Pineal gland N-acetyltransferase (NAT) activity and pineal and serum levels of melatonin declined linearly in albino rats acutely exposed to different intensities of red light (600 nm or higher; low, 140 microW/cm2; moderate, 690 microW/cm2; high, 1200 microW/cm2) during the middle of the night. The high intensity red light was as effective as white light (780 microW/cm2) in suppressing NAT activity and pineal and circulating melatonin. Red-light-inhibited nighttime NAT activity and suppressed nocturnal melatonin levels in both retinally degenerate and normal rats. Pretreatment with the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (10 mg/kg intraperitoneally) completely prevented the red-light-induced inhibition of nighttime melatonin synthesis. Magnesium chloride (300 mg/kg intraperitoneally) reduced the inhibitory effects of low and moderate intensities of red light but was ineffective when high red-light intensity was used. However, both agents failed to antagonize the suppression of nighttime melatonin synthesis elicted by the exposure to white light. Since retinally degenerate and retinally normal animals respond in the same way to both red-light and pharmacological intervention with the NMDA receptor blocker MK-801, the findings indicate that the activation of central hypothalamic NMDA receptors might mediate the photic inhibition of nocturnal melatonin synthesis in the pineal gland elicited by the exposure to red light at night. Red-light-induced suppression of nocturnal melatonin synthesis possibly can be used to investigate the biochemical mechanisms by which light entrains melatonin synthesis and to study the pharmacological and physiological effects of endogenous and synthetic agents that antagonize the NMDA receptor response.