Glibenclamide blockade of CFTR chloride channels.

The cystic fibrosis transmembrane conductance regulator (CFTR) is a protein kinase A- and ATP-regulated Cl- channel located in the apical membranes of epithelial cells. Previously Sheppard and Welsh (J. Gen. Physiol. 100: 573-591, 1992) showed that glibenclamide, a compound which binds to the sulfonylurea receptor and thus blocks nucleotide-dependent K+ channels, reduced CFTR whole cell current. The aim of this study was to identify the mechanism underlying this inhibition in cell-free membrane patches containing CFTR Cl- channels. Exposure to gliben-clamide caused a reversible reduction in current carried by CFTR which was paralleled by a decrease in channel open probability (Po). The decrease in Po was concentration dependent, and half-maximum inhibition (ki) occurred at 30 microM. Fluctuation analysis indicated a flickery-type block of open CFTR channels. Event duration analysis supported this notion by showing that the glibenclamide-induced decrease in Po was accompanied by interruptions of open bursts [i.e., an apparent reduction in the burst duration (Tburst)] with only a slight reduction in closed time (Tc). The plot of the corresponding open-to-closed (Tburst-1) and closed-to-open (Tc-1) rates as a function of glibenclamide concentration were consistent with a pseudo-first-order open-blocked mechanism and provided estimates of the on rate (kon = 1.17 microM-1S-1), the off rate (koff = 16 s-1), and the dissociation constant (Kd = 14 microM). The difference between the Ki (30 microM) and the Kd (14 microM) is the result expected for a closed-open-blocked model with an initial Po of 0.47. Since the initial Po was 0.50 +/- 0.02 (n = 12), we can conclude that glibenclamide blocks CFTR by a closed-open-blocked mechanism.