Negative feedback of estrogen was investigated in ovariectomized female guinea pigs. Two weeks after ovariectomy, indwelling catheters were inserted into the jugular vein, and 3 days later, blood samples were taken every 10 min to determine the pattern of luteinizing hormone (LH) secretion. LH secretion in these guinea pigs was episodic, with a mean pulse period of 32 min. The mean pulse amplitude was 2.1 ng/ml, with mean plasma LH levels of 1.8 ng/ml. Twenty-five micrograms 17 beta-estradiol (E2), given i.v., caused a pronounced inhibition of pulsatile LH release. Twenty-five microliters of 100% ethanol (vehicle) had no effect on plasma LH values. In a second set of experiments, ovariectomized female guinea pigs were given two injections of luteinizing hormone-releasing hormone (LHRH) (1 microgram/kg BW, i.v.) separated by 30 min. Sharp rises in serum LH values were detected after each injection. A third injection of LHRH was administered after an injection of either 25 micrograms E2 or 25 microliters vehicle. In the presence of E2, the LH response was significantly (p less than 0.005) diminished, whereas the vehicle did not change the LH response to LHRH. These rapid effects of E2 on LH secretion and the pituitary responsiveness to LHRH infusion indicate that in the ovariectomized guinea pig E2 can directly block gonadotropin secretion. These findings are consistent with the hypothesis that negative feedback actions of E2 are directly on the membrane of the gonadotrope.