Removal of extraplatelet Na+ eliminates indomethacin-sensitive secretion from human platelets stimulated by epinephrine, ADP, and thrombin.

We have previously observed that removal of extraplatelet Na+ markedly diminishes human platelet aggregation and secretion in response to epinephrine. The present studies demonstrate that this effect of the removal of extraplatelet Na+ on platelet function is not unique to activation of platelets by alpha 2-adrenergic agents but represents a phenomenon also evident for other platelet stimuli. Thus, platelet aggregation and secretion in response to maximal concentrations of ADP and lower concentrations of thrombin (less than 0.04 unit/ml) were also markedly reduced in platelets in "Na+-free" medium, suggesting that these agents share an effector mechanism that is similarly inhibited by the removal of extraplatelet Na+. In contrast, platelet aggregation and secretion in response to higher concentrations of thrombin (greater than or equal to 0.04 unit/ml) and to 0.04-1.0 microM (15S)-hydroxy-11 alpha, 9 alpha-(epoxymethano)prosta-5Z,13E-dienoic acid (U46619), an endoperoxide analog, were identical in control platelets and in those suspended in "Na+-free" medium, indicating that platelets suspended in "Na+-free" medium are functionally intact, at least in response to some stimuli. Furthermore, the observation that U46619 can elicit platelet aggregation and secretion independently of extraplatelet Na+ indicates that the loss of platelet responsiveness to epinephrine, ADP, and low concentrations of thrombin cannot be attributed to a loss of sensitivity to the stimulus-provoked secondary mediator(s) of platelet function, endoperoxides or thromboxane A2. Treatment with indomethacin to block the secondary aggregation and secretion pathways of platelets reduced the aggregatory and secretory responses of control platelets induced by epinephrine, ADP, and low concentrations of thrombin to those characteristic of platelets suspended in "Na+-free" medium. In contrast, indomethacin did not alter the functional responses induced by these agents in platelets suspended in "Na+-free" medium, suggesting that "primary" aggregation is intact but that the "secondary" aggregation and secretion mediated by arachidonic acid metabolites are eliminated by removal of extraplatelet Na+. Consistent with this interpretation is the observation that the indomethacin-insensitive aggregation and secretion induced by U46619 and higher concentrations of thrombin were retained in platelets suspended in "Na+-free" medium. Thus, the responses eliminated by removal of extraplatelet Na+ are those eliminated by treating control platelets with indomethacin, suggesting a strong link between the presence of extraplatelet Na+ and the operation of platelet function mediated by the cyclooxygenase pathway.