GPIb-IX– and Rac-1–dependent activation of LIMK1 and the roles of LIMK1 in promoting thrombin-induced platelet activation but negatively regulating PAR4-activating peptide-induced platelet activation. Aggregation and ATP release of washed wild-type (WT) or LIMK1^−/− mouse platelets in response to increasing doses of (A-C) thrombin or (D-F) PAR4-activating peptide–induced (PAR4-AP) stimulation. (A,D) Typical aggregation traces. (B,E) Quantification of the aggregation (mean ± SEM, 4 experiments). (C,F) ATP release (mean ± SEM, 4 experiments). **P < .01 and ***P < .001, 1-way ANOVA. Western blot analysis of LIMK1 phosphorylation using a phospho-Thr^505/508–dependent anti-LIMK1 antibody in (G) control CHO cells, 1b9, and Δ605 cells; (H) washed mouse platelets pretreated with either MPαC or MCsC; and (I) wild-type or Rac1^−/− mouse platelets, which were stimulated with low doses of thrombin. Loading levels were determined using a phosphorylation-independent LIMK1 antibody.