Claude Colard

Insulin-induced hypoglycemia increases preprohypocretin (orexin) mRNA in the rat lateral hypothalamic area

The recent identification of two peptides named hypocretins (Hcrt), and expressed in neurons of the rat tuberal lateral hypothalamus (LHA) previously detected by an ovine prolactin antiserum, led us to revisit some experimental procedures intented to understand the physiological roles of these neurons. In the present study, rats received intraperitoneal injections of insulin and/or glucose. Immunocytochemical observations and quantitation of in situ hybridization signals pointed out a clear stimulation of Hcrt neurons following the sole injection of insulin in hypoglycemic but not in hyperglycemic conditions. This result, together with the robust appetite boosting effect of Hcrt reported elsewhere, suggests the involvement of Hcrt neurons in the control of food intake.

Immunocytochemical localization of insulin-related peptide(s) in the central nervous system of the snail Helix aspersa Müller: involvement in growth control.

Summary1.The presence of insulin-like substances has been demonstrated by immunocytochemistry in the central nervous system of the snailHelix aspersa.2.The immunopositivity has been observed especially in the large perikarya of the mesocerebral green cells [the cerebral green cells (CeGC) stained in green by the alcian blue: alcian yellow technique].3.The removal of either the mesocerebrum or the CeGC stops the growth of the snail and induces the increase of the glycogen content in the mantle edge.4.Our results show the existence of insulin-like material in the neurosecretory cells. Previous data having demonstrated the presence of specific binding sites to insulin in the cephalic ganglia ofHelix aspersa, one may suggest that insulin could play a neuromodulatory or a neurotransmittory role in the central nervous system and might control the growth.

Evidence for the expression of dynorphin gene in the prolactin-immunoreactive neurons of the rat lateral hypothalamus

Dynorphin B (DYN B) immunoreactivity was recently reported in a population of prolactin (PRL)-immunoreactive neurons of the rat lateral hypothalamus (LH). By coupling immunocytochemistry and in situ hybridization using two synthetic oligonucleotide probes complementary to DYN mRNA, a hybridization signal was observed over the neurons of the LH exhibiting both DYN- and PRL-like immunoreactivities. Our results clearly demonstrate that these neurons contain the mRNA encoding preproDYN and are able to synthesize authentic DYN B in colocalization with a peptide related to PRL.

Activation of the rat melanin-concentrating hormone neurons by ventromedial hypothalamic lesions

The occurrence of a melanin-concentrating hormone-like peptide (MCH) was previously reported in the lateral hypothalamus of the rat. The sequence of this peptide was determined but its role as well as its regulation remain unclear. In the present study, we examined the effects of minor electrolytic lesions of the ventromedial nuclei (VMN) on MCH neurons by using immunocytochemical and in situ hybridization procedures. We report that VMN lesions resulted in (1) a clear elevation in the number and staining intensity of MCH immunoreactive perikarya and fibres, (2) a significant increase in the level of hybridocytochemical signal obtained with an oligonucleotide probe complementary to rMCH mRNA. These data provide evidence for a role of VMN in modulating the MCH gene a peptide expression.

The synthesis of melanin-concentrating hormone is stimulated by ventromedial hypothalamic lesions in the rat lateral hypothalamus: a time-course study

The activity of melanin-concentrating (MCH) neurons, was investigated by immunocytochemical and hybridocytochemical techniques in male rats bearing limited lesions of the ventromedial hypothalamic nuclei (VMN). 2 days after operation, the abundance of immunoreactive cell bodies and fibres and the intensity of labelling seemed slightly decreased in lesioned rats as compared to controls while no significant difference could be detected in MCH gene expression. After 8 days, synthesis, storage and transport of MCH appeared strongly stimulated and this stimulation lasted until the end of the experiment (day 35), suggesting that VMN plays a physiological role in controlling MCH neuron activity.

Induction of Fos-like immunoreactivity in rat oxytocin neurons following insulin injections

Double immunostaining for oxytocin (OT) and Fos was used to study the oxytocinergic system of the rat hypothalamic paraventricular nucleus (PVN) following intraperitoneal insulin injections. The expression of c-fos in the PVN appeared about 3 h after insulin treatment and was very high after 5 h while no labelling was observed in isotonic saline-injected animals. Twelve to 18% of OT neurons expressed Fos-like immunoreactivity and these activated neurons were found in both the magno- and the parvocellular compartments of the PVN suggesting that the OT neuron responses to insulin induced disturbances are complex and involve hormonal as well as autonomic pathways.

Polyethylene glycol-induced hypovolemia affects the expression of MCH mRNA, but not dynorphin or secretogranin II mRNAs, in the rat lateral hypothalamus.

Two prominent neuron populations of the rat lateral hypothalamus express genes encoding respectively the prepromelanin-concentrating hormone (MCH) or dynorphin (DYN) and secretogranin II (SGII). Their roles remain hypothetical in mammals. In the present study, we examined the changes in MCH, DYN and SGII gene expression in dehydrated rats compared to controls. Dehydration was obtained by subcutaneous injection of polyethylene glycol (PEG) resulting in a large reduction of the extracellular fluid volume. Using competitive semi-quantitative RT-PCR and in situ hybridization methods, PEG-injected animals showed a significant increase of MCH mRNA level but no variation of DYN and SGII mRNA levels. These results confirm previous observations suggesting that intra- and extracellular dehydration challenges affect different regulation circuits; they indicate that both neuron populations could be involved in the maintenance of body fluid homeostasis, directly, or indirectly, as integrators of various information leading to goal-oriented behaviour.

Ontogenic development of prolactin immunoreactive neurons in the rat lateral hypothalamus

The present study investigated the ontogenic expression of a prolactin-like substance (oPRL-ir) in rat hypothalamus from embryonic day (E) 17 to postnatal day (P) 29. By immunocytochemistry, the oPRL-ir peptide was only detected from P3. As in adults, labeled neurons were found exclusively in the lateral hypothalamic area. By in situ hybridization, with a cocktail of oligonucleotides complementary to the PRL mRNA, no labeling was observed in the hypothalamus, although dense labeling was obtained over the pituitary. With reverse-transcriptase polymerase chain reaction, a 408 bp band, presumably corresponding to an oPRL mRNA, was detected from PO in the LHA, but also in other brain regions. These results suggest that the oPRL-ir neurons do not contain oPRL. The nature of the oPRL-ir peptide is still unknown, but its late onset of expression may be related to its putative involvement in feeding behavior.

Immunocytochemical detection of different (neuro)peptides in the central nervous system of the scallop Pecten maximus

Abstract 1. Immunocytochemical investigations with antibodies raised against several biologically active peptides have been carried out on cerebro-pedal and visceral ganglia of the scallop Pecten maximus, at different stages of reproductive cycle. 2. All antisera used (anti-FMRFa, anti-methionine-enkephalin, anti-somatostatin, anti-insulin, anti-APGWa and anti-αCDCP) revealed immunoreactive structures at any investigated stage. 3. Description and localization of positive neurons and fibres are given for each antiserum. 4. The highest numbers of reacting cells were found with anti-FMRFa and methionine-enkephalin whereas αCDCP-like material was the less abundant.