Growth hormone-releasing hexapeptide (GHRP-6) increases intracellular calcium concentrations in cultured cells from human pituitary adenomas of different types Andrea Lania, Emilia Ballare ´, Sabrina Corbetta, Marcello Filopanti, Luca Persani 1 and Anna Spada Institute of Endocrine Sciences, Maggiore Hospital IRCCS, University of Milan, Italy and 1 Italian Auxological Institute IRCCS, Milan, Italy (Correspondence should be addressed to A Spada, Institute of Endocrine Sciences, Universityof Milan, Ospedale Maggiore IRCCS, via F. Sforza 35, 20122 Milan, Italy) Abstract Objective: The GH-releasing peptide GHRP-6, has been found to interact with specific receptors in somatotrophs, causing cytosolic Ca 2þ ([Ca 2þ ] i ) rise and GH release. Moreover, this peptide has been demonstrated to stimulate the secretion of pituitary hormones other than GH, i.e. ACTH and prolactin, this effect being generally attributed to a central action. In this study we evaluated whether the pituitary action of this peptide is restricted to cell type of somatotroph lineage. Methods: The effect opf GHRP-6 on [Ca 2þ ] i was tested in cell preparations obtained from a series of human pituitary adenomas (9 GH-secreting adenomas, 7 nonfunctioning adenomas, 3 ACTH- secreting adenomas, 2 TSH-secreting adenomas and 1 prolactinoma) loaded with the Ca 2þ indicator fura-2. Results: GHRP-6, at concentrations higher than 1 nmol/l, significantly increased [Ca 2þ ] i in all tumours, with the exception of the 3 ACTH-secreting adenomas in which the peptide was ineffective at any concentration tested (from 1 nmol/l to 1 mmol/l). By contrast, in all ACTH-secreting adenomas, both corticotrophin-releasing hormone and pituitary adenylate cyclase activating peptide caused a marked [Ca 2þ ] i increase. In tumours responsive to GHRP-6, the peptide caused a typical biphasic [Ca 2þ ] i rise due to Ca 2þ mobilization from the intracellular stores and Ca 2þ influx through voltage- dependent Ca 2þ channels. Conclusions: These data indicate that almost all tumoral pituitary cell types are targets of GHRP-6 action, the only exception being corticotrophs. European Journal of Endocrinology 139 343–348 Introduction The secretion of growth hormone (GH) is under the complex control of both stimulatory and inhibitory agents. Although evidence indicates that growth hormone-releasing hormone (GHRH) is the physiologi- cal stimulator of GH secretion, other peptides have been demonstrated to influence GH secretion both in vivo and in vitro. In particular, synthetic peptides derived from enkephalins, such as the GH-releasing peptide, GHRP-6, have been found specifically to stimulate GH secretion both in animals and in humans (1–6). Although endo- genous GHRPs have not yet been identified, the presence of peptides with GH releasing properties has been further suggested by the recent cloning of a G-protein- coupled receptor specifically activated by GHRPs and expressed in the pituitary and hypothalamus (7–9). The action of GHRP-6 seems to be mediated via activation of phospholipase C and phosphoinositide breakdown, together with increased Ca 2þ influx through voltage- dependent calcium channels (1, 10–12). In addition to the stimulatory action at the hypothalamic level, in vitro studies with synthetic GHPR-6 demonstrated that this compound may act specifically on somatotrophs to stimulate GH release (10, 12). Moreover, it has been observed that GHRPs may stimulate the secretion of other pituitary hormones – adrenocorticotrophic hor- mone (ACTH) and prolactin (PRL) – this effect being generally attributed to a central action of these compounds (1, 13–16). It is well established that human pituitary adenomas express receptors for a large number of hypothalamic neuropeptides (17). In particular, GHRH is effective in increasing the production of cyclic AMP and the in vitro release of GH from 50% of GH-secreting adenomas (18), the lack of GHRH effect being, at least in part, due to mutations of the Gsa gene, leading to constitutive activation of adenylyl cyclase (19). Similarly, these tumours are sensitive to GHRP-6, as this agent has been reported to induce an increase in the formation of inositol 1,4,5-trisphosphate (IP 3 ) and intracellular calcium concentrations in tumoral somatotrophs (20). European Journal of Endocrinology (1998) 139 343–348 ISSN 0804-4643  1998 Society of the European Journal of Endocrinology