The involvement of Na+/K(+)-ATPase in the development of platelet procoagulant response.

Abstract

In circulation, platelets may come into contact with both exogenous (cardiac glycoside treatment) and endogenously produced inhibitors of Na+/K(+)-ATPase. We examined whether blocking of platelet Na+/K(+)-ATPase by ouabain results in generation of procoagulant activity. It was shown that an in vitro treatment of platelets with ouabain (20-200 microM for 20 to 60 min) is associated with an intracellular accumulation of sodium ([Na+](i)), generation of a weak calcium signal, and expression of procoagulant activity. The ouabain-induced procoagulant response was dose- and time-related, less pronounced than that evoked by collagen and similar to that produced by gramicidin, not affected by EDTA or aspirin, and strongly reduced in the absence of extracellular Na+ or by hyperosmolality. Flow cytometry studies revealed that ouabain treatment results in a unimodal left shift in the forward and side scatter of the entire platelet population indicating morphological changes of the plasma membrane. The shift was dose related, weaker than that evoked by collagen and similar to that produced by gramicidin. Ouabain-treated platelets express phosphatidylserine (PS). The ouabain-evoked PS expression was dose- and time-dependent, weaker than that produced by collagen and similar to that evoked by gramicidin. Electronic cell sizing measurements showed a dose-dependent increase in mean platelet volume upon treatment with ouabain. Hypoosmotically-evoked platelet swelling resulted in the appearance of procoagulant activity. Thromboelastography measurements indicate that, in whole blood, nanomolar (50-1000 nM, 15 min) concentrations of ouabain significantly accelerate the rate of clot formation initiated by contact and high extracellular concentration of calcium. We conclude that inefficiently operating platelet Na+/K(+)-ATPase results in a rise in [Na+](i). An increase in [Na+](i) and the swelling associated with it may produce PS exposure and a rise in membrane curvature leading to the generation of a procoagulant activity.

Flow Cytometric Monitoring of Dynamic Cytosolic Calcium, Sodium, and Potassium Fluxes Following Platelet Activation

“…This work is the first detailed representation of flow-cytometric sodium kinetics with ING-2 during platelet activation, and it was performed with a larger panel of agonists and combination thereof. Our method confirms the importance of extracellular sodium for platelet activation (69,(72)(73)(74) and offers an easy tool for further analyses.…”

The plasma membrane calcium ATPase modulates calcium homeostasis, intracellular signaling events and function in platelets

“…Critical to our argument, the validity of these functional observations has been confirmed by studies in PMCA4 –/– and transgenic lines [11,15,27,28], suggesting a predominant action of CE at PMCA in a range of cell types. CE does, however, inhibit Na + /K + ‐ATPase activity at micromolar concentrations [17], and effects on transients during Ca 2+ repletion [19] and procoagulant effects [29] have been reported. We could find no published evidence for a role of this pump in modulating platelet aggregation, and demonstrated here that the Na + /K + ‐ATPase inhibitor ouabain had no effect on platelet aggregation under our experimental conditions.…”

Sodium–Calcium Exchanger Reverse Mode Sustains Dichotomous Ion Fluxes Required for Procoagulant COAT Platelet Formation

“…16 Our (see Colucci et al 15 and ►Supplementary Fig. S1, available in the online version) and others' 16,36 works highlight the importance of cytosolic sodium for the procoagulant response.…”