Hypothalamic serotonin (5-HT) is believed to have an inhibitory effect on food intake in a variety of species. To define more precisely the nature of this effect, this study investigated the effects of medial hypothalamic 5-HT injection on natural patterns of macronutrient intake in freely feeding rats. Serotonin (5-20 nmol) was injected directly into the paraventricular nucleus (PVN) of brain-cannulated rats maintained ad libitum on pure macronutrient diets, protein, carbohydrate and fat, and measurements of nutrient intake were taken one hour later. To assess whether the action of 5-HT on macronutrient intake varies across the light-dark cycle, these tests were conducted at 3 different times in the nocturnal feeding period, during hours 1, 6 and 11 after lights out. The results demonstrate that the suppressive effect of PVN 5-HT on food intake is dose dependent, nutrient selective, as well as time dependent. Specifically, PVN injection of 5-HT, at all doses tested, was effective at only one time of the nocturnal cycle, namely, at the onset of the active, dark period. While no behavioral effect of 5-HT was detected in the middle and late phases of the dark, a strong, dose-dependent reduction of nutrient intake was revealed immediately after lights out. This suppressive effect was characterized by a highly selective decrease in carbohydrate intake, along with a significant enhancement in preference for protein, as well as for fat, and little change in total caloric intake.(ABSTRACT TRUNCATED AT 250 WORDS)