Fig. 8. NE-induced translocation of cPLA₂ to the nuclear envelope depends on its phosphorylation and not its activity. VSMCs were treated with inhibitors of cPLA₂ activity arachidonyltrifluoromethyl ketone (AACOCF₃; 10 µM) and methyl arachidonyl fluorophosphonate (MAFP; 10 µM) or their vehicle (VEH). (A) cPLA₂ translocation to the nuclear envelope in cells treated with 10 µM NE or its vehicle (V) was visualized by confocal microscopy (100x magnification). (B) Density of fluorescence around the nuclear envelope was quantified (n=5). ^*Value significantly different from that obtained with V of NE (P<0.05). (C) cPLA₂ activity in VSMCs treated with AACOCF₃, MAFP or VEH in the presence of NE (10 µM) or its vehicle (V) as described in Materials and Methods (n=5). (D) Phosphorylation of cPLA₂ as determined by ³²P incorporation in VSMCs treated with AACOCF₃, MAFP or their vehicle (VEH) and treated with NE (10 µM) or its vehicle (V). The figures shows a representative of three experiments performed with each agent and its vehicle on different batches of cells grown in 100 mm tissue culture dishes. (E) Density of cPLA₂ phosphorylation was quantified using NIH Image 1.62 (densitometric analysis of D).