In the pineal gland and retina of chickens, light exposure at night when serotonin N-acetyltransferase (NAT) activity levels are high causes a 4--5-fold decrease in NAT activity. The t1/2 of NAT inactivation is 10 min and the kinetics of inactivation are similar in the pineal gland and retina. A brief pulse of light can initiate the process of NAT inactivation which continues in the dark for 30 min before a partial recovery of NAT activity occurs. In blinded chicks, there is less inactivation of pineal NAT by light than in intact chicks, indicating that the eyes are involved in the pineal's response to light. In chicks that have had their superior cervical ganglia removed inactivation of pineal NAT by light is intermediate between that of intact and blind chicks, indicating that ganglionectomy does not completely mimic the effects of blinding. The pineal gland itself is light-sensitive in culture. Light causes a 4--5-fold decrease in NAT activity in static organ culture, and inhibits melatonin release in flow-through organ culture. Drugs that increase cyclic nucleotide levels in cells (cholera toxin, RO 20-1724, monobutyryl cyclic AMP, monobutyryl cyclic GMP) block the NAT decrease by light, whereas high potassium or EGTA do not block this light-induced NAT inactivation.