Corticotropin-releasing Factor Immunoreactive Neurons Research Articles

Corticotropin-releasing factor potentiates glutamatergic input and excitability of presympathetic neurons in the hypothalamus in spontaneously hypertensive rats

Hyperactivity of presympathetic neurons in the hypothalamic paraventricular nucleus (PVN) plays a key role in generating excess sympathetic output in hypertension. However, the mechanisms driving hyperactivity of PVN presympathetic neurons in hypertension are unclear. In this study, we determined the role of corticotropin-releasing factor (CRF) in the PVN in augmented glutamatergic input, neuronal excitability and sympathetic outflow in hypertension. The number of CRF or c-Fos immunoreactive neurons and CRF/c-Fos double-labeled neurons in the PVN was significantly greater in spontaneously hypertensive rats (SHRs) than in normotensive Wistar-Kyoto (WKY) rats. Blocking glutamatergic input reduced the CRF-potentiated excitability of spinally projecting PVN neurons. Furthermore, CRF knockdown via Crispr/Cas9 in the PVN decreased the frequencies of spontaneous firing and miniature excitatory postsynaptic currents (mEPSCs) in spinally projecting PVN neurons in SHRs. In addition, the mRNA and protein levels of CRFR1, but not CRFR2, in the PVN were significantly higher in SHRs than in WKY rats. Blocking CRFR1 with NBI-35965, but not blocking CRFR2 with Antisauvagine-30, reduced the frequencies of spontaneous firing and mEPSCs of spinally projecting PVN neurons in SHRs. Also, microinjection of NBI-35965 into the PVN significantly reduced arterial blood pressure (ABP) and renal sympathetic nerve activity (RSNA) in anesthetized SHRs, but not in WKY rats. However, microinjection of Antisauvagine-30 into the PVN had no effect on ABP or RSNA in WKY rats and SHRs. Our findings suggest that endogenous CRF in the PVN potentiates glutamatergic input and firing activity of PVN presympathetic neurons via CRFR1, resulting in augmented sympathetic outflow in hypertension.

Effects of neonatal rearing by different types of foster mother on the distribution of corticotropin-releasing factor neurons in the central amygdaloid nucleus in rats.

The mother-child relationship of newborns plays an essential role in the development of the central nervous system, and an inadequate relationship, such as mother-child separation, can cause deficits of mental function in adulthood. However, insufficient research has examined the effects of foster mothers. We assigned some neonatal rats to one of two foster mothers: one that was lactating and feeding her first litter (FL group) and one that had one previous experience of childbirth and feeding but no current litter (FE group). Other pups were raised by their own mother (OM group) or subjected to maternal separation (MS group). Pups were placed with the foster mother (FL and FE groups) or separated from their mother (MS group) for 3h/day on postnatal days 1-20. At age 6weeks, each group was divided into two subgroups, one with 30min of acute restraint stress loading (FL-R, FE-R, OM-R, and MS-R) and one without it (FL, FE, OM, and MS). Then, we compared the density of corticotropin-releasing factor-immunoreactive (CRF-ir) neurons in the central amygdaloid nucleus (CeA). The density of CRF-ir neurons in the CeA was significantly lower in the FL-R and MS-R subgroups than in the FL and MS subgroups, respectively. The results suggest that differences in care received during the neonatal period affect maturation of CRF neurons in the CeA and may have negative effects on the synthesis and release of CRF.

