α-Actinin-1 deficiency inhibits platelet activation, actin polymerization, calcium mobilization, and ROS generation. Flow cytometric analyses of JON/A binding (for activated integrin αIIbβ3) on washed platelets from Actn1^f/f and PF4-Actn1^−/− mice after stimulation with different agonists, including (A) ADP (0, 10, 20, and 40 μmol/L; n = 6 mice per group), (B) ADP + epinephrine (Epi; 0, 10, 20, and 40 μmol/L; n = 4-10 mice per group), (C) thrombin (0, 0.025, 0.05, and 0.1 U/mL; n = 6-8 mice per group), (D) collagen (0, 2, 4, and 8 μg/mL; n = 10 mice per group), and (E) convulxin (0, 100, 200, and 400 ng/mL; n = 4-6 mice per group). Flow cytometric analyses of CD62P (for exposure) on washed platelets from Actn1^f/f and PF4-Actn1^−/− mice after stimulation with (F) ADP (0, 10, 20, and 40 μmol/L; n = 6 mice per group), (G) ADP + Epi (0, 10, 20, and 40 μmol/L; n = 4-10 mice per group), (H) thrombin (0, 0.025, 0.05, and 0.1 U/mL; n = 6-10 mice per group), (I) collagen (0, 2, 4, and 8 μg/mL; n = 10 mice per group), or (J) convulxin (0, 100, 200, and 400 ng/mL; n = 4-6 mice per group). Actn1^f/f and PF4-Actn1^−/− platelets were stimulated with thrombin (n = 4-7 mice per group) and collagen (n = 7 mice per group) at the indicated concentrations, and the relative F-actin content (K) was determined by flow cytometry. (L) Representative traces of changes in global calcium content (n = 6 mice per group). (M) The levels of intracellular ROS in Actn1^f/f and PF4-Actn1^−/− platelets stimulated with thrombin (n = 4-8 mice per group) and collagen (n = 4-7 mice per group) at the indicated concentrations were determined by flow cytometry. The data are presented as the mean ± SD. ∗P < .05; ∗∗P < .01; ∗∗∗P < .005. Fluo-4 AM, Fluo-4 acetoxymethyl ester; MFI, mean fluorescence intensity; ns, not significant.