Day-night differences in the concentrations of melatonin and serotonin (5HT) were measured in several regions of the chicken brain, pineal gland and serum. Melatonin concentrations are higher at midnight than at midday in 8 of the 10 tissues studied although the amplitudes of these rhythms varied greatly. Day-night differences in the pineal, hypothalamus, thalamus, retina and pons-midbrain regions had the highest amplitudes. 5HT concentrations were rhythmic in only 3 of the tissues studied: the hypothalamus, thalamus and retina. These were also the areas of highest 5HT concentration. Exogenous melatonin, injected at midday, was taken up with similar patterns; the pineal, hypothalamus, thalamus and pons-midbrain contained more melatonin 20 min after injection than did other tissues. The rate of decline of melatonin concentration varied little among all tissues studied, suggesting that the differences among tissue concentrations were due to selective uptake mechanisms rather than specialized degradation pathways. The effects of exogenous melatonin on 5HT concentration were restricted to hypothalamus, thalamus, pons-midbrain, retina and pineal. No effect was seen in cerebellum, optic tectum, neostriatum, hippocampus and medulla oblongata. Together, these data strongly suggest that pineal (and exogenous) melatonin is selectively taken up primarily by three brain regions, hypothalamus, thalamus and pons-midbrain, in which it produces increases in 5HT concentrations. Regional selectivity of uptake may be the mechanism by means of which the effects of melatonin on 5HT-mediated function are restricted to specific brain areas.