Activity-based anorexia activates CRF immunoreactive neurons in female rats

Activity-based anorexia (ABA) is a well-established animal model mimicking the eating disorder anorexia nervosa (AN). Since the pathophysiology of AN is yet poorly understood and specific drug treatments are lacking so far, animal models might be useful to further understand this disease. ABA consists of time-restricted access to food for 1.5 h/day and the possibility to exercise in a running wheel for 24 h/day. This combination leads to robust body weight loss as observed in AN. Here, we investigated the activation of brain corticotropin-releasing factor (CRF) neurons, a transmitter involved in the response to stress, emotional processes and also food intake. After development of ABA, rat brains were processed for c-Fos and CRF double immunohistochemistry. ABA increased the number of c-Fos/CRF double labeled neurons in the paraventricular nucleus (PVN) and the dorsomedial hypothalamic nucleus (DMH) compared to the ad libitum (AL, ad libitum fed, no running wheel) and activity (AC, ad libitum fed, running wheel, p < 0.05) but not to the restricted feeding (RF, food for 1.5 h/day, no running wheel, p > 0.05) group. Also the number of CRF neurons was increased in the DMH of ABA rats compared to AL and AC (p < 0.05). In the Edinger-Westphal nucleus (EW) the number of c-Fos positive neurons was increased in ABA and RF compared to AC (p < 0.05), while the number of double labeled neurons was not different (p > 0.05). Taken together, brain CRF activated under conditions of ABA might play a role in the development and maintenance of this animal model and possibly also in human AN.

Delta opioid receptors colocalize with corticotropin releasing factor in hippocampal interneurons

The hippocampal formation (HF) is an important site at which stress circuits and endogenous opioid systems intersect, likely playing a critical role in the interaction between stress and drug addiction. Prior study findings suggest that the stress-related neuropeptide corticotropin releasing factor (CRF) and the delta opioid receptor (DOR) may localize to similar neuronal populations within HF lamina. Here, hippocampal sections of male and cycling female adult Sprague–Dawley rats were processed for immunolabeling using antisera directed against the DOR and CRF peptide, as well as interneuron subtype markers somatostatin or parvalbumin, and analyzed by fluorescence and electron microscopy. Both DOR- and CRF-labeling was observed in interneurons in the CA1, CA3, and dentate hilus. Males and normal cycling females displayed a similar number of CRF immunoreactive neurons co-labeled with DOR and a similar average number of CRF-labeled neurons in the dentate hilus and stratum oriens of CA1 and CA3. In addition, 70% of DOR/CRF dual-labeled neurons in the hilar region co-labeled with somatostatin, suggesting a role for these interneurons in regulating perforant path input to dentate granule cells. Ultrastructural analysis of CRF-labeled axon terminals within the hilar region revealed that proestrus females have a similar number of CRF-labeled axon terminals that contain DORs compared to males but an increased number of CRF-labeled axon terminals without DORs. Taken together, these findings suggest that while DORs are anatomically positioned to modulate CRF immunoreactive interneuron activity and CRF peptide release, their ability to exert such regulatory activity may be compromised in females when estrogen levels are high.

Morphology, size and distribution of corticotropin releasing factor (CRF) immunoreactive neurons in the central nucleus of the rat amygdaloid complex

The amygdaloid complex (AC) is a heterogenous group of cortical and nuclear structures. As an important component of the limbic system contains numerous neurotransmitters including neuropeptides. In this study we have investigated the types, distribution, and morphometric characteristics of corticotropin releasing factor (CRF) immunoreactive neurons in the central nucleus (Ce) of the AC. We have also identified CRF- immunoreactive fibers in this nucleus. This research was performed on 5 adult rats kept in standard conditions. Animals were perfused 48 hours after application of colhicin. The removed brains were postfixed and cut into free floating sections. The sections were treated with antibodies against the rabbit CRF and ABC immunohistochemstry was applied. The neurons were measured and drawn with a camera lucida. We found uniform distribution of CRF immunoreactive neurons in the Ce of AC of rat. According to morphological type dominant were triangular (50%), while bipolar (25%) and multipolar (25%) types were also present. The most of the CRF immunoreactive neurons had 2-5 primary dendrites. These findings will contribute to understanding the role of CRF neurons in rat Ce during stress and in other behavioral functions.

