G_12/13 signaling potentiates Akt phosphorylation mediated by G_i or G_z signaling in human platelets. (A) Aspirin-treated, washed human platelets preincubated with 10 μM Y-27632 were stimulated with 60 μM YFLLRNP, and platelet aggregation was measured as described in “Materials and methods.” Arrows indicate the addition of agonist. (B) Washed platelets were stimulated at 37°C for 5 minutes with YFLLRNP (60 μM), 2-MeSADP (1 μM), or epinephrine (Epi; 10 μM). Platelets stimulated with 2-MeSADP were preincubated with 10 μM MRS-2179. The addition of YFLLRNP (60 μM) was made as indicated. The reaction was stopped by the addition of 3 × SDS sample buffer. Samples were separated by SDS-PAGE, Western blotted, and probed with anti-phospho-Akt (Ser⁴⁷³) antibody. The data shown are representative of 3 experiments.