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

摘要: Abstract The cystic fibrosis transmembrane conductance regulator (CFTR) is an epithelial Cl- channel regulated by protein kinase A. The most common mutation in (CF), deletion of Phe-508 (delta F508-CFTR), reduces secretion, but the fatal consequences CF have been difficult to rationalize solely terms this defect. aim study was determine role CFTR HCO3- transport across cell membranes. permeability assessed from measurements intracellular pH [pHi; spectrofluorimetry pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5-(and -6)carboxyfluorescein] and activity (patch clamp; attached isolated, inside-out patches) on NIH 3T3 fibroblasts C127 mammary cells transfected with wild-type (WT-CFTR) or delta F508-CFTR, also mock-transfected cells. When WT-CFTR-transfected were acidified (pulsed NH4Cl) incubated Na(+)-free (N-methyl-D-glucamine substitution) solutions (to block Na(+)-dependent pHi regulatory mechanisms), remained acidic (pH approximately 6.5) until treated 20 microM forskolin (increases cellular [cAMP]); then increased toward (but not completely to) control level (pHi 7.2) at a rate 0.055 unit/min. Forskolin had no effect recovery F508 This Na(+)-independent, forskolin-dependent observed HCO3-/CO2-free medium. Forskolin-treated F508-CFTR mock-transfected) Cl(-)-containing, HCO3(-)-free showed channels linear I/V relationship 10.4 +/- 0.5 pS symmetrical 150 mM Cl-. different [Cl-] [HCO3-] inside outside, Cl-/HCO3- ratio (determined reversal potentials curves) 3.8 1.0 (mean SEM; n = 9); conductances 3.9 (at 127 HCO3-. We conclude that WT-CFTR functions as base loader allowing cAMP-dependent influx through conduct about one-quarter efficiently it conducts Under physiological conditions, electrochemical gradients for both are directed outward, so likely contributes secretion ions. may be important controlling luminal, probably cytoplasmic, fluid CFTR-containing epithelia. In CF, decreased lead luminal fluid.