Distribution and chemical coding of corticotropin-releasing factor-immunoreactive neurons in the guinea pig enteric nervous system

Immunofluorescence was used to study immunoreactivity (IR) for corticotropin-releasing factor (CRF) in the guinea pig enteric nervous system. CRF-IR was expressed in both the myenteric and the submucosal plexuses of all regions of the large and small intestine and the myenteric plexus of the stomach. CRF-IR nerve fibers were present in the myenteric and submucosal plexuses, in the circular muscle coat, and surrounding submucosal arterioles. Most of the CRF-IR fibers persisted in the myenteric and submucosal plexuses after 7 days in organotypic culture. CRF-IR was not coexpressed with tyrosine hydroxylase-IR or calcitonin gene-related peptide-IR fibers. The proportions of CRF-IR cell bodies in the myenteric plexus increased progressively from the stomach (0.6%) to the distal colon (2.8%). Most of the CRF-IR myenteric neurons (95%) had uniaxonal morphology; the remainder had Dogiel type II multipolar morphology. CRF-IR cell bodies in the myenteric plexus of the ileum expressed IR for choline acetyltransferase (56.9%), substance P (55.0%), and nitric oxide synthase (37.9%). CRF-IR never colocalized with IR for calbindin, calretinin, neuropeptide Y, serotonin, or somatostatin in the myenteric plexus. CRF-IR cell bodies were more abundant in the submucosal plexus (29.9-38.0%) than in the myenteric plexus. All CRF-IR neurons in submucosal ganglia expressed vasoactive intestinal peptide-IR and were likely to be secretomotor/vasodilator neurons. CRF-IR neurons did not express IR for the CRF(1) receptor. CRF(1)-IR was expressed in neuronal neighbors of those with CRF-IR. Collective evidence suggests that VIPergic secretomotor neurons might provide synaptic input to neighboring cholinergic neurons.

Interrelations between monoaminergic afferents and corticotropin-releasing factor-immunoreactive neurons in the rat central amygdaloid nucleus: ultrastructural evidence for dopaminergic control of amygdaloid stress systems.

Ample evidence implicates corticotropin-releasing factor (CRF)-producing neurons of the central amygdaloid nucleus (CeA) in vegetative, endocrine, and behavioral responses to stress and anxiety in laboratory rats. Monoaminergic systems are involved in modulating these responses. In the present paper, interrelations between CRF-immunoreactive (ir) neurons, and noradrenergic, serotonergic, and dopaminergic afferents were studied using single and double immunolabeling for light and electron microscopy in the rat CeA. Dopaminergic axons formed dense plexus in the CeA overlapping with the localization of CRF-ir neurons, and their terminals formed frequent associations with CRF-ir somata. Contacts of serotonergic axons on CRF-ir neurons were few, and contacts of noradrenergic axons were the exception. Ultrastructurally, symmetric synapses of dopaminergic terminals on CRF-ir somata and dendrites were found. More than 83% of CRF-ir somata were contacted in single ultrathin sections. About half of these possessed two or more contacts. Of non-ir somata, 37% were contacted by dopaminergic terminals, and only 13% of these had two or more contacts. Correlative in situ hybridization indicated that CeA CRF-ir neurons may express receptor subtype dopamine receptor subtype 2. In conclusion, dopaminergic afferents appear to specifically target CeA CRF neurons. They are thus in a position to exert significant influence on the rat amygdaloid CRF stress system.

Effect of 6-hydroxydopamine on neuropeptide Y and corticotropin-releasing factor expression in rat amygdala

The influence of dopaminergic denervation on neuropeptide Y and corticotropin-releasing factor-containing neurons in the amygdala was investigated in rats by examining the effects of a selective, unilateral 6-hydroxydopamine lesion of mesencephalic dopaminergic neurons in both the substantia nigra and the ventral tegmental area on these peptides and their messenger RNA expression, observed eight to 10 days after the lesion. The studies were conducted by immunocytochemical and in situ hybridization methods. Neuropeptide Y or corticotropin-releasing factor-immunoreactive neurons were counted in sections of the amygdala under a microscope, and the messenger RNA expression was measured as optical density units in autoradiograms. A significant increase in both neuropeptide Y and corticotropin-releasing factor messenger RNA expression was found in the amygdala on the lesioned side in comparison with the contralateral one, as well as with the ipsilateral side of vehicle-injected controls. Immunohistochemical studies showed that the number of neuropeptide Y-immunoreactive neurons increased in the whole amygdala on the lesioned side. At the same time, the number of corticotropin-releasing factor-immunoreactive neurons grouped in the central amygdaloid nucleus declined, and so did the staining intensity. The obtained results indicate that dopaminergic denervation stimulates the synthesis of neuropeptide Y and corticotropin-releasing factor in rat amygdala, but the peptide levels are differently regulated, which points to a diverse release of these peptides.

