GABAA receptor epsilon subunit expression in identified peptidergic neurons of the rat hypothalamus.

Dual-labeling immunohistochemical or in situ hybridization studies for the recently cloned epsilon-subunit and several neuropeptides were performed in the rat hypothalamus. We revealed an extensive co-expression (>90%) with hypocretin (Hcrt), oxytocin (OT), the gonadotropin-releasing hormone (GnRH), and the melanin-concentrating hormone (MCH) peptides, whereas occasional co-expression (<10%) with cocaine-amphetamine-regulated transcript (CART) was found. Our results suggest that novel GABA(A) receptor subtypes comprising epsilon-subunit are important for metabolic and neuroendocrine functions.

Paraventricular nucleus neurons producing neurotensin after lipopolysaccharide treatment project to the median eminence.

Inflammation consists in secretion of cytokines that stimulate the hypothalamo-pituitary-adrenal (HPA) axis to release the anti-inflammatory corticosterone. Upstream in this axis are corticotropin-releasing hormone (CRH) neurons in the paraventricular nucleus (PVN) whose multipeptidergic phenotype changes: both corticotropin-releasing hormone mRNAs and neurotensin mRNAs are up-regulated. Combining in situ hybridization with a retrograde neuronal marker, we demonstrated that neurotensin-containing neurons in the paraventricular nucleus project to the median eminence.

Brain-endocrine interactions: a microvascular route in the mediobasal hypothalamus.

Blood-borne hormones acting in the mediobasal hypothalamus, like those controlling food intake, require relatively direct access to target chemosensory neurons of the arcuate nucleus (ARC). An anatomical substrate for this is a permeable microvasculature with fenestrated endothelial cells in the ARC, a system that has awaited comprehensive documentation. Here, the immunofluorescent detection of endothelial fenestral diaphragms in the rat ARC allowed us to quantitate permeable microvessels throughout its rostrocaudal extent. We have determined that permeable microvessels are part of the subependymal plexus irrigating exclusively the ventromedial (vm) ARC from the subadjacent neuroendocrine median eminence. Unexpectedly, permeable microvessels were concentrated proximal to the pituitary stalk. This marked topography strongly supports the functional importance of retrograde blood flow from the pituitary to the vmARC, therefore making a functional relationship between peripheral long-loop, pituitary short-loop, and neuroendocrine ultra-short loop feedback, altogether converging for integration in the vmARC (formerly known as the hypophysiotrophic area), thereby so pivotal as a multicompetent brain endocrinostat.

Glucocorticoids down-regulate lipopolysaccharide-induced de novo production of neurotensin mRNA in the rat hypothalamic, paraventricular, corticotrophin-releasing hormone neurons.

OBJECTIVE: Intraperitoneal injection of the endotoxin lipopolysaccharide (LPS) produces inflammation accompanied by activation of the immune system and the secretion of cytokines. Cytokines stimulate the hypothalamo-pituitary-adrenal (HPA) axis to release the anti-inflammatory corticosterone which controls its own production by acting on the HPA axis. Upstream in the HPA axis are neuroendocrine corticotrophin-releasing hormone (CRH) neurons located in the paraventricular nucleus (PVN), whose multipeptidergic phenotype changes during inflammation: while CRH mRNA is up-regulated in these conditions, neurotensin (NT) mRNA expression is induced de novo. The negative feedback control of glucocorticoids on CRH production is well documented; however, their action on NT production in the PVN of the hypothalamus is poorly documented. The aim of this study was to determine if glucocorticoids modulate the de novo production of NT during inflammation. METHODS: Using quantitative in situ hybridization histochemistry, we examined whether the absence (adrenalectomy) or excess (corticosterone implants) of glucocorticoids modulate de novo production of NT mRNA in the PVN during inflammation induced by LPS treatment. RESULTS: A relatively low dose of LPS (50 microg/kg) that is not efficient to induce NT mRNA production in the PVN becomes efficient after adrenalectomy. Moreover, corticosterone excess reduces LPS-induced production of NT mRNA in the PVN. CONCLUSION: Glucocorticoids exert a negative control on NT mRNA production in the PVN of the hypothalamus, and this effect requires that NT mRNA production be triggered, such as during inflammation.

Tilapia (Oreochromis niloticus) vitellogenins: development of homologous and heterologous ELISAs and analysis of vitellogenin pathway through the ovarian follicle.

Vitellogenin (VTG) of Oreochromis niloticus was again purified, due to the conflicting results found in the literature. Three purification processes have been used: electrophoresis and electro-elution, double chromatography (gel filtration and ion-exchange chromatography) and single ion-exchange chromatography. Using SDS-PAGE we confirmed in all cases the presence of two polypeptidic forms of plasma VTG of 130 kDa (VTG1) and 170 kDa (VTG2). We raised polyclonal antibodies against each VTG form and we demonstrated the complete cross-reactivity of each antibody with both forms of VTG by Enzyme Immuno-Assay (EIA) and Western blots. The homologous ELISAs developed exhibited a detection limit of 6 ng x ml(-1), equivalent to 60 ng x ml(-1) of plasma VTG and allowed us to quantify the total plasma VTG of O. niloticus with high specificity and sensitivity. Using photonic and electron immunomicroscopy, we followed the pathway of VTG into the ovarian follicle (OF) demonstrating that VTG enters the oocyte at stage 3 of OF development, at the same time as cortical alveoli and lipid globules appear. Heterologous ELISAs performed on other cichlid species allowed us to quantify plasma VTG in Oreochromis aureus and Sarotherodon melanotheron and to detect it in Hemichromis fasciatus, Hemichromis bimaculatus and Tilapia zillii, constituting a reliable tool for monitoring the presence of xeno-estrogens in the environment of these fish species.

In vivo activation of the interleukin-6 receptor/gp130 signaling pathway in pituitary corticotropes of lipopolysaccharide-treated rats.

Adrenocorticotropic hormone (ACTH) release from anterior pituitary corticotropes is greatly increased during peripheral inflammation induced by lipopolysaccharide (LPS) administration. Interleukin-6 (IL-6) is thought to participate in LPS-induced ACTH release, but whether or not corticotropes are directly targeted by this cytokine is unclear. Therefore, we investigated the expression and activation of IL-6 signaling components in the pituitary of rats 2 and 4 h after administration of LPS (250 microg/kg). Intraperitoneal LPS treatment provoked the nuclear translocation of signal transducer and activator of transcription 3 (STAT-3) and Fos expression in the anterior pituitary lobe, as demonstrated by immunohistochemistry. By using in situ hybridization, we demonstrated that suppressor of cytokine signaling 3 (SOCS-3) and c-fos mRNAs were significantly induced by the LPS treatment in the anterior lobe of the pituitary. Dual in situ hybridization revealed that most corticotropes expressed IL-6 receptor and gp130 mRNAs, and that 2 h after LPS treatment, SOCS-3 and c-fos mRNAs were induced in corticotropes. Our results suggest that LPS-induced IL-6 could regulate the hypothalamo-pituitary-adrenal axis by directly targeting corticotropes during peripheral inflammation.