Comparison of the latency and time course of thrombin-evoked Ca²⁺ release, Mn²⁺ entry, and actin reorganization. (A) Fura-2-loaded human platelets were rapidly mixed with thrombin (at 0 seconds) at a final concentration of 0.1 U/mL in the presence of 100 μM of EGTA and 200 μM MnCl₂. Fura-2 fluorescence was recorded at excitation wavelengths of 340 nm (black trace, left axis) and 360 nm (gray trace, right axis). Traces are representative of 20 runs made on cell preparations from 10 donors. (B) Human platelets were rapidly mixed with 0.1 U/mL thrombin (•) or with HBS (dashed line) and incubated at 37°C for various time periods (1-5 seconds) before mixing with formaldehyde (3% in PBS). Actin filament content was determined as described in “Materials and methods.” Results shown are presented as percentage of the F-actin content in resting cells and expressed as mean ± SE of 21 runs made on cell preparations from 12 donors. Vertical dashed lines represent the starting times for Ca²⁺ release and Mn²⁺ entry.