The amygdala: corticotropin-releasing factor, steroids, and stress.

The possible function of corticotropin-releasing factor (CRF), adrenal steroids, and gonadal steroids in amygdala-mediated responses to anxiogenic or stressful stimuli is reviewed. The amygdala is part of an endogenous CRF circuitry within the brain that mediates neuroendocrine, autonomic, and behavioral changes in response to stress. The amygdala contains CRF-expressing neurons that communicate with widespread regions of the neural axis. High densities of CRF, CRF-binding protein, and CRF receptors are located in the amygdala. Direct injections of CRF into the amygdala produce anxiety-like behaviors. Release of endogenous CRF can be measured in the amygdala during stress. Potent anxiolytic actions are observed when CRF receptor antagonists are administered into the amygdala. CRF-containing neurons of the amygdala can be directly modulated by alterations in circulating glucocorticoids through glucocorticoid receptors, which are expressed in amygdaloid CRF-containing neurons. Gonadal steroid hormone receptors are found in the amygdala. They are not located in CRF immunoreactive neurons, but they are located adjacent to CRF-expressing neurons and in amygdaloid neurons that are likely to participate in central autonomic and neuroendocrine circuitry. Differences are noted between the steroid influences in the amygdala of male and female animals. Also, evidence is reviewed suggesting a modulatory role in the amygdala for gonadal and adrenal steroids in behavioral, autonomic, and neuroendocrine responses to anxiogenic stimuli.

Alteration of corticotropin‐releasing factor immunoreactivity in MPTP‐treated rats

A decrease of corticotropin-releasing factor (CRF) concentration has been reported in patients with Parkinson's disease (PD). The present study further examined the role of CRF in an animal model of parkinsonism induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Results indicated that both subchronic (2 days) and chronic (7 days) MPTP treatments decreased the number of CRF immunoreactive neurons in both the paraventricular nucleus (PVN) of the hypothalamus and the central nuelcus of the amygdala (ACN). This effect lasted for almost a month after withdrawal of chronic MPTP injections. In addition, nomifensine pretreatment protected against MPTP's toxicity on DA neurons, as assessed by tyrosine hydroxylase immunoreactivity in the substantia nigra. However, the same treatment did not prevent the toxicity of MPTP on CRF neurons. Further, no significant difference was notable in the number of CRF immunoreactive neurons between normal young adult and normal middle-aged rats in both the PVN and the ACN. These results suggest that MPTP also produces a neurotoxicity on CRF neurons, and this effect is not secondary to MPTP's effect on DA neurons. Besides, altered CRF neuronal activity is involved in the process of pathological ageing, but not physiological ageing. Further, reduced CRF immunoreactivity in the PVN and ACN may imply alterations of neuroendocrine, autonomic as well as central functions caused by MPTP.

Distribution of corticotropin-releasing factor immunoreactive neurons in the brain of the tigerfrog, Rana tigrina

The distribution of immunoreactive (ir) neurons containing corticotropin-releasing factor (CRF) is described in the brain of the tigerfrog, Rana tigrina. The olfactory bulb, medial pallium, nucleus of the diagonal band of Broca and medial area of the amygdala of the telencephalon showed ir-CRF perikarya. The anterior and ventromedial thalamic nuclei, and the magnocellular nucleus preopticus (NPO) revealed several ir cells; a few NPO neurons were cerebrospinal fluid contacting in nature. The nucleus hypothalamicus ventromedialis contained a few cells, but the nucleus infundibularis ventralis of the infundibulum revealed several diffusely distributed perikarya. Individual ir-CRF perikarya were visualized in the optic tectum and interpeduncular nucleus. Extensive fiber terminals were present in the median eminence, but no fibers were discerned either in the neural lobe or in the pituitary gland.

