and pituitary were studied by in situ hybridization in the rat using a complementary RNA probe directed toward the coding region of the type 1 CRH receptor. Eighteen hours after adrenalectomy, CRH receptor messenger RNA (mRNA) expression in the PVN was significantly increased, whereas longer term adrenalectomy (4 and 6 days) had no effect. This transient effect of adrenalectomy was prevented by glucocorticoid replacement. In intact rats, 4 h after immobilization for 1 h or a single ip hypertonic saline injection, CRH receptor mRNA in the PVN markedly increased (P < 0.01), an effect that was unchanged by adrenalectomy (4 or 6 days) or dexamethasone injection (100 micrograms at -14 and 50 micrograms at -1 h) before stress. In the pituitary, CRH receptor mRNA levels decreased transiently after adrenalectomy (-62% after 18 h), returning to basal levels 4 or 6 days after adrenalectomy. The early decrease was prevented by glucocorticoid replacement. In intact rats, dexamethasone (100 micrograms, sc) caused a significant decrease in pituitary CRH receptor mRNA levels 2-10 h after injection, returning to basal levels after 15 h. On the other hand, dexamethasone (5-300 micrograms, sc) had no effect on pituitary CRH receptor mRNA levels 18 h after injection. The data show that although stress stimulation of CRH mRNA in the PVN is glucocorticoid independent, basal levels are likely to be under dual, transcriptional and posttranscriptional, control by glucocorticoids. In the pituitary, changes in hypothalamic CRFs probably play a major role in the control of CRH receptor mRNA levels during manipulations of circulating glucocorticoids levels. In addition, the inability of long term adrenalectomy and glucocorticoid administration to modify pituitary CRH receptor mRNA levels suggests that CRH receptor down-regulation observed under these experimental conditions depends mainly on translational and post-translational events rather than receptor mRNA levels.

Regulation of hypothalamic and pituitary corticotropin-releasing hormone receptor messenger ribonucleic acid by adrenalectomy and glucocorticoids

The effects of adrenalectomy and glucocorticoids on the regulation of corticotropin-releasing hormone (CRH) receptor expression in the hypothalamic paraventricular nucleus (PVN) and pituitary were studied by in situ hybridization in the rat using a complementary RNA probe directed toward the coding region of the type 1 CRH receptor. Eighteen hours after adrenalectomy, CRH receptor messenger RNA (mRNA) expression in the PVN was significantly increased, whereas longer term adrenalectomy (4 and 6 days) had no effect. This transient effect of adrenalectomy was prevented by glucocorticoid replacement. In intact rats, 4 h after immobilization for 1 h or a single ip hypertonic saline injection, CRH receptor mRNA in the PVN markedly increased (P < 0.01), an effect that was unchanged by adrenalectomy (4 or 6 days) or dexamethasone injection (100 micrograms at -14 and 50 micrograms at -1 h) before stress. In the pituitary, CRH receptor mRNA levels decreased transiently after adrenalectomy (-62% after 18 h), returning to basal levels 4 or 6 days after adrenalectomy. The early decrease was prevented by glucocorticoid replacement. In intact rats, dexamethasone (100 micrograms, sc) caused a significant decrease in pituitary CRH receptor mRNA levels 2-10 h after injection, returning to basal levels after 15 h. On the other hand, dexamethasone (5-300 micrograms, sc) had no effect on pituitary CRH receptor mRNA levels 18 h after injection. The data show that although stress stimulation of CRH mRNA in the PVN is glucocorticoid independent, basal levels are likely to be under dual, transcriptional and posttranscriptional, control by glucocorticoids. In the pituitary, changes in hypothalamic CRFs probably play a major role in the control of CRH receptor mRNA levels during manipulations of circulating glucocorticoids levels. In addition, the inability of long term adrenalectomy and glucocorticoid administration to modify pituitary CRH receptor mRNA levels suggests that CRH receptor down-regulation observed under these experimental conditions depends mainly on translational and post-translational events rather than receptor mRNA levels.

