Agonist-evoked alkaline shift in the cytosolic pH set point for activation of Na⁺/H⁺ antiport in human platelets. The role of cytosolic Ca²⁺ and protein kinase C

This work examines the roles of elevated cytosolic Ca²⁺ and stimulation of protein kinase C in agonist and non-agonist-mediated alkaline shift of the cytosolic pH set point for activation of Na⁺/H⁺ antiport in human platelets. Ca²⁺-depleted and control platelets were subjected to phorbol 12-myristate 13-acetate, thrombin, vasopressin, and ionomycin in 1 mM or Ca²⁺ free, nominally bicarbonate free media. To measure the cytosolic pH set point for Na⁺/H⁺ antiport activation, cells were acidified to different levels using the sodium propionate method. In some experiments protein kinase C was inhibited by staurosporine. Both protein kinase C stimulation and elevation of cytosolic Ca²⁺ can produce an alkaline shift in the pH set point for activation of Na⁺/H⁺ antiport in human platelets. However, the effect of Ca²⁺(i) on the pH set point predominates that of protein kinase C stimulation. Cytosolic Ca²⁺ is a prerequisite for agonist-evoked alkaline shift in the cytosolic pH set point for activation of Na⁺/H⁺ antiport. The cytosolic Ca²⁺ level is also essential for maintaining the basal cytosolic pH. These findings underscore the central role of cytosolic Ca²⁺ under basal and stimulated states in regulating cytosolic pH of human platelets.