THE CONVERSION OF CORTICOSTERONE TO ALDOSTERONE IS THE SITE OF THE OXYGEN SENSITIVITY OF THE BOVINE ADRENAL ZONA GLOMERULOSA

The dissociation of renin and aldosterone observed during hypoxia in vivo has been attributed to a direct inhibition of low oxygen on adrenal zona glomerulosa function. We have demonstrated that the adrenal zona glomerulosa production of aldosterone in vitro is directly proportional to a wide range of oxygen concentrations in the physiological range but that cortisol production from coincubated fasciculata cells is not oxygen sensitive. The present study examined the hypothesis that the sensitivity to O2 is limited to the aldosteronogenic late pathway. In order to localize the site of oxygen sensitivity, we measured endogenous pregnenolone production (early pathway) and the conversion of exogenous corticosterone to aldosterone (ALDO) (i.e. 18-hydroxylase activity) in adrenal cells treated with cyanoketone (3-beta-hydroxy-steroid dehydrogenase inhibitor). Acutely dispersed bovine adrenal glomerulosa cells (four experiments in pentuplicate) were incubated under low (5%) vs. normal (21%) O2 in the presence of cyanoketone (CK; 1-mu-M) and/or the following: corticosterone (500 ng/ml), angiotensin II (ANG II; 10 nM), or dibutyryl cAMP (1 mM). Conversion of exogenous corticosterone to ALDO in the presence of CK was inhibited by 41 +/- 1% under low O2. This was similar to the inhibitory effect of low O2 on ANG II-stimulated aldosterone production from endogenous precursors in the absence of CK (52 +/- 11% inhibition). Basal, ANG II-, and cAMP-stimulated endogenous pregnenolone production was not significantly reduced by low O2. In another experiment, glomerulosa cells were incubated under 5, 13, or 50% vs. 21% O2 in the presence of CK (1-mu-M) and different concentrations of corticosterone (10-100 ng/ml). ALDO production was significantly inhibited by low O2 when corticosterone was greater-than-or-equal-to 500 ng/ml and ALDO was significantly augmented by high O2 when added corticosterone was 1000 ng/ml. We conclude that the conversion of corticosterone to ALDO (i.e. 18-hydroxylase) appears to be the primary site of oxygen sensitivity since 1) pregnenolone production was unaffected and 2) the magnitude of the inhibition of the conversion of corticosterone to ALDO by low O2 in the presence of CK was similar to the inhibition of ALDO production from endogenous precursors in the absence of CK. These studies demonstrate that oxygen sensitivity of the steroidogenic pathway is a unique, constitutive property of 18-hydroxylase, the enzyme which catalyzes the conversion of corticosterone to ALDO. We propose that the sensitivity of 18-hydroxylase to oxygen accounts for the dissociation of renin and aldosterone during hypoxia in vivo.