No changes were found in the concentration of tryptophan (Trp), 5-hydroxytryptophan (5-HTP), 5-hydroxytryptamine (5-HT) or 5-hydroxyindole acetic acid (5-HIAA) in the cerebrospinal fluid (CSF) of male rats either before sexual activity, immediately after ejaculation of after the postejaculatory refractory period (PEI). Injection of the Trp hydroxylase inhibitor p-chlorophenylalanine (PCPA, 25 or 100 mg/kg i.p. for 3 days) in combination with an injection of the monoamine oxidase inhibitor pargyline (100 mg/kg i.p.) increased the concentration of Trp while decreasing the concentration of 5-HTP, 5-HT and 5-HIAA in the CSF. Furthermore 100 (but not 25) mg/kg PCPA in combination with pargyline caused a significant reduction in the latency to ejaculation. Injection of probenecid (200 mg/kg i.p.), an inhibitor of the transport of 5-HIAA from the CSF, increased the concentration of 5-HIAA in the CSF and slightly prolonged the latency to ejaculation. Sexual activity caused no further increase in CSF 5-HIAA levels in the probenecid-treated rats. Since drug-induced changes in sexual behavior are associated with marked alterations in 5-HT metabolism in the CSF, whereas the changes in the behavior which occur normally are not, these results question the physiological significance of the proposed inhibitory role of 5-HT in male rat sexual behavior.