The serotonin receptors involved in the secretion of adrenocorticotropin hormone (ACTH) were investigated in conscious adult male rats. Administration of serotonin (5-HT), 5-hydroxytryptophan (5-HTP) in combination with the serotonin reuptake inhibitor fluoxetine (Flx), or of the 5-HT agonists 8-OH-DPAT (5-HT1A), 5-carboxamido-tryptamine (5-HT1A+1B+5A+7), RU 24969 (5-HT1B+1A), DOI (5-HT2A+2c), S-alpha-methyl-5-HT (5-HT2A+2B+2c), MK212 (5-HT2B+2c), or methyl-chlorophenyl-piperazine (5-HT2A+2c) dose-dependently stimulated ACTH secretion. The 5-HT3 agonist 2-methyl-5-HT had no effect. Administration of a 5-HT1 agonist in combination with any of the 5-HT2 agonists DOI, S-alpha-methyl-5-HT or MK212 had an additive effect on the plasma concentration of ACTH. The ACTH stimulating effect of each of the 5-HT agonists was inhibited by pretreatment with antagonists with corresponding 5-HT receptor affinity. The ACTH response to 5-HT or 5-HTP/Flx was inhibited by injection with the 5-HT1A+2A+2c+5A+7 antagonist methysergide, the 5-HT2A antagonist ketanserine and the 5-HT2C+2A antagonist LY 53857. The 5-HT1A antagonist WAY 100635 enhanced 5-HT- and 5-HTP/Flx-induced ACTH secretion, suggesting a presynaptic 5-HT1A autoreceptor effect of the drug. The 5-HT3 antagonist ondansetrone had no effect on the either of the 5-HT agonists. The 5-HT3+4 antagonist tropisetrone attenuated the effect of 5-HTP/Flx, which may suggest a stimulation of ACTH secretion via 5-HT4 receptors. It is concluded that 5-HT1A, 5-HT2A+2C, and to a lesser extent 5-HT1B receptors, but not 5-HT3 receptors are involved in the effects of serotonin agonists on ACTH secretion. Furthermore, an involvement of the 5-HT5A and the 5-HT7 receptor is possible.