Dissociation between calcium uptake and release and calcium effect on steroid biosynthesis in isolated mitochondria from adrenal zona fasciculata

Adrenal cortex cells are activated by ACTtt to generate corticoids via the cAMP signal transduction cascade. Calcium has been postulated to regulate the rate-limting step that is located at the mitochondria, tlowev(,r, studies are lacking exploring a direct relationship of mitochondrial (a+ levels and steroid synthesis. We study the nptake of Ca²⁺ on isolated rat adrenal mitochondria using the indicator Fura-2 and its effect on steroid synthesis. Our results show that the inlramitochondrial calcium concentration can be controlled by varying the extramitochondrial concentration of the ion. Increments on calcium concentration failed to stimulate steroid production on calciumdepleted mitochondria (8.8 ± 0.9 vs 10.1 ± 0.7 ng/incubation [ir (1 and 500 nM Ca²⁺ respectively). Ca2+-depleting conditions reduced the stimulation of progesterone synthesis achieved by cAMP-dependent factors (60.2 ± 5.3 vs 25.0 ± 2.5 ng/incubation for control and 0 Ca2+ respectively), an effect that could not be overcome by readdition of Ca2+.Ca2+-depletion had no effect on steroid production when the rate-limiting step was by-passed by incubation of the mitochondria with a cholesterol permeable analogue or when the ratelimiting step was stimulated by in vivo treatment with ACTII. Our results suggest then that calcium per se is not capable to stimulate the rare-limiting step in steroidogenesis and indicate a regulatory role for calcium in the control of hormone, induced steroid synthesis at the level of cholesterol transport.