Identification of a sodium-bicarbonate symport in human platelets

Intracellular pH (pH(i)) was measured in human platelets using fluorescent probes. Basal pH(i) was higher in HCO₃^--buffered solutions (7.33 ± 0.01) than in nominally HCO₃^- free, Hepes-buffered solutions (7.16 ± 0.01, P < 0.05). Addition of EIPA caused a fall in Hepes, but did not inhibit the increase of pH(i) when platelets maintained in Hepes were transferred to a CO₂/HCO₃^- buffer. After an intracellular acidosis induced by an NH₄Cl prepulse, the initial velocity of recovery (d(pH)/dt(i), in pH units/min) was 3.32 ± 0.69 in Hepes-buffered solution and 2.85 ± 0.88 in HCO₃^- media. Taking into account the differences in buffer capacity, the efflux of acid equivalents after 1.2 min was twice as much in the presence of bicarbonate. The addition of 30 μmol/l EIPA effectively blocked acid efflux (d(pH)/dt(i) = 0.08 ± 0.03) in a nominally HCO₃^- -free solution, whereas the recovery was reduced but not abolished (d(pH)/dt(i) = 0.37 ± 0.10, P < 0.05) in the presence of bicarbonate. The stilbene derivative SITS further inhibited the ETPA-resistant pH(i) recovery. Removal of external Na⁺ inhibited the HCO₃^- -dependent recovery whereas depletion of internal Cl^-, did not suppress it. Depolarization of the membrane had no effect on this recovery. The results suggest the contribution of an electroneutral Na⁺/HCO₃^- cotransport in the recovery of pH(i) following an acid load. Both the Na⁺/H⁺ antiport and the HCO₃^--dependent mechanism contribute approx. 50% each to the total acid equivalent efflux during the recovery from a pH(i) 6.46 ± 0.14 to the basal pH(i) in human platelets.