Inhibin, a member of the transforming growth factor (TGF)- superfamily, has been proposed to act as an inhibitor of activin and BMP by sequestering their type II receptors in non-signaling complexes with betaglycan. This mechanism of inhibin action was tested in a mouse adrenocortical (AC) cell line by examining the effects of inhibins A and B on cytochrome P450 17-hydroxylase 17,20-lyase (Cyp17) expression and 17-hydroxylase activity, measured by progesterone 17-hydroxylation, in the absence and presence of activin or BMP isoforms. Cyp17 mRNA endogenously expressed by AC cells was suppressed by activins A and B and BMP-2, -6 and -7, and each ligand accordingly inhibited 17-hydroxyprogesterone production (IC50 of 0.24, 0.27, 0.4, 0.51 and 2.2 nM, respectively). Neither inhibin A nor inhibin B alone affected Cyp17 expression or 17-hydroxyprogesterone production. Both inhibin A and inhibin B blocked the inhibitory actions of activins A and B in AC cells, supporting the anti-activin model of inhibin action. Inhibin A provided more potent and effective antagonism of both activins than did inhibin B, and activin A was less subject to antagonism by either inhibin than was activin B. In contrast to the major antagonism of activin by both inhibins, only inhibin A antagonized the actions of BMP-2, BMP-6 and BMP-7, whereas inhibin B was ineffective against all tested BMP isoforms except BMP-7 at high concentrations. These results provide limited support for the anti-BMP model of inhibin action and reveal that, relative to inhibin A, inhibin B essentially behaves as a selective activin antagonist in AC cells. In conclusion, inhibins A and B differentially antagonize the actions of activins and BMPs to control adrenocortical C₁₉ steroid production.