ABSTRACr We investigated the role of sodium in megakaryocyte spreading induced by thrombin and ADP. We found that if extracellular sodium was replaced by lithium, potassium, or choline, spreading was inhibited. When extracellular sodium was present, amiloride or tetrodotoxin inhibited spreading. Using intracellular recording we found spreading to be associated with a permanent membrane depolarization. The extent and rate of thrombin-induced depolarization was reduced when lithium replaced sodium. Unspread cells had an average membrane potential of -44.8 mV. Spread cells had an average membrane potential of -18.46 mV. When choline replaced sodium, or when in the presence of tetrodotoxin and amiloride, the spread cells repolarized, indicating that the depolarization is due to an increase in sodium permeability. Similar treatments did not change the membrane potential of unspread cells. Incubation of megakaryocytes with A23187 together with monensin or methylamine induced spreading. Methylamine occasionally caused spreading by itself, but neither ionophore alone caused spreading. These results indicate that megakaryocyte spreading induced by ADP and thrombin depends on an increase in sodium conductance.Platelets and megakaryocytes, the platelets' precursor cells, have a number of physiological characteristics in common. Both are activated by the platelet agonists ADP, thrombin, and arachidonic acid, and the activation of these ceils is inhibited by prostaglandin E1 (PGE1), tetracaine, and dibutyryl cAMP (1-5). The first step in platelet activation is a change in cell shape: f'dopodia and membrane protrusions form. Isolated, cultured megakaryocytes also respond to platelet agonists by a change in cell shape; fdopodia and a ruffled membrane form and the cells, resting on a surface, become adherent and spread out over the substratum much as platelets spread on glass (4). In both cases, the shape change involves the formation of microfdament nets and bundles from an amorphous precursor material (6; R. M. Leven and V. T. Nachmias, manuscript submitted for publication). The mechanism by which various agents, acting at the cell surface, cause the state of the cytoplasm to change in this way is unknown. However, the close similarities in the responses of these two cell types, and the fact that platelets are themselves fragments of the megakaryocyte cytoplasm, suggest strongly that the mechanism of activation of both platelets and megakaryocytes is very similar, if not the same.Experiments with platelets have recently suggested a possible role for sodium and pH changes as stimuli for the cytoskeletal reorganization that underlies shape change. Feinberg et al. (7) have shown that ADP causes an increased influx of sodium into platelets, although the correlation of shape change with sodium influx was not studied. Simons et al. (8), using membrane potential and pH sensitive dyes, have shown that the intracellular pH increases -0.3 pH unit in thrombin-stimulated platelets and that the membrane potential depolarizes from a restin...
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