Quantitative Analysis of Tuberoinfundibular Tyrosine Hydroxylase- and Corticotropin-Releasing Factor-Immunoreactive Neurons in Monkeys Raised with Differential Rearing Conditions

A major goal in assessing biological determinants of behavior lies in studying the effect(s) of rearing on the development of the central nervous system. Specifically, a series of neuroanatomic analyses have been undertaken to identify potential neuropathological changes seen in monkeys exposed to early social deprivation, which leads to profound psychopathology and inappropriate responses to stress. The animals used in this study were either raised with their mother and peers (socially reared) or raised without maternal/peer contact (socially deprived). Within this context, the distribution of tuberoinfundibular dopaminergic neurons in the hypothalamic paraventricular nucleus and arcuate nucleus of rhesus monkeys was determined by immunohistochemistry using an antibody against the enzyme tyrosine hydroxylase, a marker for dopamine-containing systems. Additionally, the distribution of corticotropin-releasing factor-containing neurons in the paraventricular nucleus was assessed immunohistochemically. The majority (97.5%) of dopaminergic neurons in the paraventricular nucleus were parvicellular, with a small (2.5%), but consistently observed population of magnocellular neurons immunoreactive for tyrosine hydroxylase. Within the arcuate nucleus, tyrosine hydroxylase-immunoreactive neurons were similar in morphology to the parvicellular neurons of the paraventricular nucleus. Qualitative assessment of corticotropin-releasing factor-immunoreactive neurons in the paraventricular nucleus revealed a parvicellular population of neurons located in medial aspects of the nucleus, similar to what has been observed in the rat. Quantitative analysis revealed no differences in the number of tyrosine hydroxylase- and corticotropin-releasing factor-immunoreactive neurons between rearing conditions, suggesting that these neurons were not affected, in terms of overall cell counts, by the early environmental insult of social deprivation.

Coexistence of c-Fos and glucocorticoid receptor immunoreactivities in the CRF immunoreactive neurons of the paraventricular hypothalamic nucleus of the rat after acute immobilization stress

By means of triple immunofluorescence procedures the codistribution and coexistence of glucocorticoid receptor (GR), c-Fos and corticotropin-releasing factor (CRF) immunoreactive (IR) nerve cell profiles have been analyzed within the paraventricular hypothalamic nucleus at various rostrocaudal levels following acute immobilization stress (2 h) in the male rat. One population of CRF IR neurons is shown to contain both nuclear GR and c-Fos immunoreactivities following the stress, while another contains GR immunoreactivity alone. These results indicate heterogeneities among parvocellular CRF paraventricular hypothalamic nerve cell populations in the response to acute immobilization stress.

Distribution and cerebellar projections of cholinergic and corticotropin-releasing factor-containing neurons in the caudal vestibular nuclear complex and adjacent brainstem structures

