The effects of ionizing radiation on pineal melatonin and on key enzymes of its metabolism have been studied in our laboratory. After adaptation to an artificial light/dark cycle of 12:12 h, male Wistar rats were fractionally whole-body irradiated with a dose of 2.4 Gy of gamma-rays twice a week up to total doses of 4.8, 9.6 or 14.4 Gy. Irradiation and sham-irradiation were performed in the late afternoon. The rats were sacrificed at 24:00 to 01:00 h in darkness, 6 h, 3 or 5 days after the last exposure. Pineal and serum melatonin concentrations, pineal activities of serotonin N-acetyltransferase (NAT) and of monoamine oxidase (MAO) were determined. The NAT activities in the rats irradiated with 4.8 and 9.6 Gy decreased at some intervals without changes of melatonin concentration. Irradiation with a total dose of 14.4 Gy decreased NAT activity and the concentration of pineal and serum melatonin 6 h and 3 days after the last exposure. The activity of MAO, estimated only in the rats irradiated with the dose of 14.4 Gy, increased significantly 3 days after irradiation. The fractionated irradiation up to the dose of 14.4 Gy caused a transient decrease in pineal melatonin synthesis. This could be the consequence of preferential oxidative deamination of serotonin in comparison with its N-acetylation, leading to melatonin biosynthesis.