Unsaturated fatty acids prevent desensitization of the human growth hormone secretagogue receptor by blocking its internalization.

The composition of the plasma membrane affects the responsiveness of cells to metabolically important hormones such as insulin and vasoactive intestinal peptide. Ghrelin is a metabolically regulated hormone that activates the G protein-coupled receptor GH secretagogue receptor type 1a (GHSR) not only in the pituitary gland but also in peripheral tissues such as the pancreas, stomach, and T cells in the circulation. We have investigated the effects of lipids and altered plasma membrane composition on GHSR activation. Oligounsaturated fatty acids (OFAs) disrupt the structure of membranes and make them more fluid. Prolonged (96 h), but not acute, treatment of the GHSR cells with the 18C OFAs oleic and linoleic acid caused a significant increase in sensitivity of the receptor to ghrelin (EC(50) reduced by a factor of 2.4 and 2.9 at 60 and 120 microM OFAs, respectively). OFAs were found to block the inhibitory effects of ghrelin pretreatment on subsequent ghrelin responsiveness, suggesting that OFAs suppress desensitization of GHSR. Radioligand displacement studies did not show a significant shift in receptor binding after incubation with OFAs. However, it was found that OFA treatment suppressed GHSR internalization, likely explaining OFA-induced refractoriness to ligand-induced desensitization. The involvement of lipid rafts in this process was indicated by the altered responsiveness of GHSR under conditions that alter membrane cholesterol. In conclusion, our findings demonstrate the importance of membrane composition for GHSR activation and desensitization and indicate at least part of the mechanism through which OFAs and cholesterol could affect ghrelin's activity in vivo.

Bolus administration of obestatin does not change glucose and insulin levels neither in the systemic nor in the portal circulation of the rat.

Obestatin is a second peptide derived from the preproghrelin polypeptide. It was originally thought to have anorexigenic effects, thereby functioning as an antagonist of ghrelin. However, this has been a subject of debate ever since. Since acylated ghrelin strongly induces insulin resistance, it could be hypothesized that obestatin plays a role in glucose homeostasis as well. In the present study we evaluated the effect of obestatin on glucose and insulin metabolism in the systemic and portal circulation. Obestatin 200 nmol/kg was administered systemically as a single intravenous bolus injection to fasted pentobarbital anesthetized adult male Wistar rats. Up to 50 min after administration, blood samples were taken to measure glucose and insulin concentrations, both in the portal and in the systemic circulation. The effect of obestatin was evaluated in fasted and in glucose-stimulated conditions (IVGTT) and compared to control groups treated with saline or IVGTT, respectively. Intravenous administration of obestatin did not have any effect on glucose and insulin concentrations, neither systemic nor portal, when compared to the control groups. Only the glucose peak 1 min after administration of IVGTT was slightly higher in the obestatin treated rats: 605.8+/-106.3% vs. 522.2+/-47.1% in the portal circulation, respectively (NS), and 800.7+/-78.7% vs. 549.6+/-37.0% in the systemic circulation, respectively (P<0.02), but it can be debated whether this has any clinical relevance. In the present study, we demonstrated that intravenously administered obestatin does not influence glucose and insulin concentrations, neither in the portal nor in the systemic circulation.

Unacylated ghrelin is not a functional antagonist but a full agonist of the type 1a growth hormone secretagogue receptor (GHS-R).

Recent findings demonstrate that the effects of ghrelin can be abrogated by co-administered unacylated ghrelin (UAG). Since the general consensus is that UAG does not interact with the type 1a growth hormone secretagogue receptor (GHS-R), a possible mechanism of action for this antagonistic effect is via another receptor. However, functional antagonism of the GHS-R by UAG has not been explored extensively. In this study we used human GHS-R and aequorin expressing CHO-K1 cells to measure [Ca(2+)](i) following treatment with UAG. UAG at up to 10(-5)M did not antagonize ghrelin induced [Ca(2+)](i). However, UAG was found to be a full agonist of the GHS-R with an EC(50) of between 1.6 and 2 microM using this in vitro system. Correspondingly, UAG displaced radio-labeled ghrelin from the GHS-R with an IC(50) of 13 microM. In addition, GHS-R antagonists were found to block UAG induced [Ca(2+)](i) with approximately similar potency to their effect on ghrelin activation of the GHS-R, suggesting a similar mode of action. These findings demonstrate in a defined system that UAG does not antagonize activation of the GHS-R by ghrelin. But our findings also emphasize the importance of assessing the concentration of UAG used in both in vitro and in vivo experimental systems that are aimed at examining GHS-R independent effects. Where local concentrations of UAG may reach the high nanomolar to micromolar range, assignment of GHS-R independent effects should be made with caution.

Unacylated ghrelin acts as a potent insulin secretagogue in glucose-stimulated conditions.

Acylated and unacylated ghrelin (AG and UAG) are gut hormones that exert pleiotropic actions, including regulation of insulin secretion and glucose metabolism. In this study, we investigated whether AG and UAG differentially regulate portal and systemic insulin levels after a glucose load. We studied the effects of the administration of AG (30 nmol/kg), UAG (3 and 30 nmol/kg), the ghrelin receptor antagonist [D-Lys(3)]GHRP-6 (1 micromol/kg), or various combinations of these compounds on portal and systemic levels of glucose and insulin after an intravenous glucose tolerance test (IVGTT, d-glucose 1 g/kg) in anesthetized fasted Wistar rats. UAG administration potently and dose-dependently enhanced the rise of insulin concentration induced by IVGTT in the portal and, to a lesser extent, the systemic circulation. This UAG-induced effect was completely blocked by the coadministration of exogenous AG at equimolar concentrations. Similarly to UAG, [D-Lys(3)]GHRP-6, alone or in combination with AG and UAG, strongly enhanced the portal insulin response to IVGTT, whereas exogenous AG alone did not exert any further effect. Our data demonstrate that, in glucose-stimulated conditions, exogenous UAG acts as a potent insulin secretagogue, whereas endogenous AG exerts a maximal tonic inhibition on glucose-induced insulin release.