A 2-CELL TYPE THEORY FOR ALDOSTERONE BIOSYNTHESIS - THE ROLES OF 11-BETA-HYDROXYLASE AND ALDOSTERONE SYNTHASE, AND A HIGH-CAPACITY TIGHTLY BINDING STEROID CARRIER FOR 18-HYDROXYDEOXYCORTICOSTERONE IN RAT ADRENALS

Several lines of experimentation suggest that a tissue-sequestered poo l of 18-hydroxydeoxycorticosterone (18-OH-DOC) in the rat adrenal may be mobilized as an aldosterone precursor. We show here that this stero id is maintained in a non-extractable form in the membranes of collage nase-dispersed fasciculata/reticularis cells. Because of this stabilit y, the complex can be identified by immunocytochemistry and also, in I EF gels of solubilized inner adrenocortical zone membrane preparations , by immunoblotting. However, the complexed steroid cannot be extracte d from the gels into organic solvent unless first treated with trypsin . Preincubation of viable whole glandular tissue with trypsin signific antly enhanced aldosterone output and eliminated the trypsin-releasabl e 18-OH-DOC pool in IEF gels of solubilized inner zone membranes. Both prior sodium depletion and acute trypsin stimulation of whole glands enhanced extractable 18-OH-DOC in glomerulosa tissue membranes. Other experiments using in situ hybridization show that mRNA coding for 11 b eta-hydroxylase (which generates 18-OH-DOC) is confined to the inner a drenocortical zones, whereas aldosterone synthase (which does not) is transcribed exclusively in the glomerulosa. The data suggest that a po ol of 18-OH-DOC in inner zone membranes can be mobilized for utilizati on as an aldosterone precursor in the glomerulosa. The results also in dicate the existence of an entirely novel tightly binding steroid carr ier from which steroid cannot be extracted by organic solvent unless f irst subjected to proteolytic degradation.