Subcellular compartmentation of steroidogenesis in the zona fasciculata/reticularis of the rat adrenal cortex

The secretion of corticosterone and 18-hydroxydeoxycorticosterone (18-OH-DOC) by fasciculata/reticularis tissue or cells from the rat adrenal cortex is markedly dissociated under certain conditions. For this reason, it has been suggested that the two steroids may be synthesised in discrete subcellular compartments. Using subcellular fractions obtained by centrifugation, and with measurement of steroids by g.l.c., compartmentation of steroidogenesis in this tissue has been reinvestigated by measuring steroids produced under different incubation conditions from endogenous precursors and from added progesterone and deoxycorticosterone (DOC). In accordance with the literature, major sites for 18 and 11β-hydroxylation are mitochondrial and 21-hydroxylation is largely microsomal. However, all three activities also occur in other sites throughout the cell. Although there is some cross contamination of fractions, steroid profiles are sufficiently distinct in different compartments to suggest different roles in the elaboration of the secreted product. Three sites for steroidogenesis may be distinguished. In the first, apparently purely intramitochondrial, 18-OH-DOC is produced in amounts much greater than corticosterone. This site is revealed by incubation of mitochondria with isocitrate, but in the absence of NADP + or NADPH, and thus steroidogenesis, from endogenous precursors, relies on intramitochondrial generation of reduced nucleotide alone. Secondly, there is a more superficial site of mitochondrial steroidogenesis in which corticosterone and 18-OH-DOC are produced in similar amounts. This profile is shown in products from both added and endogenous precursors, and in this case steroidogenesis may be supported by added NADPH or isocitrate with NADP +. Finally, purely extramitochondrial steroidogenesis results in production of much more corticosterone than 18-OH-DOC from both added and endogenous precursors. Changes in ratio of secreted corticosterone and 18-OH-DOC under physiological conditions may therefore arise through changes in the intracellular locus of steroidogenesis. In particular, the effect of ACTH stimulation on the steroid profile would be replicated by a switch from the purely intramitochondrial to the superficial mitochondrial site of steroidogenesis. In vivo this might arise through movement of steroid between the sites, or alternatively through increased extramitochondrial generation of reducing equivalents.