TRANSFER OF TRIS BUFFER AND EFFECTS ON K+ LOSS IN HUMAN RED-BLOOD-CELLS AND RECONSTITUTED GHOSTS

The rate of volume changes of human red blood cells in the presence of Tris-HCl is pH-dependent. At 37.degree. C, t1/2 is 25-30 min at pH 7.4 and 10-20 min at pH 8.4. Hemolysis in Tris-HCl is delayed by H2 DIDS [4,4''-diisothiocyanodihydrostilbene-2,2''-disulfonate] but is promoted by low concentrations of bicarbonate. This bicarbonate effect was reversed by inhibiting carbonic anhydrase with acetazolamide. K+ loss of red blood cells is increased at 37.degree. C in isotonic NaCl solutions containing in addition Tris-HCl. This Tris effect is enhanced from pH 6.4 to 8.4. At pH 8.4 K+ loss is stimulated about 3-fold by addition of 160 mM Tris-HCl. The onset of the Tris effect is delayed at pH 7.4 and below, but not at pH 8.4. Such a delay is absent after preincubation of the cells with Tris-HCl. After binding H2DIDS to red cells, no Tris-dependent increase of K+ loss was observed. K+ loss of reconstituted red cell ghosts with equal internal and external chloride concentrations remained unaffected by Tris-HCl added to the external solution. In ghosts containing sucrose for isotonicity instead of choline chloride K+ loss is smaller but is stimulated by Tris-HCl approaching the rate in those ghosts with equal internal and external chloride concentrations. The transfer of Tris-HCl into red blood cells depends on the pH and on the chloride shift. As there is evidence that Tris-HCl raises the intracellular pH and reduces the Donnan potential at the membrane, K+ loss of red cells may be increased following an intracellular buffer interaction of Hb and Tris-HCl.