The effects of the synthetic GH-releasing peptides, GHRP-2 and GHRP-6, on phosphatidylinositol (PI) hydrolysis and cAMP production have been examined in human pituitary somatotropinomas with and without adenylyl cyclase-activating gsp oncogenes. Both peptides dose-dependently stimulated the rate of PI hydrolysis and GH secretion by cell cultures of both types of somatotropinoma. GHRP-2 was considerably more potent than GHRP-6. The effects on GH secretion were reduced or abolished by phloretin, an inhibitor of protein kinase C, and W7, an inhibitor of calmodulin. However, antagonism of the GHRH-receptor and of protein kinase A with (N-Ac-Tyr1,D-Arg2)GRF-(1-29)-NH2 and Rp-adenosine-3',5'-cyclic monophosphothioate, respectively, did not alter the stimulatory effects of GHRP-2 and GHRP-6 on GH secretion. The effect of GHRP-2 and/or GHRP-6 on cAMP production was studied in 15 tumors, seven of which possessed constitutive adenylyl cyclase activity as evidenced by presence of gsp oncogenes. Both peptides stimulated cAMP production in the latter but not former types of tumor. Moreover, GHRP-2 and GHRP-6 potentiated the stimulation of cAMP production induced by GHRH and pituitary adenylate cyclase-activating polypeptide in tumors without gsp oncogenes. These results demonstrate that GHRP-2 and GHRP-6 exert identical effects on human pituitary somatotropinomas, except for differences in potency. Additionally, under conditions of adenylyl cyclase activity above basal levels (i.e. through stimulation of G2-protein coupled receptors or because of gsp oncogene expression), cAMP production can be increased even further by GHRP, providing evidence for cross-talk between the PI and adenylyl cyclase transduction systems in pituitary cells.