Growth Hormone Releasing Hexapeptide (GHRP-6) Activates the Inositol (1,4,5)-Trisphosphate/Diacylglycerol Pathway in Rat Anterior Pituitary Cells

Abstract

Growth hormone-releasing hexapeptide (GHRP-6) is known to stimulate secretion of growth hormone (GH) in vivo and in vitro in a variety of species. However, the cellular effects of GHRP-6 remain largely unknown. We have tested the influence of GHRP-6 on the inositol phospholipid second messenger system in cultured anterior pituitary cells. Cultured pituitary cells responded upon challenge with GHRP-6 with a dose-dependent release of GH. Moreover, incubation of GHRP-6 with pituitary cell cultures labelled with nrvo-[³H]inositol resulted in a dose-dependent rise in [³H]inositol phosphates. Brief stimulation of pituitary cells with GHRP-6 increased phosphorylation of MBP_4-14, a specific protein kinase C substrate, when incubated with the cytosol- or plasma membrane fraction from the stimulated cells. Furthermore, introduction of MBP_4-14, into the cytosol in digitonin permeabilized pituitary cells caused increased phosphorylation of this substrate. GHRP-6 induced a rise in intracellular Ca²⁺ in individual somatotrophs loaded with the Ca²⁺ indicator, Fura-2. Preincubation (3 min) with somatostatin (SRIF) diminished the Ca²⁺ spike elicited by GHRP-6, while no effect of SRIF was observed when added simultaneously with GHRP-6. These results indicate that GHRP-6-stimulated GH-secretion involves the diacylglycerol/inositol(1,4,5)trisphosphate pathway with a resulting rise in cytosolic Ca²⁺