Influence of ionized calcium on thrombin-induced down regulation of GPIb/IX receptors on human platelets.

The influence of ionized calcium on the down-regulation of GPIb/IX receptors on human platelets was evaluated by flow cytometry using monoclonal antibodies. Addition of EDTA alone to a washed platelet suspension did not cause decreased monoclonal antibody binding to the cells. However, introduction of thrombin to the washed platelets containing EDTA resulted in a marked decrease in binding of monoclonal antibodies to the GPIb/IX receptors. If calcium at 1-3 mmol/L was added to buffered platelets instead of EDTA before thrombin, down-regulation was prevented or significantly reduced. Restoring calcium to EDTA platelets after the thrombin-stimulated down-regulation had been in progress for 1-3 minutes caused reversal of decreased antibody binding by GPIb/IX to near resting levels. Results demonstrate that extracellular calcium is a major factor regulating thrombin-induced down-regulation.