MAP kinase inhibition does not block activation of αIIbβ3 by VWF. (Ai) Structure of the novel MEK inhibitor 5-bromo-2-[2-chloro-4-iodo-phenylamino]-N-cyclopropylmethoxy-3,4-difluoro-benzamide (PD184161). (Aii) Human washed platelets (5 × 10⁸/mL) were preincubated for 5 minutes with vehicle (DMSO 0.1%) or the MEK inhibitor PD184161 at the indicated concentration followed by stimulation with thrombin (0.05 IU/mL). Reactions were stopped by addition of equal volumes of sample buffer, and proteins were separated by SDS-PAGE and Western blotted for phosphorylated p42/44 MAP kinase. Blots are representative of 4 experiments. (Bi) Human platelet-rich plasma (3 × 10⁸ platelets/mL) and (ii) human washed platelets (3 × 10⁸ platelets/mL) were preincubated for 5 minutes with vehicle (DMSO 0.1%) or the MEK inhibitor PD184161 (1-10 μM) followed by stimulation with ristocetin (Rist; 1.5 mg/mL) or thrombin (Thr; 0.05 IU/mL). Experiments were conducted in aggregometer cuvettes with stirring at 37°C. Arrows indicate addition of agonist. Traces are representative of 3 experiments.