These studies investigated the effects of neuropeptide Y (NPY) on in vitro release of luteinizing hormone-releasing hormone (LHRH) from the medial basal hypothalmus (MBH) and tested whether ovarian steroids modulate the LHRH response to NPY. Ovariectomized rats were implanted with 20-mm-long Silastic capsules containing a low concentration of estradiol (E2) (150 µg/ml oil), a high concentration of E2 (250 µg/ml oil), or sesame oil vehicle. Additional animals received high-dose E2 capsules plus an injection of progesterone (15 mg) concomitantly. Two days later, individual MBH fragments were incubated in medium alone for a 30-min period to obtain the basal rate of LHRH release, followed by a second 30-min period in medium containing NPY or saline. Exposure to NPY (10–6M) increased the release of LHRH from MBH of ovarian hormone-treated, but not from hormonally untreated rats. The LHRH response was most pronounced from the MBH of rats treated with either high-dose E2 or E2 plus progesterone. The increase in LHRHrelease was also elicited by 10–7M, but not by 10–8M NPY concentrations, using MBH from E2 plus progesterone-treated rats. In addition, NPY markedly potentiated the KCl-evoked release of LHRH from MBH of ovariectomized, hormonally untreated or low-dose E2-treated rats, under conditions when there was little or no effect of NPY on the basal LHRH release. Further, the release of LHRH stimulated by NPY was not accompanied by increase in the release of norepinephrine or of dopamine. These results strongly suggest that the NPY stimulation of LH secretion after intracerebral administration in steroid-primed rats occurs, in part, as a result of increased release of LHRH. NPY may act at the level of the LHRH neurosecretory terminals in the median eminence as well as at additional sites containing LHRH pe