1. Either electrical stimulation of midbrain raphe nuclei or administration of 5-hydroxytryptamine (5-HT; serotonin) into the preoptic anterior hypothalamus caused hypothermia in conscious rats at ambient temperatures (Ta) of both 8 degrees C and 22 degrees C. The hypothermia was due to decreased metabolic heat production at Ta = 8 degrees C, while at Ta = 22 degrees C the hypothermia was due to both decreased metabolism and increased heat loss (cutaneous vasodilatation). However, at Ta = 30 degrees C, electrical stimulation of midbrain raphe or intrahypothalamic injection of 5-HT caused an insignificant change in the thermoregulatory responses. There was no changes in respiratory evaporative heat loss in response to these treatments at various Ta's. 2. Direct administration of the serotonergic receptor antagonists such as cyproheptadine and methysergide into the preoptic anterior hypothalamus caused hyperthermia in conscious rats at Ta's of 8 degrees C, 22 degrees C and 30 degrees C. The hyperthermia was due to increased metabolism and cutaneous vasoconstriction. 3. The hypothermia induced by intrahypothalamic administration of 5-HT was antagonized by pretreatment with an intrahypothalamic dose of either cyproheptadine or methysergide in rats at Ta = 22 degrees C. 4. Inhibition of 5-HT neuronal activity with administration of 5-HT into the midbrain raphe regions also caused hyperthermia, increased metabolism and cutaneous vasoconstriction in rats at Ta's of 8 degrees C, 22 degrees C and 30 degrees C. 5. These observations tend to suggest that the functional activity of serotonergic receptors in the preoptic anterior hypothalamus mediates thermoregulatory responses in the rat. Activation of serotonergic receptors in the hypothalamus decreases heat production and/or increases heat loss, while inhibition of serotonergic receptors in the hypothalamus increases heat production and/or decreases heat loss in the rat.