The long-term goal of the proposed study is to understand the long-term effect of angiotensin II and the endogenous renin-angiotensin system (ERAS) in relation to the mechanism of regulation of the neurotransmitters release and the gene expression of adrenal cell proteins (proenkephalin, tyrosine hydroxylase, and PNMT). The specific goal of this project was to determine if the release of the neurotransmitters (catecholamines and met-enkephalin) and gene expression by long-term exposure to angiotensin II (AgII) is mediated by protein kinase C (PKC) or arachidontic acid (AA) and its metabolite (eg. PGE2). We have used primary cultured bovine adrenal chromaffin cells. We have found that AgII, AA and PGE2 increased the secretion of met-enkephalin (ME) after continuous stimulation for 12 hrs. The secretion of ME by AgII, AA and PGE2 was completely inhibited by staurosporin but not by K252a, indicating that PKC plays an important role in mediating the secretion of ME. The action of staurosporin seems to be selective since saturosporin did not affect the secretion induced by potassium and veratridine. We also found that the secretion of ME induced by AgII and AA was not inhibited by indomethacin (a cycloxygenase inhibitor) and caffeic acid (a lipooxygenase inhibitor), indicating that secretion of Me induced by AGII and AA is not mediated by PGE2. Currently, we are trying to determine if the secretion of ME induced by AgII is mediated by AA, using PLA2 inhibitor. We are also studying the effects of saturosporin, K252a, indomethacin and caffeic acid on the gene expression of proenkephalin induced by AgII, AA or PGE2. This study will delineate the mechanism underlying the regulation of adrenal medullary function of sympathetic neurotransmission. This work should provide rational means for intervening in cardiovascular diseases in which blood pressure and heart rate may be modified by AgII.
