Role of Na+/H+ exchange in thrombin- and arachidonic acid-induced Ca2+ influx in platelets.
Platelet activation is accompanied by an increase of cytosolic free Ca2+ concentration, [Ca2+]i, (due to both extracellular Ca2+ influx and Ca2+ movements from the dense tubular system) and an Na+ influx associated with H+ extrusion. The latter event is attributable to the activation of Na+/H+ exchange, which requires Na+ in the extracellular medium and is inhibited by amiloride and its analogs. The present study was carried out to determine whether a link exists between Ca2+ transients (measured by the quin2 method and the 45CaCl2 technique) and Na+/H+ exchange activation (studied with the pH-sensitive intracellular probe, 6-carboxyfluorescein) during platelet stimulation. Washed human platelets, stimulated with thrombin and arachidonic acid, showed: (1) a large and rapid [Ca2+]i rise, mostly due to a Ca2+ influx through the plasma membrane; (2) a marked intracellular alkalinization. Both phenomena were markedly inhibited in the absence of extracellular Na+ or in the presence of an amiloride analog (EIPA). Monensin, a cation exchanger which elicits Na+ influx and alkalinization, and NH4Cl, which induces alkalinization only, were able to evoke an increase in [Ca2+]i, mostly as an influx from the extracellular medium. Our results suggest that Ca2+ influx induced by thrombin and arachidonic acid in human platelets is strictly dependent on Na+/H+-exchange activation.
Ghigo
D
D
Dipartimento di Genetica, Biologia e Chimica Medica, Universita' di Torino, Italy.
Treves
S
S
Turrini
F
F
Pannocchia
A
A
Pescarmona
G
G
Bosia
A
A
D000643:Ammonium Chloride / Q000494:pharmacology; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000097:blood*; D002132:Calcium Radioisotopes; D002352:Carrier Proteins / Q000097:blood*; D003600:Cytosol / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008985:Monensin / Q000494:pharmacology; D010974:Platelet Aggregation; D017923:Sodium-Hydrogen Exchangers; D013917:Thrombin / Q000502:physiology*
D001095:Arachidonic Acids; D002132:Calcium Radioisotopes; D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D000643:Ammonium Chloride; D016718:Arachidonic Acid; D008985:Monensin; D013917:Thrombin; D002118:Calcium