Effect of cAMP-PK activation (using BIMPS) on cation influx in platelets mediated by thrombin or OAG.

Fura-2–loaded platelets were incubated at 37°C with additions (marked with arrows or overhead bars), and the elevations of [Ca⁺⁺]_i or [Ba²⁺]_iwere monitored using the 340:380 nm ratio. (A) In the absence of extracellular Ca⁺⁺ (addition of 100 μM EGTA), release of Ca⁺⁺ from stores was activated by the addition of 0.5 U/mL thrombin and subsequent addition of 250 μM BIMPS (red line) or dimethyl sulfoxide (DMSO) vehicle (black line). Influx was recorded with the addition of 1 mM Ca⁺⁺. Peak values upon addition of extracellular Ca⁺⁺ were 0.46 ± 0.015 (n = 3) and 1.06 ± 0.009 (n = 4), BIMPS and vehicle, respectively;P < .001. (B) As in (A) above, with the addition of 1 mM Ba²⁺, 5-minute plateau values were 1.065 ± 0.171 (n = 4) and 0.874 ± 0.167 (n = 5), BIMPS and vehicle, respectively; P = .46. (C) As in (B), except BIMPS or vehicle was added 2 minutes prior to thrombin, 4-minute plateau values of [Ba²⁺]_i were 0.62 ± 0.01 (n = 3) and 1.13 ± 0.02 (n = 3) BIMPS and vehicle, respectively;P < .001. (D) In the presence of 1 mM extracellular Ba²⁺, cells were incubated with 250 μM BIMPS (red line) or DMSO vehicle (black line) before addition of 60 μM OAG. Ratio values at 5 minutes were 0.61 ± 0.057 (n = 3) and 0.54 ± 0.059 (n = 3), BIMPS and vehicle, respectively;P = .44.