Rac1 regulates shear-induced PS exposure and microvesiculation independent of its role in stimulating platelet secretion and aggregation. (A) Representative aggregation and ATP secretion traces of washed human platelets activated with 0.035 U/mL thrombin following treatment with 2 µg/mL C3-toxin, 100 µM NSC23766, or vehicle control. (B) Washed human platelets were treated with vehicle, 2 µg/mL C3-toxin (C3), or 100 μM NSC23766 (NSC), stimulated with 0.025 U/mL thrombin, subjected to shear, and analyzed for PS exposure and MV release. Data are expressed as mean ± SEM, n = 3. (C) Quantification of ATP secretion (left y-axis) and PS exposure (right y-axis) in washed mouse platelets, either left static or subjected to shear, in the presence or absence of thrombin (0.03 U/mL) (mean ± SEM, n = 3). (D) Aggregation of NSC23766- or DMSO-treated human platelets following stimulation with 60 µM AYPGKF (PAR4) or 2 µM SFLLRN (PAR1) in the presence or absence of 1 µM ADP; 1 µM ADP was added alone as a control. (E) Platelets treated under the same conditions as C were analyzed for the fold increase in shear-induced MV release from the resting baseline (mean ± SEM, n = 8). For all data, *, **, and *** represent statistical significance of P < .05, .01, and .001, respectively.