Role of Rho kinase in Akt phosphorylation induced by combined G_12/13 and G_i/G_z signaling in platelets. Washed platelets preincubated with 1 μM AR-C69931MX, 25 μM LY294002, or 10 μM Y-27632 were stimulated at 37°C for 5 minutes with either YFLLRNP (60 μM) + 2-MeSADP (1 μM) or YFLLRNP (60 μM) + epinephrine (10 μM). Platelets stimulated with 2-MeSADP were preincubated with 10 μM MRS-2179. The reaction was stopped by the addition of 3 × SDS sample buffer. Equal amounts of proteins were analyzed by Western blot analysis with anti-Ser(P)⁴⁷³ antibody. The Western blot analysis shown is a representative of 3 independent experiments.