The effect of central, short-term melatonin administration on daily GnRH and LH secretion was studied in ewes during seasonal anestrus. Melatonin, in a total dose of 32 micrograms and the vehicle were perfused for 4 hours into the mediobasal hypothalamus/median eminence (MBH/ME). The mean GnRH concentration during perfusion with melatonin decreased significantly (P < 0.05), as compared to the concentration during the preceding perfusion with the vehicle only. This change resulted from high variations in GnRH concentration noted during the initial phase of perfusion rather than from an action of melatonin. Melatonin perfused into the MBH/ME did not significantly affect LH secretion. A higher dose of melatonin and vehicle were also infused intracerebroventricularly (icv.) in either intact (300 micrograms for 3 hours) or ovariectomized (OVX) ewes (400 micrograms for 4 hours, 100 micrograms/100 microliters/h). In the intact animals, melatonin did not significantly affect LH secretion. Interestingly, melatonin significantly decreased (P < 0.05) the number of LH peaks in OVX ewes. These results demonstrate that melatonin delivered for a few hours directly into the central nervous system did not affect either daily hypothalamic GnRH release or pituitary LH secretion in intact ewes during seasonal anestrus, but did modify pulsatile LH secretion in ewes deprived of the negative feedback of estradiol.