Delta F508-CFTR channels: Kinetics, activation by forskolin, and potentiation by xanthines

Trafficking, activation, and kinetics of Delta F508-cystic fibrosis transmembrane conductance regulator (CFTR) and CFTR were compared in stably transduced C127I mouse mammary epithelial cells. Western blots detected a small amount of fully glycosylated Delta F508-CFTR. Efflux of I-125 was stimulated by forskolin with the same mean effective concentration (EC(50); similar to 0.5 mu M) for CFTR and Delta F508-CFTR cells, but the maximum response was reduced more than fivefold and its latency increased approximately threefold in Delta F508-CFTR cells. In Delta F508-CFTR cells, 3-isobutyl-1-methylxanthine (IBMX; EC(50) = 1.45 mM) and 8-cyclopentyl-1,3-dipropylxanthine (CPX; EC(50) = 58 mu M) increased the peak forskolin-stimulated efflux rate similar to 2.5-fold and decreased the time to peak. A sevenfold increase in intracellular adenosine 3',5'-cyclic monophosphate (cAMP) levels accompanied potentiation of forskolin-induced I-125 efflux by IBMX but not by CPX. Elevation of intracellular cAMP increased linear voltage-independent whole cell currents 30-fold in CFTR and 4-fold in Delta F508-CFTR cells; the response rate in Delta F508-CFTR cells was much slower. Single-channel currents were detected in 57 of 68 cell-attached patches from forskolin-prestimulated CFTR cells vs. 6 of 35 patches in Delta F508-CFTR cells. Mean number of active channels per patch was 4.1 for CFTR [open probability (P-o) = 0.34] and 0.2 for Delta F508-CFTR (P-o = 0.11). The lower P-o of Delta F508-CFTR resulted from an approximately threefold longer mean interburst interval. We estimate that forskolin-stimulated chloride conductance of Delta F508-CFTR C127I cells is <5% of CFTR cells. CPX is similar to 25-fold more potent than IBMX in potentiating Delta F508-CFTR and may operate by a mechanism other than elevation of cAMP.