Proceedings of the National Academy of Sciences, Vol 91, 5340-5344, Copyright © 1994 by National Academy of Sciences
Bicarbonate Conductance and pH Regulatory Capability of Cystic Fibrosis Transmembrane Conductance Regulator
JH Poulsen, H Fischer, B Illek and TE Machen
The cystic fibrosis transmembrane conductance regulator (CFTR) is an
epithelial Cl- channel regulated by protein kinase A. The most
common mutation in cystic fibrosis (CF), deletion of Phe-508 (
F508-CFTR), reduces
Cl- secretion, but the fatal consequences of CF have been
difficult to rationalize solely in terms of this defect. The aim of this
study was to determine the role of CFTR in HCO-3
transport across cell membranes. HCO-3 permeability
was assessed from measurements of intracellular pH [pHi; from
spectrofluorimetry of the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5-(and
-6)carboxyfluorescein] and of channel activity (patch clamp; cell attached
and isolated, inside-out patches) on NIH 3T3 fibroblasts and C127 mammary
epithelial cells transfected with wild-type CFTR (WT-CFTR) or F508-CFTR, and also on
mock-transfected cells. When WT-CFTR-transfected cells were acidified
(pulsed with NH4Cl) and incubated in Na+-free
(N-methyl-D-glucamine substitution) solutions (to block
Na+-dependent pHi regulatory mechanisms),
pHi remained acidic (pH 
6.5) until the cells were treated with 20 µM forskolin
(increases cellular [cAMP]); pHi then increased toward (but not
completely to) control level (pHi 7.2) at a rate of 0.055 pH
unit/min. Forskolin had no effect on rate of pHi recovery in
F508 and
mock-transfected cells. This Na+-independent,
forskolin-dependent pHi recovery was not observed in
HCO-3/CO2-free medium. Forskolin-treated
WT-CFTR-transfected (but not F508-CFTR or mock-transfected) cells in
Cl--containing, HCO-3-free solutions
showed Cl- channels with a linear I/V relationship and a
conductance of 10.4 ± 0.5 pS in symmetrical 150 mM Cl-.
When channels were incubated with different [Cl-] and
[HCO-3] on the inside and outside, the
Cl-/HCO-3 permeability ratio (determined
from reversal potentials of I/V curves) was 3.8 ± 1.0 (mean ±
SEM; n = 9); the ratio of conductances was 3.9 ± 0.5 (at 150 mM
Cl- and 127 mM HCO-3. We conclude that in
acidified cells the WT-CFTR functions as a base loader by allowing a
cAMP-dependent influx of HCO-3 through channels that
conduct HCO-3 about one-quarter as efficiently as it
conducts Cl-. Under physiological conditions, the
electrochemical gradients for both Cl- and
HCO-3 are directed outward, so CFTR likely
contributes to the epithelial secretion of both ions.
HCO-3 secretion may be important for controlling pH
of the luminal, but probably not the cytoplasmic, fluid in CFTR-containing
epithelia. In CF, a decreased secretion of HCO-3 may
lead to decreased pH of the luminal fluid.
