Enucleated and enucleated+Harderianectomized (E+H) 12-day-old rats were exposed to an additional 5 h of light or placed in the dark at 17.00. Body temperatures were lower in both groups in the dark in comparison to rats with comparable treatments in the light. These results suggest that removal of the Harderian gland does not alter the animal’s response to environmental temperature. Intact, suckling animals were placed in the dark at 7, 22, and 34°C for 4 h or left with their mother. Pineal N-acetyltransferase was significantly higher in the group at 22 °C than in any other group, suggesting that the activity of this enzyme is influenced by environmental temperature in suckling rats. Pineal serotonin (5-HT) shows little change under these conditions. However, if rats at these temperatures also are exposed to overhead lighting, dramatic increases occur in pineal 5-HT at both 22 and 34 °C. N-acetyltransferase levels are significantly lowered at 34°C only; pineal HIOMT does not change. These findings are compatible with the hypothesis that light and temperature both regulate pineal function in the suckling rat, probably by different mechanisms. The diurnal variation in pineal 5-HT in 12-day-old E+H rats is abolished by constant light for 3 days but not by constant dark. This pattern is similar to that found in intact adults in constant light or constant dark.