The peptide oxytocin (OT) is secreted by hypothalamic neurons and exerts numerous actions related to reproduction. OT stimulation of prolactin secretion in female rats is important during the estrous cycle, pregnancy, and lactation. Here we report that OT also stimulates transients of intracellular Ca(2+) concentration in somatotrophs and gonadotrophs as well as the release of GH and LH in a dose-dependent manner with EC50 values that closely correspond to the ligand affinity of the OT receptor (OTR). Remarkably, the hormone-releasing effect of OT in these two cell types is 2 orders of magnitude more sensitive than that in lactotrophs. The specific OTR agonist [Thr(4),Gly(7)]-oxytocin acutely stimulated the release of LH, GH, and prolactin from female rat pituitary cells in primary culture and increased intracellular Ca(2+) concentration in gonadotrophs, somatotrophs, and lactotrophs. In these three cell types, the effects on hormone release and intracellular Ca(2+) of both OT and [Thr(4),Gly(7)]oxytocin were abolished by the specific OT receptor antagonist desGly-NH2-d(CH2)5[D-Tyr(2),Thr(4)]OVT but not by the highly selective vasopressin V1a receptor antagonist, d(CH2)5[Tyr(Me)(2),Dab(5)]AVP. Furthermore, 10 nM arginine vasopressin stimulated LH and GH release comparably with a dose of OT that was at least 10 times lower. Finally, the presence of the OTR-like immunoreactivity could be observed in all three cell types. Taken together, these results show that OT directly stimulates gonadotrophs, somatotrophs, and lactotrophs through OT receptors and suggest that OT signaling may serve to coordinate the release of different pituitary hormones during specific physiological conditions.