By using immunohistochemistry combined with lesioning and retrograde neuronal labeling techniques, cholinergic neurons and corticotropin-releasing factor-immunoreactive neurons were examined for their distribution, coincidence and cerebellar projections in feline vestibular nuclear complex and adjacent brainstem structures. Cholinergic neurons as revealed here with choline acetyltransferase immunoreactivity were found massively in the abducens and hypoglossal nuclei, dorsal motor nucleus of the vagus nerve and nucleus of Roller; less numerously in the medial vestibular, prepositus hypoglossi and solitary nuclei and the caudal two-thirds of descending vestibular nucleus; and only occasionally in the intercalated and supravestibular nuclei and cell groups f, x and z. Corticotropin-releasing factor-immunoreactive neurons were found clustered in the prepositus hypoglossi nucleus and also in cell groups f and x and the rostral two-thirds of descending vestibular nucleus, less numerously in the medial vestibular, intercalated and solitary nuclei and nucleus of Roller, and only occasionally in the caudal one-third of descending vestibular nucleus, the dorsal motor nucleus of the vagus nerve, supravestibular nucleus and cell group z. The lateral and superior vestibular nuclei did not contain either type of neuron. The two types of immunopositive neurons observed in most of the brainstem nuclei differed in cell size, distribution-pattern and rostrocaudal level of occurrence. While there were many regions which exhibited both types of immunopositive neurons, perikarya colocalizing the cholinergic and peptide markers were not detected in the brainstem. Following unilateral, partial lesioning of the vestibular nuclear complex, corticotropin-releasing factor-immunoreactive mossy fiber terminals (rosettes) disappeared from the ipsilateral flocculus. However, such lesions did not produce clear-cut changes of cholinergic terminals in the vermis. Following retrograde neuronal labeling combined with immunohistochemistry, the two types of immunopositive neurons observed in most of the brainstem sites were found to project to the vermal lobules I–III, IX and X. On comparison of these immunopositive projection neurons with non-immunoreactive, retrogradely labeled neurons, the cholinergic neurons and the peptide-immunoreactive neurons were found to constitute a major part of the total vestibulocerebellar neuronal population. The results indicate chemical heterogeneity in vestibular nuclear complex and cerebellar afferents.

Repeated stress enhances vasopressin synthesis in corticotropin releasing factor neurons in the paraventricular nucleus

The effect of repeated stress on corticotropin-releasing factor (CRF) and vasopressin (AVP) synthesis in parvocellular neurons of the paraventricular nucleus (PVN) was studied by means of double label immunocytochemistry. Once daily immobilization of male rats for 16 days leads to a 75% increase in the number of CRF immunoreactive neurons, but a 5-fold elevation in the number of AVP containing CRF cell bodies. These results demonstrate that repeated stress activates AVP synthesis in CRF neurons.

The distribution of corticotropin-releasing factor-immunoreactive neurons and nerve fibers in the brain of the snake, Natrix maura

The anatomical distribution of neurons and nerve fibers containing corticotropin-releasing factor (CRF) has been studied in the brain of the snake, Natrix maura, by means of immunocytochemistry using an antiserum against rat CRF. To test the possible coexistence of CRF with the neurohypophysial peptides arginine vasotocin (AVT) and mesotocin (MST) adjacent sections were stained with antisera against the two latter peptides. CRF-immunoreactive (CRF-IR) neurons exist in the paraventricular nucleus (PVN). In some neurons of the PVN, coexistence of CRF with MST or of CRF with AVT has been shown. Numerous CRF-IR fibers run along the hypothalamo-hypophysial tract and end in the outer layer of the median eminence. In addition, some fibers reach the neural lobe of the hypophysis. CRF-IR perikarya have also been identified in the following locations: dorsal cortex, nucleus accumbens, amygdala, subfornical organ, lamina terminalis, nucleus of the paraventricular organ, nucleus of the oculomotor nerve, nucleus of the trigeminal nerve, and reticular formation. In addition to all these locations CRF-IR fibers were also observed in the lateral septum, supraoptic nucleus, habenula, lateral forebrain bundle, paraventricular organ, hypothalamic ventromedial nucleus, raphe and interpeduncular nuclei.

Coexistence of corticotropin‐releasing factor and enkephalin in the paraventricular nucleus of the rat

Corticotropin-releasing factor and enkephalin-containing neurons are found in the paraventricular nucleus of the rat hypothalamus. Their immunocytochemical distribution suggests that a subpopulation of neurons of the paraventricular nucleus might contain both peptides. The present study describes the coexistence of corticotropin-releasing factor and enkephalin in parvicellular neurons of the paraventricular nucleus. Immunocytochemical labeling of peptides was combined with in situ hybridization of mRNA to visualize peptide and mRNA labeling in the same tissue section. This resulted in several observations: (1) neurons labeling for the peptide corticotropin-releasing factor also contain the mRNA to synthesize corticotropin-releasing factor, (2) neurons labeling for the peptide enkephalin also contain the mRNA to synthesize the peptide enkephalin, (3) a subpopulation of corticotropin-releasing factor neurons contains the mRNA to synthesize enkephalin, and (4) a subpopulation of enkephalin neurons contains the mRNA to synthesize corticotropin-releasing factor. A large percentage of enkephalin immunoreactive neurons contains corticotropin-releasing factor mRNA, whereas a smaller percentage of corticotropin-releasing factor immunoreactive neurons contains enkephalin mRNA. These double-labeled neurons are present throughout the rostral-caudal extent of the paraventricular nucleus; the majority of these neurons is present in the medial parvicellular paraventricular nucleus.

