Mechanism of collagen activation in human platelets.
The mechanism of collagen-induced human platelet activation was examined using Ca2+, Na+, and the pH-sensitive fluorescent dyes calcium green/fura red, sodium-binding benzofuran isophthalate, and 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Administration of a moderate dose of collagen (10 microg/ml) to human platelets resulted in an increase in [Ca2+](i) and platelet aggregation. The majority of this increase in [Ca2+](i) resulted from the influx of calcium from the extracellular milieu via the Na+/Ca2+ exchanger (NCX) functioning in the reverse mode and was reduced in a dose-dependent manner by the NCX inhibitors 5-(4-chlorobenzyl)-2',4'-dimethylbenzamil (KD(50) = 4.7 +/- 1.1 microm) and KB-R7943 (KD(50) = 35.1 +/- 4.8 microm). Collagen-induced platelet aggregation was dependent on an increase in [Ca2+](i) and could be inhibited by chelation of intra- and extracellular calcium through the administration of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) (BAPTA-AM) and EGTA, respectively, or via the administration of BAPTA-AM to platelets suspended in no-Na+/HEPES buffer. Collagen induced an increase in [Ca2+](i) (23.2 +/- 7.6 mm) via the actions of thromboxane A(2) and, to a lesser extent, of the Na+/H+ exchanger. This study demonstrates that the collagen-induced increase in [Ca2+](i) is dependent on the concentration of Na+ in the extracellular milieu, indicating that the collagen-induced increase in [Ca2+](i) causes the reversal of the NCX, ultimately resulting in an increase in [Ca2+](i) and platelet aggregation.
Department of Pharmacology and Therapeutics, University of Manitoba, 753 McDermot Avenue, Winnipeg, Manitoba R3E 0W3, Canada.
Diane E
DE
Roberts
Archibald
A
McNicol
Ratna
R
Bose
D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D001241:Aspirin / Q000494:pharmacology; D001572:Benzofurans / Q000494:pharmacology; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D003094:Collagen / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology; D005452:Fluoresceins / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008954:Models, Biological; D009930:Organic Chemicals; D015539:Platelet Activation / Q000187:drug effects; D010974:Platelet Aggregation; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000502:physiology; D013048:Specimen Handling; D013696:Temperature; D013928:Thromboxane A2 / Q000494:pharmacology; D013997:Time Factors; D014662:Vasoconstrictor Agents / Q000494:pharmacology
D001572:Benzofurans; D004988:Ethers, Cyclic; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007093:Imidazoles; D009930:Organic Chemicals; D019831:Sodium-Calcium Exchanger; D014662:Vasoconstrictor Agents; C061665:sodium-binding benzofuran isophthalate; C071336:calcium green; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; C082877:fura red; C024098:6-carboxyfluorescein; D004533:Egtazic Acid; D013928:Thromboxane A2; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D015544:Inositol 1,4,5-Trisphosphate; D003094:Collagen; D012964:Sodium; D001241:Aspirin; D002118:Calcium