Inhibition of human platelet aggregation and cytoplasmic calcium response by calcium antagonists: studies with aequorin and quin2.
Calcium antagonists inhibit platelet aggregation, but whether this action is due to inhibition of the effect of agonists on cytoplasmic ionized calcium concentration is unknown. We studied this problem by loading gel-filtered platelets with either quin2 or aequorin and stimulating them with epinephrine, arachidonate, thrombin, the calcium ionophore A23187, 1-oleoyl-2-acetyl glycerol, or adenosine diphosphate in media with or without extracellular calcium. In response to all of these agonists, aequorin indicated an increase in cytoplasmic calcium that accompanied or preceded platelet aggregation. In calcium-containing media, verapamil, nifedipine, and diltiazem inhibited these effects in a concentration-dependent fashion, except for those produced by thrombin and A23187. Removal of extracellular calcium with EGTA reduced the calcium response to arachidonate, adenosine diphosphate, and 1-oleoyl-2-acetyl glycerol, and the calcium response and aggregation were further inhibited by the calcium antagonists. In general, strong inhibition of the aequorin cytoplasmic calcium signal by approximately 100 microM concentrations of nifedipine, verapamil, and diltiazem was correlated with inhibition of platelet aggregation, but high concentrations of the inhibitors were required. Since inhibition by the calcium antagonists of the cytoplasmic calcium response and aggregation exceeded the effect of simple removal of extracellular calcium, these drugs may affect internal redistribution of calcium in human platelets.
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000331:Aequorin; D000634:Aminoquinolines; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D003470:Culture Media; D003593:Cytoplasm / Q000378:metabolism; D004075:Diglycerides / Q000494:pharmacology; D004110:Diltiazem / Q000494:pharmacology; D004837:Epinephrine / Q000494:pharmacology; D005456:Fluorescent Dyes; D006801:Humans; D008164:Luminescent Proteins; D009543:Nifedipine / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects*; D013917:Thrombin / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
D000634:Aminoquinolines; D001095:Arachidonic Acids; D002121:Calcium Channel Blockers; D003470:Culture Media; D004075:Diglycerides; D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D016718:Arachidonic Acid; D000001:Calcimycin; D000331:Aequorin; D000244:Adenosine Diphosphate; C035738:Quin2-acetoxymethyl ester; C038237:1-oleoyl-2-acetylglycerol; D014700:Verapamil; D013917:Thrombin; D004110:Diltiazem; D009543:Nifedipine; D002118:Calcium; D004837:Epinephrine