Pineal serotonin N-acetyltransferase activity (NAT) and melatonin concentrations were determined at various intervals in prepubertal (35 days old) and adult male hamsters (74 day old) throughout a 24 hour period with the animals kept in a light:dark cycle of 6:18 (lights on at 0600 h and off at 1200 h). In prepubertal animals, daytime pineal NAT activity of 0.20-0.28 nmoles 14C-N-acetyltryptamine/pineal/hour was maintained for 8 hours after the initiation of darkness. Peak pineal NAT activity of 0.46 +/- 0.64 nmoles 14C-N-acetyltryptamine/pineal/hour occurred 13 hours after the onset of darkness and remained significantly elevated until 0400 h (p less than 0.001). Daytime pineal melatonin concentrations of 78-194 pg/pineal gland also were maintained for 8 hours after the initiation of darkness. At 13 hours into the dark period, pineal melatonin concentrations rose to 788 +/- 150 pg/pineal gland (p less than 0.01 vs all other time points except 0230 h and 0400 h). At one hour before the onset of light both the pineal NAT activity and pineal melatonin concentrations returned to daytime values. Adult male hamsters had diurnal pineal NAT and melatonin rhythms which are indistinguishable from those found in the prepubertal animals.