Stress-specific regulation of corticotropin releasing hormone receptor expression in the paraventricular and supraoptic nuclei of the hypothalamus in the rat.

Corticotropin releasing hormone (CRH), a major regulator of pituitary ACTH secretion, also acts as a neurotransmitter in the brain. To determine whether CRH is involved in the regulation of hypothalamic function during stress, CRH receptor binding and CRH receptor mRNA levels were studied in the hypothalamus of rats subjected to different stress paradigms: immobilization, a physical-psychological model; water deprivation and 2% saline intake, osmotic models; and i.p. hypertonic saline injection, a combined physical-psychological and osmotic model. In agreement with the distribution of CRH receptor binding in the brain, in situ hybridization studies using 35S-labeled cRNA probes revealed low levels of CRH receptor mRNA in the anterior hypothalamic area, which were unaffected after acute or chronic exposure to any of the stress paradigms used. Under basal conditions, there was no CRH binding or CRH receptor mRNA in the supraoptic (SON) or paraventricular (PVN) nuclei. However, 2 h after the initiation of acute immobilization, CRH receptor mRNA hybridization became evident in the parvicellular division of the PVN, with levels substantially increasing from 2 to 4 h, decreasing at 8 h and disappearing by 24 h. Identical hybridization patterns of CRH receptor mRNA were found in the parvicellular PVN after repeated immobilization; levels were similar to those after 2 h single stress following immobilization at 8-hourly intervals for 24 h (3 times), and very low, but clearly detectable 24 h after 8 or 14 days daily immobilization for 2 h. On the other hand, water deprivation for 24 or 60 h and intake of 2% NaCl for 12 days induced expression of CRH receptor mRNA in the SON and magnocellular PVN, but not in the parvicellular pars of the PVN.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of corticotropin-releasing hormone (CRH) receptor mRNA in adult rat brain by in situ hybridization histochemistry

Even though functional CRH receptors have been identified in several brain regions by ligand binding, the identity of brain areas expressing the CRH receptor gene has not been described. The recent cloning of the rat CRH receptor gene has permitted us to conduct an in situ hybridization histochemistry study to localize CRH receptor mRNA in brain, using an antisense 35S-labeled riboprobe and autoradiography. In virus- and pathogen-free, unstressed, adult male Sprague-Dawley rats we observed CRH receptor gene expression in several brain regions, most of which had been previously shown to bind radiolabeled CRH. Those regions include the pituitary, olfactory bulb, hippocampal formation, cerebral and cerebellar cortexes, hypothalamus, median eminence, amygdala, olfactory tubercle, choroid plexus, thalamus, and inferior colliculus. Further studies are needed to determine the cell types expressing both CRH receptor mRNA and the CRH receptor peptide in nervous system as well as in peripheral tissues.

Localization of corticotropin-releasing hormone (CRH) receptor mRNA in adult rat brain by in situ hybridization histochemistry.

Even though functional CRH receptors have been identified in several brain regions by ligand binding, the identity of brain areas expressing the CRH receptor gene has not been described. The recent cloning of the rat CRH receptor gene has permitted us to conduct an in situ hybridization histochemistry study to localize CRH receptor mRNA in brain, using an antisense 35S-labeled riboprobe and autoradiography. In virus- and pathogen-free, unstressed, adult male Sprague-Dawley rats we observed CRH receptor gene expression in several brain regions, most of which had been previously shown to bind radiolabeled CRH. Those regions include the pituitary, olfactory bulb, hippocampal formation, cerebral and cerebellar cortexes, hypothalamus, median eminence, amygdala, olfactory tubercle, choroid plexus, thalamus, and inferior colliculus. Further studies are needed to determine the cell types expressing both CRH receptor mRNA and the CRH receptor peptide in nervous system as well as in peripheral tissues.