Progesterone (P) powerfully inhibits the neuroendocrine reproductive axis, but the mechanisms and site or sites of action of this steroid remain poorly understood. Progesterone exposure during the luteal phase also alters the responsiveness of the hypothalamus to increased concentrations of estrogen (E) during the follicular phase. Using an ovariectomized ovine follicular phase model, we investigated whether the amplitude and duration of the luteal phase increase in circulating P affects the E-induced surge in LH. Treatment of ewes for 10 days with two, one, or half an intravaginal P-releasing implant or with an empty implant demonstrated that P concentrations significantly (P < 0.0001) delayed the time to surge onset upon exposure to an equal concentration of E. This delay was not due to a time-related difference in responsiveness to E after P clearance because the time of surge onset was not different when E treatment began 6, 12, or 24 h after the withdrawal of two P implants that had been present for 10 days. The final study demonstrated that the duration of P before treatment (5, 10, or 30 days) significantly (P < 0.0001) delayed the responsiveness of the estradiol-dependent surge-generating system. There was no effect on surge amplitude or duration in any experiment. Thus, the amplitude and duration of exposure to luteal phase P significantly affect the neural elements targeted by E to induce the preovulatory LH surge.