Hebert, Richard L.,Lal, Mark A.2009-03-192009-03-1919971997Source: Masters Abstracts International, Volume: 36-06, page: 1533.9780612284333http://hdl.handle.net/10393/4489http://dx.doi.org/10.20381/ruor-13883The mammalian kidney, in particular the collecting duct segment of the nephron, is a major site of renal sodium and water reabsorption. One of the many agonists known to regulate collecting duct function is bradykinin (BK), a nonapeptide which exerts natriuretic and diuretic effects through its ability to increase arachidonic acid (AA) availability and prostaglandin production. Evidence presented herein indicates that 85 kDa cytosolic phospholipase A$\sb2$ (cPLA$\sb2)$ is the enzyme accountable for BK-mediated AA. This is based on the following observations: RCCD cells express cPLA$\sb2$ protein, an arachidonyl trifluoromethyl ketone inhibitor of cPLA$\sb2$ completely blunted BK-stimulated AA release, both basal and BK-stimulated PLA$\sb2$ activities were significantly diminished following treatment with an antibody to this enzyme, and finally, PLA$\sb2$ in vitro activity could be stimulated by submillimolar calcium concentrations and was dithiothreitol (DTT)-insensitive. Strategies were also employed to reveal the involvement of protein kinase C (PKC) in BK-mediated AA release, cPLA$\sb2$ activation and cPLA$\sb2$ phosphorylation. Chemical inhibition of PKC reduced both BK-stimulated AA release and cPLA$\sb2$ activity; however, BK-stimulated AA release appeared limited by calcium mobilization rather than by PKC. Cytosolic PLA$\sb2$ activity could be stimulated by phorbol ester (PMA) and the magnitude of this activation was similar to that caused by BK. A direct correlation between the level of cPLA$\sb2$ phosphorylation and activation was confirmed by the phosphatase-mediated reversal of both BK- and PMA-stimulated responses. Furthermore, the increase in cPLA$\sb2$ phosphorylation was found to occur on serine residues. (Abstract shortened by UMI.)115 p.Biology, Cell.Thesis