Cation influx in response to oleoyl acetyl glycerol (OAG), thapsigargin (Tg), and thrombin.
Fura-2–loaded platelets were incubated at 37°C with additions (marked with arrows), and the ratio (340:380 nm) fluorescence increases were measured in experiments using extracellular Ca++(continuous line) or Ba2+ (dashed line). (A) 1 mM Ca++ or Ba2+ was added to platelets followed by 60 μM OAG 2 minutes later. The increase in ratio fluorescence 4 minutes after OAG addition was 0.588 ± 0.072 (n = 13) and 0.58 ± 0.056 (n = 9) ratio units, for Ca++ and Ba2+, respectively; P = .93. (B) 100 μM EGTA was added to platelets, followed by 3 μM Tg 1 minute later. 1 mM Ca++ or 1 mM Ba2+ was added 3 minutes later, and fluorescence recordings continued for a further 4 minutes. Increase of ratio fluorescence 4 minutes after cation addition was 3.24 ± 0.354 (n = 4) and 0.46 ± 0.05 (n = 6) for Ca++ and Ba2+, respectively;P < .001 (C) 1 U/mL thrombin was added 1 minute after 100 μM EGTA. 2 minutes later either 1 mM Ca++ or Ba2+ was added. Ratio fluorescence increases with 1 mM Ca++ were at peak (0.5 minutes) 1.2 ± 0.082 and plateau (4 minutes) 0.46 ± 0.138 (n = 4); and for Ba2+ at 0.5 minutes was 0.29 ± 0.02 and at plateau 0.66 ± 0.053 (n = 6); at 0.5 minutes P < .001, at 4 minutesP = .153.
