Complementary roles of JNK1 and ERK2 in platelet aggregation and TXA₂ synthesis. The levels of ERK2 expression (A) and phosphorylation were evaluated in WT and JNK1^−/− platelets after activation by collagen (1 μg/mL; B) or thrombin (65 mU/mL; C) for 3 minutes, with stirring. Stimulated platelets were lysed and samples analyzed by Western blotting with appropriate antibodies directed against total ERK2 (A) or phosphorylated ERK2 (B-C). Tubulin was used as a loading control. The Western blot shown is representative of at least 3 independent experiments. The complementary roles of JNK1 and ERK2 were evaluated for platelet aggregation (D-E) and for TXA₂ synthesis (measured by evaluating production of TXB₂; F-G), in WT or JNK1^−/− platelets (10⁸) stimulated with collagen (1 μg/mL) or thrombin (65 mU/mL) for 3 minutes, with stirring. Platelets were first incubated for 10 minutes at 37°C with DMSO (0.1%, vol/vol), as a control, or the MEK inhibitor U0126 (10μM) plus or minus indomethacin (5μM). Thromboxane synthesis was determined in the supernatants of aggregated platelets, as described in “Methods.” Each bar represents the mean plus or minus SEM of 3 experiments. **P < .01, ***P < .001 (unpaired Student t test). Traces are representative of at least 3 independent experiments.