INHIBITION OF STEROIDOGENESIS IN RAT ADRENAL-CELLS BY 18-ETHYNYLDEOXYCORTICOSTERONE - EVIDENCE FOR AN ALTERNATIVE PATHWAY OF ALDOSTERONE BIOSYNTHESIS

The effect of the mechanism-based inhibitor 18-ethynyldeoxycorticoster one (18-E-DOC) on the late steps of the aldosterone biosynthetic pathw ay was examined in freshly isolated cells of the zona glomerulosa (ZG) and fasciculata (ZF) from rat adrenal glands. ZG synthesis of aldoste rone was inhibited by 18-E-DOC in a time-and concentration-dependent m anner with a K-i of approximately 0.05 mu M The maximal degree of inhi bition of ZG production of aldosterone and 18-hydroxycorticosterone (1 8-OH-B) was approximately 80%. ZI; cells, perhaps surprisingly, were f ound to secrete 18-OH-B at levels approximately one-third to one-fourt h those of ZG cells and the Ki of 18-E-DOC inhibition of 18-OH-B secre tion was approximately :LO mu M for ZF cells, 200-fold higher than for ZG cells. The inhibitor had no effect on the secretion of corticoster one by either ZG or ZF, and the secretion of 18-hydroxydeoxycorticoste rone (18-OH-DOC) by both the ZG and ZF was inhibited only to a minor d egree. 18-E-DOC inhibited the biosynthesis of aldosterone by ZG cells incubated with 10 mu M added DOC or 18-OH-DOC by approximately 75%, si milar to the degree of inhibition of aldosterone biosynthesis from end ogenous substrate, whereas ZF biosynthesis of 18-OH-B from either subs trate was inhibited by less than 40%. ZI; cells do not express aldoste rone synthase, the only enzyme known to convert 18-OH-DOG into 18-OH-B . Incubation of MA-10 cells stably transfected with the cDNA of the ra t aldosterone synthase with 18-E-DOC resulted in a complete inhibition of the conversion of DOC to aldosterone with a K-i of approximately 0 .02 mu M. In addition, transfected cells expressing 11 beta-hydroxylas e convert DOC to 18-OH-B in very small quantities only and cannot conv ert 18-OH-DOG to 18-OH-B. These data suggest that neither 11 beta-hydr oxylase nor aldosterone synthase are responsible for the biosynthesis of 18-OH-B by ZF cells from DOC or 18-OH-DOC, that 20% of aldosterone synthesis appears not to be attributable to the actions of aldosterone synthase and that an unknown CYP11B enzyme is also involved in the bi osynthesis of 18-OH-B. (C) 1997 Elsevier Science Ltd.