Figure 3.

Effects of chronic ET-1 treatment on insulin-stimulated PI3-kinase activity. Serum-starved 3T3-L1 adipocytes were incubated in the absence or presence of insulin (I; 100 ng/ml) or PDGF (P; 50 ng/ml) for 10 minutes, after treatment with or without ET-1 (10 nM) for 24 hours. Cell lysates were immunoprecipitated with anti-p110α antibody, and the immunecomplexes were assayed for their ability to phosphorylate phosphatidylinositol. Reaction products (PI3P) were analyzed by thin layer chromatography, and signals were quantitated on a PhosphorImager as described in Methods. A representative experiment is shown in a, and data shown in the bar graph in b are mean ± SE from three independent experiments.