The effects of angiotensin II (AII) on prolactin release and arachidonate liberation were studied in anterior pituitary cells preincubated with [3H]arachidonate to label the cellular phospholipids. AII increased prolactin release and [3H]arachidonate liberation over similar concentration ranges with the dynamics of these two events proving identical. Dopamine attenuated both prolactin release and [3H]arachidonate liberation. The diacylglycerol lipase inhibitor RHC80267 decreased AII-stimulated prolactin release and arachidonate liberation. Further evidence that AII-induced release of arachidonate is mediated by a diacylglycerol lipase is suggested by the finding that AII increased [14C]stearate liberation from cells prelabeled with the fatty acid. Although arachidonate itself may have some role in prolactin secretion, it is likely that arachidonate metabolites are more directly involved because BW755c and AA861, inhibitors of arachidonate metabolite formation, increased AII-stimulated arachidonate liberation, but decreased prolactin release.