cAMP is thought to be an essential intracellular mediator in the release of GH from somatotrophs. Ca++ is required in the incubation medium to elicit GH release in vitro. We have carried out studies using a purified preparation of rat somatotrophs to see whether the Ca++ requirement procedes or follows the accumulation of cAMP induced by prostaglandin E2 (which increases adenylate cyclase activity) and 3-isobutyl-1-methylxanthine (a phosphodiesterase inhibitor). Incubation of somatotrophs in low Ca++ medium (less than 85 microM) abolished the release of GH induced by the two secretagogues, while the increase in intracellular cAMP was actually augmented. Thus, Ca++ is required in the incubation medium to express the action of intracellular cAMP.