28-day-old female rats were killed at increasing time periods after the injection of either 30 IU pregnant mares’ serum (PMS) or saline, and pituitary and plasma luteinizing hormone (LH) and prolactin levels were measured by radioimmunoassay. Hormone concentrations were measured at intervals up to one week following injection with frequent measurements made on the afternoon preceding ovulation to better characterize and evaluate the PMS-treated immature rat as an ovulation model. Peaks in both plasma LH and prolactin occurred on the afternoon preceding ovulation. Pituitary LH was found to decrease and again increase during this afternoon period before falling to a low value on the morning following ovulation. Pituitary prolactin increased to an early afternoon maximum value, fell to a low value by late afternoon, and increased again by the morning following ovulation. Saline-treated immature female rats displayed fluctuations in plasma and pituitary LH and prolactin similar to but smaller than the afternoon changes found in PMS-treated rats. Pituitary weights were significantly greater in PMS-treated rats on the afternoon preceding ovulation. The hormonal changes encountered in the PMS-treated rat prior to ovulation appear to be similar to those encountered in the proestrous cycling female rat, although more study is necessary to validate this system as a model of spontaneous ovulation in the adult animal. The data suggest that PMS causes ovulation in immature rats by amplifying or synchronizing existing hormonal rhythm