Central NUCB2/Nesfatin-1-expressing neurones belong to the hypothalamic-brainstem circuitry activated by hypoglycaemia.

Nesfatin-1 is a recently identified 82 amino acid peptide shown to have an anorexigenic effect on rodents when administrered centrally and peripherally. Nesfatin-1 is expressed not only in neurones of various brain areas, including the hypothalamic and brainstem nuclei, but also in peripheral organs, such as the stomach and the pancreas. Nesfatinergic neurones were reported to participate in the regulation of satiety signals and in the responses to other stimuli, including restraint stress, abdominal surgery, and lipopolysaccharide-induced inflammation. The present study aimed to investigate whether NUCB2/nesfatin-1 expressing neurones also take part in the central signalling activated in response to hypoglycaemia and therefore are involved in central glucose sensing. Using immunolabelling methods based on the detection of the neuronal activation marker c-Fos and of nesfatin-1, we showed that peripheral injection of insulin induced a strong activation of nesfatin-1-expressing neurones in the brain vagal-regulatory nuclei, including the arcuate nucleus, paraventricular nucleus, lateral hypothalamic area, dorsal motor nucleus of the vagus (DMNX) and nucleus of the tractus solitarius. In response to intracellular glucopaenia induced by i.p. or i.c.v. 2-deoxyglucose injection, the c-Fos/nesfatin-1 colocalisations observed at the hypothalamic and brainstem levels were similar to those observed after insulin-induced hypoglycaemia. Moreover, using Fluorogold as a retrograde tracer, we showed that nesfatinergic preganglionic DMNX neurones activated by hypoglycaemia target the stomach and the pancreas. Taken together, these results suggest that a subpopulation of nesfatinergic neurones belongs to the central network activated by hypoglycaemia, and that nesfatin-1 participates in the triggering of physiological and hormonal counter-regulations observed in response to hypoglycaemia.

Gastric electrical stimulation modulates hypothalamic corticotropin-releasing factor-producing neurons during post-operative ileus in rat.

High-frequency/low-energy gastric electrical stimulation (GES) is an efficient therapy to treat gastric emptying-related disorders but its mechanism of action remains poorly understood. We aimed to assess the effects of high-frequency/low-energy GES on corticotropin-releasing factor (CRF)-producing neurons in the paraventricular nucleus of the hypothalamus (PVN), which are involved in gastric ileus induced by laparotomy. Two electrodes were implanted in the rat gastric antrum during laparotomy, then stimulation (amplitude: 2 mA; pulse duration 330 micros; frequency: 2 Hz; 1 min ON/2 min OFF) or sham stimulation (control group) were applied. Using immunohistochemistry, the number of c-Fos protein-expressing neurons (c-Fos protein-immunoreactive cells, Fos-IR) was quantified in the PVN after 1 h of stimulation. The number of neurons expressing simultaneously c-Fos protein and CRF mRNA was measured by means of immunocytochemistry combined with in situ hybridization. Finally, c-Fos and CRF mRNA levels in the hypothalamus were determined by in situ hybridization or quantitative reverse transcriptase-polymerase chain reaction. Fos-IR in the PVN was significantly decreased 1 h after GES (P<0.05) but was not affected by sub-diaphragmatic vagotomy. The number of neurons containing c-Fos protein and CRF mRNA was lower in the GES group compared with the control group (P<0.05). In addition, c-Fos and CRF mRNA levels in the PVN were significantly decreased by GES (P

Pituitary adenylate cyclase-activating polypeptide directly modulates the activity of proopiomelanocortin neurons in the rat arcuate nucleus.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and the proopiomelanocortin (POMC)-derived peptide alpha-melanocyte-stimulating hormone (alpha-MSH) both regulate multiple neuroendocrine functions and feeding behavior. Two subtypes of PACAP receptor mRNAs, pituitary adenylate cyclase-activating polypeptide-specific receptor (PAC1-R) and pituitary adenylate cyclase-activating polypeptide/vasoactive intestinal polypeptide mutual receptor (VPAC2-R), are actively expressed in the arcuate nucleus of the hypothalamus, where POMC cell bodies are located. This observation led us to investigate the possible regulatory action of PACAP on rat POMC neurons. Double-labeling in situ hybridization histochemistry revealed that approximately 50% of POMC-producing neurons express PAC1-R and/or VPAC2-R mRNAs. The proportion of POMC neurons that also contain PAC1-R mRNA was homogeneous along the rostro-caudal axis of the arcuate nucleus while POMC-positive cell bodies expressing the VPAC2-R subtype were more abundant in the rostral region. Incubation of mediobasal hypothalamic explants with PACAP (10(-7) M; 30 min) increased POMC mRNA expression, and this effect was blocked by PACAP6-38 (10(-6) M). In contrast, incubation with vasoactive intestinal polypeptide (10(-7) M) did not affect POMC mRNA level. Incubation of hypothalamic fragments with PACAP (10(-7) M) caused a significant increase in alpha-MSH content in the tissue and in the incubation medium. Altogether, the present results reveal that exogenous PACAP, acting probably through PAC1-R, regulates the activity of POMC neurons in the rat hypothalamus. These data suggest that the effects of PACAP on the gonadotropin-releasing hormone neuroendocrine axis and the regulation of feeding behavior may be mediated, at least in part, through modulation of POMC neurons.

Adenosine A2A receptor gene disruption provokes marked changes in melanocortin content and pro-opiomelanocortin gene expression.

A2A receptor knockout (A2AR-/-) mice are more anxious and aggressive, and exhibit reduced exploratory activity than their wild-type littermates (A2AR+/+). Because alpha-melanocyte-stimulating hormone (alpha-MSH) influences anxiety, aggressiveness and motor activity, we investigated the effect of A2AR gene disruption on alpha-MSH content in discrete brain regions and pro-opiomelanocortin (POMC) expression in the hypothalamus and pituitary. No modification in alpha-MSH content was observed in the hypothalamus and medulla oblongata where POMC-expressing perikarya are located. In the arcuate nucleus of the hypothalamus, POMC mRNA levels were not affected by A2AR disruption. Conversely, in A2AR-/- mice, a significant increase in alpha-MSH content was observed in the amygdala and cerebral cortex, two regions that are innervated by POMC terminals. In the pars intermedia of the pituitary, A2AR disruption provoked a significant reduction of POMC mRNA expression associated with a decrease in alpha-MSH content. By contrast, in the anterior lobe of the pituitary, a substantial increase in POMC mRNA and adrenocorticotropin hormone concentrations was observed, and plasma corticosterone concentration was significantly higher in A2AR-/- mice, revealing hyperactivity of their pituitary-adrenocortical axis. Together, these results suggest that adenosine, acting through A2A receptors, may modulate the release of alpha-MSH in the cerebral cortex and amygdala. The data also indicate that A2A receptors are involved in the control of POMC gene expression and biosynthesis of POMC-derived peptides in pituitary melanotrophs and corticotrophs.