The Rate‐Limiting Step of Adrenal Steroidogenesis and Adenosine 3′:5′‐Monophosphate

Abstract

The rate‐limiting step of adrenal steroidogenesis is first defined in detail. The kinetics of the conversion in vitro of double‐labeled exogenous cholesterol as potential precursor by intact and ruptured bovine adrenocortical mitochondria are compared under identical conditions. The results strongly suggest that the transport of cholesterol from the outside to the inside of the mitochondrial membrane is the proper rate‐limiting process and not the subsequent enzymatic conversion of cholesterol to steroids. The involvement of a transport process is supported by findings such as increase of conversion rates in intact mitochondria by (a) preincubation of exogenous precursor with mitochondria; (b) addition of carrier proteins such as non‐defatted bovine serum albumin as against proteins strongly binding cholesterol; (c) addition of Ca 2+ concentrations sufficient to increase membrane permeability. The immediate stimulation by adrenocorticotrophic hormone of adrenocortical steroidogenesis seems to be mediated by cyclic AMP formed intracellularly. Such a messenger may be considered to act directly or indirectly upon the rate‐limiting step. It was found that the presence of cyclic AMP (1–5 mM) or cyclic monobutyryl AMP (0.04–1 mM) under optimal and various suboptimal conditions of cholesterol utilization did in no instance increase the conversion rate nor did it influence the utilization of endogenous precursor in intact mitochondria. It is concluded that cyclic AMP has no direct effect on (a) the transport process for exogenous cholesterol; (b) the intramitochondrial enzymatic conversion; (c) the utilization of endogenous steroid precursor. If cyclic AMP is involved, its action seems to have a premitochondrial localization and to be related indirectly only to a transport mechanism or a precursor supply process.