The study examined the affect of cortisol on basal and cAMP-stimulated in vitro steroidogenesis of rainbow trout (Oncorhynchus mykiss) ovarian follicles at different maturational stages. Basal and cAMP-stimulated 17?-estradiol (E2) and testosterone (T) production was highest during the mid-vitellogenic (MV) stage, compared with early- and late-vitellogenic stages. Cortisol suppressed the basal steroid production in the MV stage, and suppressed cAMP-stimulated steroid production in all three maturational stages. To determine whether cortisol exerts its effect on pregnenolone (P5) production, real time RT-PCR was used to measure the expression of genes coding for steroidogenic acute regulatory (StAR) protein, and cytochrome P450scc. There was no significant affect of cortisol on the expression of either gene. To look for possible actions of cortisol on steroidogenic enzyme function, the effect of the glucocorticoid on the biotransformation of radiolabelled P5, and T was examined; cortisol appeared to suppress the aromatization of androgens to estrogens by a mechanism that is not fully understood.