Summary. The ovulating hormone (oh) of the pituitary gland was determined in untreated, 30-day-old immature female rats given an ovulating dose of pregnant mare's serum gonadotrophin (pmsg), and in pmsg-primed rats given sodium phenobarbital (PB) before the critical period (13.30 hours) or after the critical period (16.30 hours) on Day 32. PB given at 13.30 hours blocks ovulation for 24 hr. The adenohypophysis was removed at various times from 14.00 hours on Day 32 to 07.30 hours on Day 35 and assayed for oh in 24-day-old pmsg-PB treated test rats. The number of ova ovulated by the test rats indicated the pituitary oh. In untreated animals the pituitary oh remained high. In the pmsg-primed rats, the hormone decreased during the critical period, 14.00 to 16.00 hours on Day 32, but was minimal on the morning of Day 33 and remained low through Day 35. In pmsg-treated animals injected with PB at 13.30 hours on Day 32, the oh changes were delayed for 24 hr. When PB was administered at 16.30 hours, oh decreased during the critical period on Day 32 but minimal oh on Day 33 was not as low as in the pmsg-primed rats at the same time. Instead of remaining low the hormone increased by 16.00 hours on Day 33. In ten of twelve test situations between 14.00 and 16.00 hours each day there was an oh decrease, indicating a diurnal oh secretion.