The Synergistic Effects of His-D-Trp-Ala-Trp-D-Phe- Lys-NH2on Growth Hormone (GH)-Releasing Factor- Stimulated GH Release and Intracellular Adenosine 3',5'-Monophosphate Accumulation in Rat Primary Pituitary Cell Culture

His-D-Trp-Ala-Trp-D-Phe-Lys-NH2 (GHRP-6) stimulated GH release from rat primary pituitary cells in a timeand dose-dependent manner. Stimulation was observed after a 15-min, but not a 4-h, incubation. The concentrations of GHRP- 6 required for half-maximal and maximal stimulation were 7 X 10-9 and 10-7 M, respectively. GH release induced by GHRP-6 was not affected by the addition of either naloxone or the GRF antagonist [AT-Ac-Tyr1,D-Arg2]GRF-(l-29)-NH2. The latter inhibited GRF-stimulated GH release by shifting the dose-response curve to the right. His-D-Trp-D-Lys-Trp-D-Phe-Lys- NH2, an analog of GHRP-6, inhibited GH release stimulated by GHRP-6 without affecting that induced by GRF. When present together at maximal concentrations, GHRP-6 and GRF produced a synergistic effect on GH release. GHRP-6 had no effect on intracellular cAMP levels, whereas GRF increased intracellular cAMP concentrations by 3-fold. Combined treatment of pituitary cells with GRF and GHRP-6 resulted in a potentiation of the GRF-induced increase in cAMP levels. Basal GH release was reduced by 30% after pretreatment with GHRP-6 (10-7 M) for 1 h. Pretreatment with GHRP-6 also decreased the subsequent response to GHRP-6, but not GRF. In contrast, pretreatment with GRF for 1 h had no effect on the subsequent action of GHRP-6 or GRF on GH release. The desensitization induced by GHRP-6 was completely reversed within 1 h after removal of the peptide. Results from this study indicate that GHRP-6 and GRF stimulated GH release from somatotrophs via different receptors and through discrete mechanisms. (Endocrinology124: 2791-2798, 1989)