Two distinct HCO3--dependent H+ efflux pathways in human vascular endothelial cells

Intracellular pH (pH(i)) regulation in human umbilical vein endothelial cel ls (HUVEC) was investigated. The pH(i) was recorded using seminaphthorhodaf luor-1 (SNARF-1). Cells were intracellularly acid loaded with NH4Cl prepuls e. In HEPES-buffered Tyrode (nominally HCO3- free), pH(i) recovery from aci d load was inhibited by 1.5 mM amiloride or Na+-free solution. Additionally , in HCO3--buffered Tyrode, a HCO3--dependent pH(i) recovery from acidosis was evident in the presence of 1.5 mM amiloride, which mediated complete re covery of pHi (7.26). In Na+-free solution, the HCO3--dependent acid extrud er mediated pH(i) recovery after an acid load but only back to 7.09. These results suggest that there are two HCO3--dependent acid extruders in the HU VEC. One is Na+ dependent, and the other is Na+ independent. The former was further shown to be completely inhibited by 0.5 mM DIDS, whereas the latte r was only inhibited by 24.6%. In Cl--free solution, both of the HCO3--depe ndent pathways were inhibited. In conclusion, one HCO3--dependent acid extr uder in the HUVEC resembles the Na+-dependent Cl-/HCO3- exchange found in o ther tissues, and the other is Cl-dependent but Na+ independent.