Effects of proteolytic enzymes on steroid release from rat adrenal zona glomerulosa tissue: Evidence for novel steroid-protein complexes

Trypsin (2mg/ml) added to conventional incubations of rat adrenal capsules (largely glomerulosa) reproducibly increases the amount of free extractable aldosterone (aldo) and 18-hydroxycorticosterone (18-OH-B) in incubation media, but has no effect on capsule cell suspensions formed by collagenase incubation. The effect is abolished by the addition of a trypsin inhibitor, but is still seen in the absence of $\text{de novo}$ steroidogenesis. Qualitatively similar results were obtained with capsule homogenates and high speed supernatant fractions, and chromatography of the high speed supernatant protein fraction on Sephadex G-50 gave a number of minor fractions and one major fraction which yielded free aldo on incubation with trypsin. The results indicate the existence of storage forms of aldo and 18-OH-B which are extremely tightly bound to protein. Such steroid-protein complexes appear to be of an entirely novel kind, and are quite distinct from the familiar receptor type complexes. The findings support previously proposed mechanisms for aldo synthesis and secretion.