HETEROGENEITY OF INTRACELLULAR PH AND OF MECHANISMS THAT REGULATE INTRACELLULAR PH IN POPULATIONS OF CULTURED-CELLS

Cells within solid tumors are known to exist in a microenvironment tha t may be acidic and depend on membrane-based mechanisms (Na+/H+ antipo rt and Na+-dependent Cl-/HCO3--exchanger) that regulate intracellular pH (pHi). We have used the fluorescent pH indicator 2',7'-bis-(2-carbo xyethyl) 5 (and 6)-carboxyfluorescein and flow cytometry to study the distribution of pHi and the activity of these pHi-regulating mechanism s among populations of murine mammary sarcoma (EMT6), human breast can cer (MCF-7), and Chinese hamster ovary cells exposed to different leve ls of extracellular pH (pHe). Cells were exposed to Na+ buffer in the presence or absence of HCO3- and of 5-(N-ethyl-N-isopropyl)-amiloride (a potent inhibitor of the Na+/H+ antiport) to determine the relative importance of each exchanger in the regulation of pHi. Our results ind icate that: (a) the distribution of pHi at any value of pHe is broader than can be accounted for by machine noise; (b) cells maintain levels of pHi that are higher than pHe under acidic conditions; (c) the dist ribution of pHi is narrower when the Na+-dependent Cl-/HCO3- exchanger is active; and (d) populations that are derived from selected cells w ith values of pHi at lower and higher ends of the pHi distribution gen erate pHi distributions that are similar to those of controls, suggest ing a stochastic variation in the activity of membrane-based mechanism s that regulate pHi. Our data suggest that the Na+-dependent Cl-/HCO3- exchanger is the dominant mechanism for regulation of pHi under moder ately acidic conditions such as may occur in the microenvironment of s olid tumors.