AJP - Cell Physiology, Vol 259, Issue 1 C110-C120, Copyright © 1990 by American Physiological Society
Regulation of intracellular pH by cultured opossum kidney cells
M. H. Montrose and H. Murer
Institute of Physiology, University of Zurich, Switzerland.
Opossum kidney (OK) cells (an epithelial cell line) were examined by flame
photometry of cellular Na+ and K+ and by microfluorometric measurements of
the intracellular pH (pHi) of single cells loaded with
2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). The work
concentrates on defining resting pHi values under different experimental
conditions and examines factors that contribute to the maintenance of
resting pHi. To use nigericin to calibrate the intracellular response of
BCECF, cellular K+ levels were measured by a null point analysis, and the
stability and magnitude of cellular Na+ and K+ levels were determined vs.
time. Resting pHi in medium without added CO2 was high when measured by
null point analysis of the population (pHi 7.6) and from measurements of
single cells that have recovered from an acid load caused by NH4 prepulse
(pHi 7.76 +/- 0.03, n = 20 cells). In single-cell measurements, addition of
CO2-HCO3- to the medium results in cellular acidification of the
steady-state pHi by 0.35 +/- 0.04 pH units. In medium equilibrated with
room air, the resting pHi is shown to be a dynamic steady state composed of
net flux due to apical Na(+)-dependent transport (Na(+)H+ exchange) plus
acidifying processes. It is concluded that although
5-[N-ethyl-N-isopropyl]amiloride (EIPA) inhibits the forward reaction of
Na(+)-H+ exchange, EIPA is either ineffective as an inhibitor of the
reverse reaction of Na(+)-H+ exchange or Na(+)-H+ exchange does not reverse
measurably in the OK cells.
