Aldosterone increases T-type calcium currents in human adrenocarcinoma (H295R) cells by inducing channel expression

In adrenal glomerulosa cells, low-threshold voltage-activated (T-type) calcium channels are known to play a crucial role in coupling physiological variations of extracellular potassium to aldosterone biosynthesis. On the other hand, aldosterone itself has been recently shown to regulate Ca(2+) currents in its target cells. In the present study, we have investigated the effect of aldosterone on Ca(2+) channels of the steroidogenic human adrenocarcinoma cell line, using both electrophysiological and molecular techniques. Cell incubation with aldosterone (1 microM) for 24 h increased by 39% the density of T-type calcium currents, as assessed with the patch clamp technique. This effect of aldosterone was not related to a modification of T channel activation and inactivation properties. In contrast, L-type calcium currents remained unaffected by aldosterone treatment. The mineralocorticoid receptor antagonist, spironolactone, blunted the aldosterone-induced increase in T-type calcium current. By RT-PCR, we detected in human adrenocarcinoma cells the presence of mRNA coding for the alpha(1) subunits of three different calcium channels: the alpha(1)H isoform of T channels and the alpha(1)C and alpha(1)D isoforms of the L channels. The presence of mRNA coding for the mineralocorticoid receptor was also found in these cells. Aldosterone treatment induced a 36% increase of mRNA coding for alpha(1)H, as assessed by real-time PCR. This aldosterone-evoked stimulation of mRNA expression was maximal at 24-48 h and reversed by spironolactone, suggesting a receptor-mediated genomic effect of aldosterone. Pregnenolone production in response to KCl stimulation was increased after aldosterone treatment, in parallel to T channel expression, confirming the essential role of these channels in the steroidogenic response to potassium. Taken together, these data indicate that, in human adrenocarcinoma cells, aldosterone increases, through an autocrine pathway, the expression of T-type calcium channels and therefore modifies the ability of these cells to respond to steroidogenic agonists.