Corticotropin-releasing factor immunoreactive neurons persist throughout the brain in Alzheimer's disease

Corticotropin-releasing factor (CRF) immunoreactivity was examined in the hippocampal formation, cerebellum and hypothalamus of normal aged and Alzheimer's disease (AD) brains. Immunoreactive non-pyramidal neurons were located in the polymorphic layer of the dentate gyrus and CA fields. A plexus of CRF terminals was seen in the supragranular layer of the dentate gyrus. In AD, occasional senile plaques contained CRF-immunoreactive fiber terminals but the pattern of staining was otherwise unchanged aside from a suggestion of increased staining intensity. Similarly, the pattern of immunoreactive cerebellar climbing fibers and paraventricular hypothalamic neurons was preserved in AD brains aside from increased perikaryal staining intensity in the hypothalamus.

Demonstration of glucocorticoid receptor-like immunoreactivity in glucocorticoid-sensitive vasopressin and corticotropin-releasing factor neurons in the hypothalamic paraventricular nucleus.

Many parvocellular neurons in the paraventricular nucleus of the hypothalamus express high levels of corticotropin releasing factor (CRF) or vasopressin following adrenalectomy. To determine whether glucocorticoids feed back directly on these neurons, a mouse monoclonal antibody directed against the rat liver glucocorticoid receptor was used in combination with polyclonal antisera directed against either vasopressin or CRF to permit simultaneous visualization of either peptide with glucocorticoid receptor-like immunoreactivity (IR). Rats were adrenalectomized (ADX) for 2 weeks to optimize numbers of vasopressin - and CRF-IR neurons. Six hours prior to sacrifice, a separate group of adrenalectomized rats was treated with corticosterone (40 mg/kg). This short-term replacement resulted in nuclear localization of glucocorticoid receptor-like-IR but did not attenuate the increased numbers of CRF- and vasopressin-IR neurons observed after adrenalectomy. It was therefore possible to visualize vasopressin- or CFR-IR and nuclear glucocorticoid receptor-like-IR simultaneously. Cell counts of double-labeled neurons in the paraventricular nucleus of the hypothalamus (PVH) demonstrated that glucocorticoid receptor-like-IR is colocalized in virtually all the CRF and vasopressin immunoreactive parvocellular neurons studied, which respond to adrenalectomy by increased peptide expression. These data suggest that a major feedback effect of glucocorticoids on the hypothalamic-pituitary-adrenal axis is exerted directly within nuclei of CRF and vasopressin neurons.

Organization of ovine corticotropin-releasing factor immunoreactive neurons in the canine hypothalamo-pituitary system

The distribution of corticotropin-releasing factor immunoreactive (CRF-i) cell bodies and varicose fibers in the hypothalamus and the pituitary of the canine brain was studied by indirect immunofluorescence. CRF-i cell bodies were demonstrated mainly in the periventricular zone of the third ventriele, while some CRF-i cell bodies were scattered throughout the ventral part of the caudomedial hypothalamus. CRF-i fibers were mostly situated in the median eminence. In addition some CRF-positive fibers were detected in the ventromedial aspect of the lateral hypothalamus forming a pathway arising from the CRF-i cell bodies, running via the median eminence through the infundibular stalk and terminating in the pars nervosa of the pituitary. The localization of the cell bodies and their projection points towards a possible (patho)physiological role of this peptide-transmitter system in the release of ACTH and β-endorphin.