Angiotensin II-activated protein kinase D mediates acute aldosterone secretion.

Dysregulation of the renin-angiotensin II (AngII)-aldosterone system can contribute to cardiovascular disease, such that an understanding of this system is critical. Diacylglycerol-sensitive serine/threonine protein kinase D (PKD) is activated by AngII in several systems, including the human adrenocortical carcinoma cell line NCI H295R, where this enzyme enhances chronic (24h) AngII-evoked aldosterone secretion. However, the role of PKD in acute AngII-elicited aldosterone secretion has not been previously examined. In primary cultures of bovine adrenal glomerulosa cells, which secrete detectable quantities of aldosterone in response to secretagogues within minutes, PKD was activated in response to AngII, but not an elevated potassium concentration or adrenocorticotrophic hormone. This activation was time- and dose-dependent and occurred through the AT1, but not the AT2, receptor. Adenovirus-mediated overexpression of constitutively active PKD resulted in enhanced AngII-induced aldosterone secretion; whereas overexpression of a dominant-negative PKD construct decreased AngII-stimulated aldosterone secretion. Thus, we demonstrate for the first time that PKD mediates acute AngII-induced aldosterone secretion.

The role of calcium influx pathways in phospholipase D activation in bovine adrenal glomerulosa cells.

The steroid hormone aldosterone maintains sodium homeostasis and is therefore important in the control of blood volume and pressure. Angiotensin II (AngII) and elevated extracellular potassium concentrations ([K(+)](e)), the prime physiologic regulators of aldosterone secretion from adrenal glomerulosa cells, activate phospholipase D (PLD) in these cells. The role of Ca(2+) in the activation by these agents is unknown, although nitrendipine, a voltage-dependent Ca(2+) channel antagonist, does not inhibit AngII-elicited PLD activation, despite the fact that this compound blocked elevated [K(+)](e)-stimulated PLD activity. PLD activation triggered by AngII was also unaffected by the T-type calcium channel inhibitor nickel. Nevertheless, Ca(2+) influx was required for AngII-induced PLD activation in both primary cultures of bovine adrenal glomerulosa cells and a glomerulosa cell model, the NCI H295R adrenocortical carcinoma cell line. The involvement of store-operated Ca(2+) (SOC) influx and Ca(2+) release-activated Ca(2+) (CRAC) influx pathways in PLD activation was investigated using thapsigargin, an endoplasmic reticulum Ca(2+) pump inhibitor that empties the store to induce SOC influx, and the SOC inhibitor YM-58483 (BTP2), as well as a CRAC inhibitor, tyrphostin A9. In bovine glomerulosa cells, tyrphostin A9 inhibited AngII-induced PLD activation without affecting elevated [K(+)](e)-stimulated enzyme activity. On the other hand, differences were observed between the bovine adrenal glomerulosa and H295R cells in the involvement of Ca(2+) influx pathways in PLD activation, with the involvement of the SOC pathway suggested in the H295R cells. In summary, our results indicate that Ca(2+) entry only through certain Ca(2+) influx pathways is linked to PLD activation.