The exposure of organ-cultured pineal glands of Syrian hamsters to forskolin, an adenylate cyclase activator, caused marked increases in melatonin levels when glands were collected in the second half of the dark period and incubated for 4 h. However, when glands were collected at the beginning of the dark period and incubated with the same drug, a significant increase in melatonin content was observed only after 6-8 h. Likewise, when glands were collected at the beginning of the light period, forskolin stimulated melatonin synthesis only after 6-8 h of incubation with the drug. These results support the existence of a relatively long lag period necessary for the induction of melatonin production in the Syrian hamster pineal gland. Experiments with actinomycin D indicate that RNA synthesis occurs during the lag period; thus, actinomycin D blocks the induction of melatonin synthesis by forskolin in glands collected at the beginning of the dark period and incubated for 8 h. In contrast, when pineal glands were collected from hamsters killed in the second half of the dark period, and incubated with forskolin for either 4 or 8 h, actinomycin D was unable to block the induction of melatonin production. These data suggest that RNA, presumably messenger RNA, which is necessary for increasing hamster pineal melatonin synthesis, is synthesized and accumulates during the first half of the night.