The relative importance of several phospholipid pathways in cyclic AMP (cAMP) metabolism and growth hormone (GH) release was determined by an indirect, pharmacological approach in cultured anterior pituitary cells. The diglyceride lipase inhibitor RHC-80267 (30–100 µM) had no significant effect on cAMP levels but markedly inhibited basal and growth hormone-releasing factor- (GRF) stimulated GH secretion. A phospholipase A2 inhibitor quinacrine (30 µM) increased cellular cAMP content while decreasing GH release. Indomethacin, which reduces cyclooxygenase activity, affected neither cAMP levels nor GRF-enhanced GH release; this drug (30–100 µM) did reduce basal GH release. The lipoxygenase inhibitors nordihydroguaiaretic acid and BW-755c both reduced basal and GRF-stimulated GH release in a concentration-dependent manner. Both agents had various effects on cAMP levels. These results suggest that phospholipid metabolism, through both the cyclooxygenase and lipoxygenase pathways, contributes to basal GH release, wihle the lipoxygenase route predominates in GRF-stimulated GH release in vitro. Interestingly, cAMP metabolism can be dissociated from GH release with some of these probes, indicating an action of phospholipid metabolites distal or lateral to the cAMP-generating