Prostaglandin E2 (PGE2) was infused into a lateral ventricle of adult male rats or infused into a hypophysial portal vessel. Intraventricularly administered PGE2 stimulated the release of LH, FSH, and prolactin but not TSH. Intraventricular infusion of the control solution did not alter the basal release of any of these hormones. PGE2 was found to be a potent stimulator of LH release, but larger quantities were needed to stimulate the release of FSH or prolactin. In addition, lateral ventricle infusion of 5 mug or 20 mug of PGE2 stimulated a 2- to 3-fold increase in the concentration of LHRH in hypophysial portal plasma but did not affect LHRH levels in arterial plasma. PGE2 infused into a hypophysial portal vessel at a rate of 0.167 mug/min for 30 min, or infusion of the control solution, resulted in similar changes in the concentration of FSH, LH, and prolactin in arterial plasma. The results of this investigation suggest that PGE2 stimulates the release of gonadotropins in vivo by enhancing the release of LHRH. The portal vessel infusion studies do not support a direct stimulatory effect of PGE2 on the pituitary gonadotrophs.