This study examines ACTH-like immunoreactivity in the pituitary pars distalis and pars intermedia of the freshwater teleost Oreochromis mossambicus (tilapia). Two products (tACTHA and tACTHB) were present in both lobes. These two products also accounted for the majority of the ACTH i.r. when in vitro pars distalis incubation medium was analyzed by HPLC. In a homologous bioassay the two tilapia ACTH-like molecules and human ACTH1-39 possessed similar corticotropic potency. The peptides were quantified using a newly validated radioimmunoassay, which was also used to measure ACTH in plasma of unstressed and stressed fish. Short-term (< 12 min) stress rapidly increased plasma cortisol, reaching levels of around 300 ng/ml in 5 min. Surprisingly, this initial elevation was not accompanied by a rise in plasma ACTH levels. A more prolonged (3 hr) confinement in pairs resulted in high plasma cortisol and ACTH levels in one fish of every pair. The second fish had control ACTH levels and only marginally elevated cortisol levels. Therefore, in this species social interactions seem to influence the reaction of the pituitary-interrenal axis to stress. The short-term cortisol response to disturbance could be abolished completely by pretreating fish in vivo with cortisol for 48 hr. This treatment did not alter circulating ACTH levels. It is concluded that tilapia did not rely on circulating ACTH for a rapid elevation of plasma cortisol levels. Both neuronal mechanisms and cortisol feedback may regulate the pituitary-interrenal axis at the level of the interrenal.