Specific receptors for synthetic GH secretagogues in the human brain and pituitary gland

Abstract

In vitro studies have been performed to demonstrate and characterize specific binding sites for synthetic GH secretagogues (sGHS) on membranes from pituitary gland and different human brain regions. A binding assay for sGHS was established using a peptidyl sGHS (Tyr-Ala-hexarelin) which had been radioiodinated to high specific activity at the Tyr residue. Specific binding sites for 125 I-labelled Tyr-Ala-hexarelin were detected mainly in membranes isolated from pituitary gland and hypothalamus, but they were also present in other brain areas such as choroid plexus, cerebral cortex, hippocampus and medulla oblongata with no sex-related differences. In contrast, negligible binding was found in the thalamus, striatum, substantia nigra, cerebellum and corpus callosum. The binding of 125 I-labelled Tyr-Ala-hexarelin to membrane-binding sites is a saturable and reversible process, depending on incubation time and pH of the buffer. Scatchard analysis of the binding revealed a finite number of binding sites in the hypothalamus and pituitary gland with a dissociation constant (K d ) of (1·5 0·3) 10 9 and (2·1 0·4) 10 9 mol/l respectively. Receptor activity is sensitive to trypsin and phospholipase C digestion, suggesting that protein and phospholipids are essential for the binding of 125 I-labelled Tyr-Ala-hexarelin. The binding of 125 Ilabelled Tyr-Ala-hexarelin to pituitary and hypothalamic membranes was displaced in a dose-dependent manner by different unlabelled synthetic peptidyl (Tyr-Ala-hexarelin, GHRP2, hexarelin, GHRP6) and non-peptidyl (MK 0677) sGHS. An inhibition of the specific binding was also observed when binding was performed in the presence of [-Arg 1 --Phe 5 --Trp 7,9 -Leu 11 ]-substance P, a substance P antagonist that has been found to inhibit GH release in response to sGHS. In contrast, no competition was observed in the presence of other neuropeptides (GHRH, somatostatin, galanin or Met-enkephalin) which have a known influence on GH release.In conclusion, the present data demonstrate that sGHS have specific receptors in human brain and pituitary gland and reinforce the hypothesis that these compounds could be the synthetic counterpart of an endogenous GH secretagogue involved in the neuroendocrine control of GH secretion and possibly in other central activities.

Pharmacological characterisation of a new oral GH secretagogue, NN703

“…In the light of recent publications by Muccioli et al (23) and Ong et al (24), where an alternative GHS receptor with high affinity for hexarelin but low affinity for MK677 has been described, it is possible to speculate whether the discrepancies between data using the cloned receptor and the rat pituitary cell assay or data from the cloned receptor and in vivo data may be explained by the involvement of such an alternative receptor. However, since no other GHS receptors have yet been characterised these speculations remain unsolved.…”

Ghrelin as a novel locally produced relaxing peptide of the iris sphincter and dilator muscles

“…Growth hormone secretagogues (GHS) promote GH release by direct action on the pituitary gland Locatelli and Torsello, 1997;Bowers, 1998;Dieguez and Casanueva, 2000), stimulating a specific G Protein coupled receptor (GHS type 1a receptor, Abbreviations: L-NA, N u -nitro-L-arginine; GHS, growth hormone somatosecretagogues; GHSR-1a, growth hormone somatosecretagogues receptor type 1a; COX, cycloxygenase; ET-1, endothelin 1; frGhr, 1e5 human ghrelin; dfrGhr, 1e5 human des-octanoyl-ghrelin; NO, nitric oxide; Pg, prostaglandins; EP 2 , prostaglandin E receptor 2. , which is different from GHRH receptor (Howard et al, 1996;Muccioli et al, 1998;Smith et al, 1999;Feighner et al, 1998). Binding sites to the somatosecretagogues have been described in several tissues such as (in order of decreasing binding activity) the myocardium, adrenal gland, gonads, arteries, lung, liver, skeletal muscle, kidney, pituitary, thyroid, adipose tissue, veins, uterus, skin and lymphnode Iglesias et al, 2004).…”

Cardiac effects of ghrelin and its endogenous derivatives des-octanoyl ghrelin and des-Gln14-ghrelin

“…GHS were discovered as a family of non-natural peptidyl and non-peptidyl molecules able to strongly stimulate GH secretion acting on a specific Gprotein coupled receptor, namely GHS type 1a receptor, that is different from the GHRH-receptor (Smith et al, 1997;Muccioli et al, 1998Muccioli et al, , 2002. Theoretically, GHS would therefore influence cardiac structure and function via enhanced GH and IGF-I secretion.…”