Appearance of Additional Ouabain Binding Sites on Platelet Surfaces Following Activation

Bork, Tracie; Mrsny, Randall J.

doi:10.1006/excr.1993.1237

Cited by 5 publications

(3 citation statements)

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“…Another possibility is that there is a change in surface expression of the Na + /K + -ATPase in thrombin-stimulated platelets. Surface expression of this transporter has been reported to increase more than ten-fold in ADP stimulated platelets as assessed by [ 3 H]-ouabain binding [57]. Given the central role of PKC in granule secretion in platelets [32], it is not inconceivable that thrombin-evoked activation of PKC may also increase the apparent activity of Na + /K + -ATPase through modulation of its expression in the platelet plasma membrane.…”

Section: Discussionmentioning

confidence: 99%

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

et al. 2015

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Rises in cytosolic Ca 2+ concentration ([Ca 2+ ] cyt ) are central in platelet activation, yet many aspects of the underlying mechanisms are poorly understood. Most studies examine how experimental manipulations affect agonist-evoked rises in [Ca 2+ ] cyt , but these only monitor the net effect of manipulations on the processes controlling [Ca 2+ ] cyt (Ca 2+ buffering, sequestration, release, entry and removal), and cannot resolve the source of the Ca 2+ or the transporters or channels affected. To investigate the effects of protein kinase C (PKC) on platelet Ca 2+ signalling, we here monitor Ca 2+ flux around the platelet by measuring net Ca 2+ fluxes to or from the extracellular space and the intracellular Ca 2+ stores, which act as the major sources and sinks for Ca 2+ influx into and efflux from the cytosol, as well as monitoring the cytosolic Na + concentration ([Na + ] cyt ), which influences platelet Ca 2+ fluxes via Na + /Ca 2+ exchange. The intracellular store Ca 2+ concentration ([Ca 2+ ] st ) was monitored using Fluo-5N, the extracellular Ca 2+ concentration ([Ca 2+ ] ext ) was monitored using Fluo-4 whilst [Ca 2+ ] cyt and [Na + ] cyt were monitored using Fura-2 and SFBI respectively. PKC inhibition using Ro-31-8220 or bisindolaemide I potentiated ADP-and thrombin-evoked rises in [Ca 2+ ] cyt in the absence of extracellular Ca 2+ . PKC inhibition potentiated ADP-evoked but reduced thrombin-evoked intracellular Ca 2+ release and Ca 2+ removal into the extracellular medium. SERCA inhibition using thapsigargin and 2,5-di(tert-butyl) l,4-benzohydroquinone abolished the effect of PKC inhibitors on ADP-evoked changes in [Ca 2+ ] cyt but only reduced the effect on thrombin-evoked responses. Thrombin evokes substantial rises in [Na + ] cyt which would be expected to reduce Ca 2+ removal via the Na + /Ca 2+ exchanger (NCX). Thrombinevoked rises in [Na + ] cyt were potentiated by PKC inhibition, an effect which was not due to altered changes in non-selective cation permeability of the plasma membrane as assessed by Mn 2+ quench of Fura-2 fluorescence. PKC inhibition was without effect on thrombin-evoked rises in [Ca 2+ ] cyt following SERCA inhibition and either removal of extracellular Na + or inhibition of Na + /K + -ATPase activity by removal of extracellular K + or treatment with digoxin. These data suggest that PKC limits ADP-evoked rises in [Ca 2+ ] cyt by acceleration of SERCA activity, whilst rises in [Ca 2+ ] cyt evoked by the stronger platelet activator thrombin are limited by PKC through acceleration of both SERCA and Na + /K + -ATPase activity, with the latter limiting the effect of thrombin on rises in [Na + ] cyt and so forward mode NCX activity. The use of selective PKC inhibitors indicated that conventional and not novel PKC isoforms are responsible for the inhibition of agonist-evoked Ca 2+ signalling.

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Section: Discussionmentioning

confidence: 99%

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

et al. 2015

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“…Platelets are likely to possess at least two isoforms of Na + /K + -ATPase, since exogenously added plant ouabain is not able to inhibit its activity totally (Marx et al, 1992). Activated platelets have been reported to express additional binding sites for ouabain, which may result in a rise in the sensitivity of their Na + /K + -ATPase to cardiac glycosides (Bork & Mrsny, 1993). Since patients with atrial fibrillation and heart failure are reported to have preactivated platelets (Kamath et al, 2001;Choudhury et al, 2007), it is likely that their Na + /K + -ATPase is also more sensitive to cardiac glycosides.…”

Section: Discussionmentioning

confidence: 99%

“…Activated platelets play a crucial role in the generation of the hypercoagulable state of blood (Heemskerk et al, 2002). Platelets are a likely target of endogenous Na + /K + -ATPase inhibitors since specific ouabain binding sites have been reported to be present on their surface (Bork & Mrsny, 1993). The ouabain-induced rise in [Na + ] i is reported to produce water accumulation in platelets and their subsequent swelling.…”

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

Tomasiak¹,

Stelmach²,

Rusak³

et al. 2007

Acta Biochim Pol

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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.

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Do cardiac glycosides affect platelet function? A flow cytometric study in healthy volunteers.

Pettersen

Hagberg²,

Lyberg³

et al. 2002

Eur J Clin Pharmacol

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Appearance of Additional Ouabain Binding Sites on Platelet Surfaces Following Activation

Cited by 5 publications

References 0 publications

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

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

Do cardiac glycosides affect platelet function? A flow cytometric study in healthy volunteers.

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