Spontaneous firing of suprachiasmatic nucleus (SCN) neurons and the effect of melatonin (MEL) were examined in hypothalamic slices made from pinealectomized rats or from the rats exposed to constant light. The results are as follows. (1) Under normal light cycle (light dark=12 12), SCN neurons displayed a circadian rhythm in spontaneous discharges. A peak about 8.3 Hz was presented at CT (circadian time) 6 8 and a trough about 3.8 Hz at CT18 20. The circadian rhythm persisted after pinealectomy, but disappeared after constant light exposure. (2) The response of SCN neurons to MEL was mainly inhibitory. Under normal light exposure, the inhibitory proportion was higher during the subjective day with the largest proportion of about 42% at CT8 10, and was lower during the subjective night with the largest proportion of about 26% at CT22 24. Under both constant light exposure and pinealectomy, the circadian rhythm of the response of SCN neurons to MEL disappeared. (3) The inhibitory effect of MEL on spontaneous discharges of SCN neurons was blocked by ML-1 receptor antagonist luzindole, but not by ML-2 receptor antagonist prazosin. The above results suggest that pineal gland, as a regulator of the organization of circadian rhythm, regulates the circadian rhythm of SCN neurons at two responsive time windows (CT8 10 and CT22 24) through high affinity ML-1 receptors in SCN.