Bicarbonate conductance and pH regulatory capability of cystic fibrosis transmembrane conductance regulator.

Abstract

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-S08 (AF508-CFTR), reduces Cl1 secretion, but the fatal consequences of CF have been difficult to rationalie solely in terms ofthis defect. The aim ofthis study was to determine the role of CFTR in HCO_ transport across cell membranes. HCO perbili was asse from measurements of intracellular pH [pHi; from spectrofluorimetry of the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-S-(and -6)carboxyfluorescein] and of channel activity (patch damp; cell attached and isolated, inside-out patches) on NIH 3T3 fibroblasts and C127 mammary epithelial cells transfected with wild-type CFTR (WT-CFTR) or AF5S08-CFTR, and also on mock-transfected cells. When WT-CFTR-transfected cells were acidified (pulsed with NH4CI) and incubated in Na+-free (N-methyl-n-glucmine substitution) solutions (to block Na+-dependent pHI, regulatory mechanisms), pH, remained acidic (pH 6.5) until the cells were treated with 20 IAM forskolin (increases cellular [cAMPI); pHi then increased toward (but not completely to) control level (pH1 7.2) at a rate of 0.055 pH unit/min. Forskolin had no effect on rate of pHi recovery in AF508 and mock-transfected cells. This Na+-independent, forskolin-dependent pHi recovery was not observed in HCO /CO2-free medium. Forskolin-treated WT-CFTRtransfected (but not AF508-CFTR or mock-transfected) cells in Cl0-containing, HCO--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 channeis were incubated with different [Cl-I and [HCOi-on the inside and outside, the Cl-/HCO; 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 HCO5. We conclude that in acidified cells the WT-CFTR functions as a base loader by allowing a cAMPdependent influx of HCO-through channeis that conduct HCOj about one-quarter as efflciently as it conducts Cl-.Under physiological conditions, the electrochemical gradients for both Cl-and HCO5 are directed outward, so CFTR likely contributes to the epithellal 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 HCO3 may lead to decreased pH of the luminal fluid.The cystic fibrosis transmembrane conductance regulator (CFTR) is an epithelial Cl-channel regulated by cAMPdependent protein kinase A (1-3). The most common mutation in CF is deletion of Phe-508 of the CFTR (AF508-CFTR), which reduces the ability ofcells to secrete Cl-(4). However, the fatal consequences in CF have been difficult to rationalize solely as a defect in Cl-secretion (5, 6). If the CFTR were also conductive to HCO-, then movements of HCO-might affect the pH of the luminal solution of some epithelia. For example, i...

CFTR mediates bicarbonate-dependent activation of miR-125b in preimplantation embryo development

“…The effect of HCO 3 − in triggering downstream signaling requires CFTR (cystic fibrosis transmembrane conductance regulator), a cAMP-activated anion channel known to conduct Cl − [29,30] and HCO 3 − [31], as the necessary HCO 3 − transport mechanism, either directly or indirectly [28,32]. This suggests that high HCO 3 − concentration may act as an environmental stimulus in initiating cellular responses through CFTR-mediated entry.…”

Gel-forming mucins form distinct morphologic structures in airways

“…Loss of CFTR reduces Cl − and HCO 3 − secretion (33,34). These defects decrease the rate of liquid secretion by submucosal glands and reduce the pH of the secreted liquid (30,31,35).…”

Activation of GABAAReceptor/Cl−Channel and Capacitation in Rat Spermatozoa: HCO3−and Cl−are Essential

“…Our previous studies have demonstrated that the involvement of CFTR in uterine bicarbonate secretion and the fertilizing capacity of sperm were important [Wang et al 2003]. Previously, some studies proposed that a crucial role of the CFTR is to conduct both Cl À and HCO 3 À [Poulsen et al 1994;Reddy and Quinton 2001]. The Cl À ion and its channel, and transportation during sperm capacitation provoked us to investigate this issue.…”