AJP - Cell Physiology, Vol 259, Issue 2 C365-C373, Copyright © 1990 by American Physiological Society
H+/base transport in principal cells characterized by confocal fluorescence imaging
X. Wang and I. Kurtz
Department of Medicine, University of California, School of Medicine, Los Angeles 90024.
A dual-excitation inverted confocal laser-scanning microscope has been
developed for measuring intracellular pH (pHi) using
2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF) in individual
cells in the isolated perfused cortical collecting tubule (CCT). This new
microscope has superior depth discrimination, which eliminates the
contribution of fluorescence information from cells outside the plane of
focus. pHi was monitored in real time from a spot 0.55 microns in diameter
within a single cell. Experiments were performed to examine the apical and
basolateral membrane H+/base transport properties of single principal
cells. The results indicate that principal cells possess a basolateral
membrane Na(+)-independent Cl-/base exchanger, a Na(+)-H+ antiporter, and a
Na+/base cotransporter. No evidence was found for an apical membrane
Na(+)-independent Cl-/base exchanger. The data provide evidence for base
efflux pathways in the principal cell and are compatible with the
hypothesis that principal cells contribute importantly to H+/base transport
in the CCT. The new methodology described in this report can be applied to
other epithelia that are optically heterogeneous in the depth dimension.
