Na-H and Cl-HCO3 exchange in rabbit oxyntic cells using fluorescence microscopy

We have used the fluorescence intensity ratio (excitation 490/439; emission 520-550 nm) of 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) to measure intracellular pH (pHi) in single oxyntic cells (OCs) within intact gastric glands isolated from rabbits. The fluorescence ratio was converted to pHi by exposing cells to high [K] Ringer plus 10(-5) M nigericin. Ratios varied linearly with pHi over the physiological range. When the bathing solution was changed from NaCl to Na gluconate Ringer, pHi increased from 7.0 to 7.4. The increase of pHi occurred equally rapidly in nominally CO2-HCO3-free solutions and in solutions containing 5% CO2 and 25 mM HCO3. This effect was reversible and blocked by 2 X 10(-4) M 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS). The DIDS block was bypassed by 10(-5) M tri-n-butyltin, a pharmacological Cl-OH exchanger. When the bathing solution was changed from NaCl to N-methyl-D-glucamine (NMG) Cl Ringer, pHi decreased reversibly from 7.0 to 6.8. It was also found that changing from NMG gluconate to Na gluconate Ringer caused pHi to increase from 7.1 to 7.3, and this alkalinization was blocked by 10(-3) M amiloride; changing from NMG gluconate to NMG Cl Ringer caused pHi to decrease to 6.7. We conclude that OCs contain separate Na-H and Cl-HCO3 exchangers. Similarities and differences between these exchangers and those present in other cell types are discussed.