Digital image processing of intracellular pH in gastric oxyntic and chief cells

Abstract

Cytosolic pH (pH_i) is a critically regulated determinant of intracellular function. Several mechanisms for pH_i regulation in different tissues have been found, such as direct proton pumping^1,2, Na/H exchange³, Cl/HCO₃ exchange⁴, NaHCO₃ cotransport⁵, and Na/H/Cl/HCO₃ obligatorily linked^6,7. All these studies have used either single cells or cell populations assumed to be behaving homogeneously. Most tissues consist of more than one cell type, so it would be desirable to examinepH_i regulation simultaneously in many identified individual cells, particularly in epithelia where disaggregation and purification of isolated cells destroys the normal distinction between luminal and serosal environments. We have used a pH-sensitive fluorescent dye, BCECF (2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein) and digital image processing to study pHi regulation simultaneously in the oxyntic cells (OC) and chief cells (CC) of gastric glands isolated from rabbit stomach. CCs become markedly more acidic upon removal of external Na (Na₀), but pH_i is restored rapidly on return to normal Na₀, with or without Cl. Oxyntic cell pH_i is much less affected by Na₀. Conversely, OCs become strongly more alkaline on removal of external Cl (Cl₀),pH_ibeing restored when Cl₀ is replaced with or without Na, whereas CCs are relatively insensitive to C1₀. Therefore, Na/H exchange is dominant over Cl/HCO₃ exchange in CCs, but in the neighbouring OCs, C1/HCO₃outweighs the Na/H mechanism, a heterogeneity that correlates with the functions of the two cell types.