The disruption of endocrine system function in wildlife species, including teleosts, by contaminants such as metals is presently of major environmental concern. Recently, it has been shown that cadmium (Cd) exposure results in significant reductions in corticosteroid secretion by fish interrenal steroidogenic cells, likely through an inhibition of intracellular cortisol synthesis. In the present study, the effects of CdCl(2) on unstimulated and stimulated interrenal steroidogenesis in rainbow trout were examined with the intention of furthering an understanding of the site(s) of Cd toxic action. CdCl(2) alone reduced cortisol secretion in minced interrenal tissues to 59% and 55% of control values when exposed to 10 and 100 microM, respectively. Incubation of interrenal tissues with 0.01 IU/mL adrenocorticotropic hormone (ACTH), which activates rate-limiting steps in steroid synthesis, resulted in significant stimulation of steroidogenesis in controls. However, ACTH-stimulated steroidogenesis was reduced when tissues were previously incubated with Cd. Maximal rates of unstimulated cortisol secretion were achieved by augmentation using 5 microM 25-hydroxycholesterol (25-OHC) or 0.8 microL/mL synthetic cholesterol [SyntheChol(SC)]. Steroidogenesis augmentation by 25-OHC was significantly reduced in tissues incubated with Cd. Interestingly, cortisol secretion was significantly higher in SC-augmented tissue exposed to 1 and 10 microM Cd when compared to augmented control tissues. The results of this study show that Cd affects both stimulated and unstimulated steroidogenesis in rainbow trout, and that one major site(s) of action of Cd in the cortisol synthesis pathway is likely prior to cytochrome P450 side chain cleavage.