ABSTRACT
Title
Correlations between endocrine and extraendocrine actions of growth hormone secretagogues mediated by GHS-R1A receptor.
Authors
L. Rizzi, L. Tamiazzo,E. Bresciani, E. Vigano, M. Ravelli, V. Locatelli, A. Torsello
Dept. of Experimental Medicine University of Milan-Bicocca, Monza
Dept. of Experimental Medicine University of Milan-Bicocca, Monza
Abstract
Growth hormone secretagogues (GHS) are synthetic peptidyl and non-peptidyl molecules, that share with ghrelin several properties; they are ligands of the GHS-R1a and modify GH release and food intake. Aim of the present study was the characterization of endocrine and extraendocrine actions of new molecules that were designed starting from a triazole scaffold. We have investigated their effects in vitro on free intracellular calcium levels and in vivo on food intake and GH secretion in the rat.
In vitro we used CHO cells transfected with the h-GHS-R1a. The effect of the GHSs were measured in terms of intracellular calcium mobilisation and compared with those of hexarelin, an hexapeptidic GHS already widely known for its GH-releasing and orexigenic activities. Ghrelin and hexarelin stimulated calcium mobilisation in a concentration-dependent fashion. This effect is followed by a massive GHS-R1a desensitisation, since a second challenge with the same compounds given 5 min later did not increase intracellular calcium levels. JMV 3002 and JMV 3012 also stimulated intracellular calcium mobilisation. At difference with the other analogs, JMV 3012 did not induce receptor desensitization. JMV 2959 did not stimulate intracellular calcium mobilisation nor induce receptor desensitization. Fluorescence microscopy showed massive receptor internalization after stimulation with ghrelin and hexarelin.
In vivo, the 10-day old rat model was used to study the GH stimulating activity of the test compounds. Hexarelin and ghrelin significantly stimulated GH secretion while all JMV compounds had no effect. Pre-treatment with the JMV3002, JMV 3012 and JMV 2959 molecules didn’t modify hexarelin stimulation of GH release.
To study the effects on food intake, we used young-adult rats. Hexarelin and ghrelin effectively stimulated feeding behaviour. JMV compounds given alone didn’t stimulate food intake whereas they attenuated the orexigenic effects of hexarelin.
In conclusion, our results demonstrate that in vitro ghrelin and synthetic GHSs can induce the internalisation of their receptor, but more studies are needed to explain the discrepancies between their endocrine and extraendocrine actions.
In vitro we used CHO cells transfected with the h-GHS-R1a. The effect of the GHSs were measured in terms of intracellular calcium mobilisation and compared with those of hexarelin, an hexapeptidic GHS already widely known for its GH-releasing and orexigenic activities. Ghrelin and hexarelin stimulated calcium mobilisation in a concentration-dependent fashion. This effect is followed by a massive GHS-R1a desensitisation, since a second challenge with the same compounds given 5 min later did not increase intracellular calcium levels. JMV 3002 and JMV 3012 also stimulated intracellular calcium mobilisation. At difference with the other analogs, JMV 3012 did not induce receptor desensitization. JMV 2959 did not stimulate intracellular calcium mobilisation nor induce receptor desensitization. Fluorescence microscopy showed massive receptor internalization after stimulation with ghrelin and hexarelin.
In vivo, the 10-day old rat model was used to study the GH stimulating activity of the test compounds. Hexarelin and ghrelin significantly stimulated GH secretion while all JMV compounds had no effect. Pre-treatment with the JMV3002, JMV 3012 and JMV 2959 molecules didn’t modify hexarelin stimulation of GH release.
To study the effects on food intake, we used young-adult rats. Hexarelin and ghrelin effectively stimulated feeding behaviour. JMV compounds given alone didn’t stimulate food intake whereas they attenuated the orexigenic effects of hexarelin.
In conclusion, our results demonstrate that in vitro ghrelin and synthetic GHSs can induce the internalisation of their receptor, but more studies are needed to explain the discrepancies between their endocrine and extraendocrine actions.