Overexpression of cannabinoid CB2 receptor in the brain induces hyperglycaemia and a lean phenotype in adult mice.

It is well known that the endocannabinoid system, through cannabinoid CB1 receptor activation, has an important role in the main aspects of energy balance (i.e. food intake, energy expenditure and glucose and fat metabolism), orchestrating all the machinery involved in body weight control and energy homeostasis. A number of studies have revealed a crucial role of brain CB1 receptors in these processes. However, functional cannabinoid CB2 receptors have also been described in the brain, with no studies addressing their putative role in body weight control and glucose homeostasis. We have tested this hypothesis by analysing fasting-induced feeding, body weight, some hypothalamic neuropeptides, glucose tolerance and plasma hormones in an animal model specifically overexpressing CB2 receptors in the central nervous system. We found that specific overexpression of CB2 receptors in the brain promoted higher basal glucose levels, decreased fasting-induced feeding and, eventually, led to a lean phenotype and glucose intolerance. These findings could not be attributed to decreased locomotor activity, increased anxiety or depressive-like behaviours. The expression of relevant neuropeptides such as pro-opiomelanocortin and galanin in the arcuate nucleus of the hypothalamus was altered but not those of the CB1 receptor. Indeed, no changes in CB1 expression were found in the liver, skeletal muscle and adipose tissue. However, cannabinoid CB1 and CB2 receptor expression in the endocrine pancreas and glucagon plasma levels were decreased. No changes in plasma adiponectin, leptin, insulin and somatostatin were found. Taken together, these results suggest a role for central cannabinoid CB2 receptors in body weight control and glucose homeostasis.

Rapid non-genomic regulation of Ca2+ signals and insulin secretion by PPAR alpha ligands in mouse pancreatic islets of Langerhans.

PPAR alpha is a ligand-activated transcription factor belonging to the nuclear receptor superfamily. PPAR alpha is involved in the regulation of in vivo triglyceride levels, presumably through its effects on fatty acid and lipoprotein metabolism. Some nuclear receptors have been involved in rapid effects mediated by non-genomic mechanisms. In this paper, we report the rapid non-genomic effects of PPAR alpha ligands on the intracellular calcium concentration ([Ca2+]i), mitochondrial function, reactive oxygen species (ROS) generation, and secretion of insulin in freshly isolated mouse islets of Langerhans. The hypolipidemic fibrate PPAR alpha agonist WY-14 643 decreased the glucose-induced calcium oscillations in intact islets. This effect was mimicked by the synthetic agonist GW7647 and the endogenous agonist oleylethanolamide. The WY-14 643 action was rapid in onset (5 min) and was still produced in the presence of protein and mRNA synthesis inhibitors, cycloheximide, and actinomycin-d. This suggests that it is independent of gene transcription. In addition, WY-14 623 impaired mitochondrial function, increased ROS formation and decreased insulin release. PPAR alpha is present in beta-cells, mainly in the cytosol and nucleus, with a small subpopulation localized in the plasma membrane. However, the presence of the PPAR alpha ligand effects in mice bearing a disrupted Ppar alpha gene raises the possibility that the rapid effects of the agonists in pancreatic beta-cells are independent of the receptor. We conclude that PPAR alpha agonists produce a decrease in glucose-induced [Ca2+]i signals and insulin secretion in beta-cells through a rapid, non-genomic mechanism.

Cannabinoid receptors regulate Ca(2+) signals and insulin secretion in pancreatic beta-cell.

Insulin is the main hormone involved in the regulation of glycaemia, its impaired secretion is a hallmark of type I and type II diabetic individuals. Additionally, insulin is involved in lipogenesis and weight gain, provoking an anorexigenic action. The endocannabinoid system contributes to the physiological regulation of energy balance, food intake and lipid and glucose metabolisms. Despite that, an experimental link between the endocannabinoid system and the endocrine pancreas has not yet been described. Using quantitative real-time PCR and immunocytochemistry, we have demonstrated the existence of both CB1 and CB2 receptors in the endocrine pancreas. While the CB1 receptor is mainly expressed in non-beta-cells, the CB2 type exists in beta- and non-beta-cells within the islet. The endocannabinoid 2-arachidonylglycerol (2-AG) through CB2 receptors regulates [Ca(2+)](i) signals in beta-cells and as a consequence, it decreases insulin secretion. This effect may be a new component involved in the orexigenic effect of endocannabinoids and constitutes a potential target for pharmacologic manipulation of the energy balance.

The subcommissural organ expresses D2, D3, D4, and D5 dopamine receptors.

Dopamine receptors have been found in certain populations of non-neuronal cells in the brain, viz., discrete areas of ciliated ependyma and the ependymal cells of the choroid plexus. We have studied the presence of both tyrosine-hydroxylase-immunoreactive nerve fibers and dopamine receptors in the subcommissural organ (SCO), an ependymal brain gland that is located in the roof of the third ventricle and that secretes, into the cerebrospinal fluid, glycoproteins that aggregate to form Reissner's fiber (RF). Antibodies against D2, D3, D4, and D5 dopamine receptors were used in immunoblots of bovine striatum, fresh SCO, and organ-cultured SCO, and in immunocytochemistry of the bovine, rat, and mouse SCO. Only a few tyrosine-hydroxylase fibers appeared to reach the SCO. However, virtually all the secretory ependymal and hypendymal cells of the SCO immunoreacted with antibodies against D2, D4, and D5 receptors, with the last-mentioned rendering the strongest reaction, especially at the ventricular cell pole of the secretory ependymocytes, suggesting that dopamine might reach the SCO via the cerebrospinal fluid. The antibodies against the four subtypes of receptors revealed corresponding bands in immunoblots of striatum and fresh SCO. Although the cultured SCO displayed dopamine receptors, dopamine had no apparent effect on the expression of the SCO-spondin gene/protein or on the release of RF-glycoproteins (SCO-spondin included) by SCO explants, suggesting that dopamine affects the function(s) of the SCO differently from the secretion of RF-glycoproteins.

Bovine subcommissural organ displays spontaneous and synchronous intracellular calcium oscillations.

The subcommissural organ (SCO) is an ependymal brain gland that secretes into the cerebrospinal fluid glycoproteins that polymerize, forming Reissner's fiber (RF). The SCO-RF complex seems to be involved in vertebrate nervous system development, although its role in adults is unknown. Furthermore, its physiology is still greatly undetermined, and little is known about the release control of SCO secretion and the underlying intracellular mechanisms. In this report, we show that up to 90% of 3-5-day-old in vitro SCO cells from both intact and partially-dispersed SCO explants displayed spontaneous cytosolic Ca2+ oscillations. The putative role of these spontaneous calcium oscillations in SCO secretory activity is discussed taking into consideration several previous findings. Two distinct subpopulations of SCO cells were detected, each one containing cells with synchronized calcium oscillations. A possible existence of different functional domains in SCO is therefore discussed. Oscillations persisted in the absence of extracellular Ca2+, indicating the major involvement of Ca2+ released from internal stores. Depolarization failed to induce intracellular calcium increases, although it disturbed the oscillation frequency, suggesting a putative modulator role of depolarizing agonists on the calcium oscillating pattern through voltage-gated calcium channels. Carbachol, a cholinergic agonist, evoked a switch in Ca2+ signaling from a calcium oscillating mode to a sustained and increased intracellular Ca2+ mode in 30% of measured cells, suggesting the involvement of acetylcholine in SCO activity, via a calcium-mediated response.