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1. The role of 5-hydroxytryptamine (5-HT) in temperature regulation and in fever in the rabbit has been investigated. 2. Intrahypothalamic microinjections of 5-HT in the conscious rabbit alters body temperature in a dose-dependent manner. 3. Low doses (5-5nmol) of 5-Ht and control saline injections produced a small, non-significant increase in temperature, with a long latency. 4. Doses of 14 nmol 5-HT produce a hyperthermia with a 45 min delay; while microinjections of 28 nmol result in a biphasic response; an initial short hypothermia is followed later by a hyperthermia. 5. Depleting the rabbit's brain of 5-HT by pretreatment with p-chlorophenylalanine (PCPA) fails to affect its body temperature at thermoneutral temperatures but significantly impairs the ability to thermoregulate against a cold stress. 6. PCPA pretreatment did not, however, impair the febrile response to bacterial pyrogen and prostaglandin E1. 7. These results reveal a dissociation between the effects of 5-HT depletion on temperature regulation, and on fever. The site of action of 5-HT in temperature regulation must be proximal to the fever input, but distal to the convengence of peripheral and hypothalamic temperature inputs.