20-HYDROXYEICOSATETRAENOIC ACID MEDIATES CALCIUM CALMODULIN-DEPENDENTPROTEIN-KINASE II-INDUCED MITOGEN-ACTIVATED PROTEIN-KINASE ACTIVATIONIN VASCULAR SMOOTH-MUSCLE CELLS/

Norepinephrine (NE) and angiotensin II (Ang II), by promoting extracel lular Ca2+ influx, increase Ca2+/calmodulin-dependent kinase II (CaMKI I) activity, leading to activation of mitogen-activated protein kinase (MAPK) and cytosolic phospholipase A(2) (cPLA(2)), resulting in relea se of arachidonic acid (AA) for prostacyclin synthesis in rabbit vascu lar smooth muscle cells. However, the mechanism by which CaMKII activa tes MAPK is unclear. The present study was conducted to determine the contribution of AA and its metabolites as possible mediators of CaMKII -induced MARK activation by NE, Ang II, and epidermal growth factor (E GF) in vascular smooth muscle cells. NE-, Ang II-, and EGF-stimulated MAPK and cPLA(2) were reduced by inhibitors of cytochrome P450 (CYP450 ) and lipoxygenase but not by cyclooxygenase. NE-, Ang II-, and EGF-in duced increases in Ras activity, measured by its translocation to plas ma membrane, were abolished by CTP450, lipoxygenase, and farnesyltrans ferase inhibitors. An AA metabolite of CYP450, 20-hydroxyeicosatetraen oic acid (20-HETE),increased the activities of MAPK and cPLA(2) and ca used translocation of Ras. These data suggest that activation of MAPK by NE, Ang II, and EGF is mediated by a signaling mechanism involving 20-HETE, which is generated by stimulation of cPLA(2) by CaMKII. Activ ation of Ras/MAPK by 20-HETE amplifies cPLA(2) activity and releases a dditional AA by a positive feedback mechanism. This mechanism of Ras/M APK activation by 20 HETE may play a central role in the regulation of other cellular signaling molecules involved in cell proliferation and growth.