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

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 125I-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 125I-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 (Kd) of (1.5 +/- 0.3) x 10(-9) and (2.1 +/- 0.4) x 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 125I-labelled Tyr-Ala-hexarelin. The binding of 125I-labelled 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 [D-Arg1-D-Phe5-D-Trp7,9-Leu11]-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.

Effects of GH and IGF-I administration on GHRH and somatostatin mRNA levels: I. A study on ad libitum fed and starved adult male rats.

The individual role played by GH and IGF-I in the regulation of hypothalamic GHRH and SRIF gene expression is still object of debate. We have investigated the effect of exogenously administered recombinant hGH (rhGH) and recombinant hIGF-I (rhIGF-I) in ad libitum fed control and starved rats, the latter an animal model which is characterized by low circulating levels of endogenous GH and IGF-I. Adult male rats were fed ad libitum (C) or food-deprived (S) for 72 hours; rats in either C or S groups were treated with systemic administration of rhGH and rhIGF-I for 3 days. GHRH, SRIF and GH mRNA levels were evaluated by Northern and slot blot hybridization. Administration of rhGH (250 micrograms/kg/twice daily, sc) induced a significant inhibition of GHRH and a significant stimulation of SRIF mRNA levels in C rats; GH treatment was, however, ineffective on both neuropeptide mRNA levels in the S group. Continuous infusion of rhIGF-I (300 micrograms/kg/day, sc) induced a significant increase of SRIF levels in both C and S rats but did not modify GHRH mRNA levels in either group. In the pituitary, GH mRNA levels followed a pattern very similar to that of GHRH. These results provide evidence for a direct role of GH in the inhibition of GHRH mRNA levels; IGF-I appears more involved in the direct stimulation of SRIF mRNA levels.