Serine proteases selectively control the output of 18-hydroxycorticosterone and aldosterone in stimulated zona glomerulosa tissue of the rat adrenal

The finding that incubation of rat adrenal capsules (largely zona glomerulosa) with trypsin reproducibly releases aldosterone and 18-hydroxycorticosterone (18-OH-B) from tightly protein-bound tissue stores leads to the hypothesis that the secretion of these steroids may be under the control of endogenous proteases. Rat adrenal capsule whole tissue and collagenase-dispersed cells were incubated under conditions of stimulation by (1–24)ACTH (10−7 mol/l), potassium (8·4 × 10−3 mol/l) or dibutyryl cyclic AMP (dbcAMP) (10−4 mol/l) with the addition in some flasks of one of the following protease inhibitors at the appropriate concentration for their known actions: Nα-p-tosyl-l-arginine methyl ester (TAME; 10−2 mol/l), 2-nitro-4-carboxyphenyl-N,N′-diphenylcarbamate (NCDC; 2×10−6 mol/l), Nα-benzoyl-l-arginine (BA; 10−2 mol/l), p-nitrophenyl-Nα-benzyloxycarbonyl-l-lysinate (CBZ-NL; 2×10−6 mol/l) and soybean trypsin inhibitor (STI; 1 mg/ml). The (1–24)ACTH-stimulated steroid output in dispersed cells was not affected by NCDC, BA or CBZ-NL. However, all of the inhibitors except STI produced selective effects on aldosterone and 18-OH-B production by whole capsule tissue under certain conditions. Thus TAME and NCDC significantly inhibited the dbcAMP-stimulated production of these two steroids (aldosterone values decreased from 328±35 to 128±15 and 157±32 ng/gland pair respectively) and furthermore NCDC elicited the same effect in potassium- or ACTH-stimulated whole tissue (e.g. in K+-stimulated tissue aldosterone decreased from 79±15 to 40±7 ng/gland pair). The reverse effect was shown by BA and CBZ-NL in potassium-stimulated whole tissue, and yields of aldosterone and 18-OH-B were significantly enhanced (aldosterone increased from 79±15 ng/pair to 151±14 ng in the presence of BA). The high molecular weight inhibitor STI had no effect on potassium-stimulated whole tissue, but enhanced the yield of extractable aldosterone from 9·±1·7 to 16·9±2·3 ng/pair when added to incubations of a cytosol preparation. The results suggest that endogenous proteases in rat adrenal whole capsule tissue play specific roles in the control of aldosterone and 18-OH-B secretion. Some (type 1) whose action is mimicked by trypsin, are inhibited by TAME and NCDC and appear to be involved in the release of these two steroids from their tight (apparently covalent) binding to protein. Others (type 2) are inhibited by BA, CBZ-NL and STI, and one interpretation of their function is that they are concerned with the activation of type 1 proteases. The mechanisms envisaged provide a selective means for control of secretion of the late-pathway steroid products in the rat adrenal zona glomerulosa. These mechanisms are not apparent in dispersed cell incubations, probably because of the loss of steroid–protein complexes in these preparations.