Glibenclamide blockade of cftr chloride channels.
Schultz, Bruce D., Albert D. G. Deroos, Charles J. Venglarik, Ashvani
K. Singh, Raymond A. Frizzell, and Robert J. Bridges.
Department of Physiology and Biophysics, University of Alabama at
Birmingham, Birmingham, AL 35294
APStracts 3:0052L, 1996.
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, 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 chloride channels.
Exposure to glibenclamide 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 occurred at 30 [mu]M (Ki). 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 a reduction
in 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 [mu]M-1s
-1), the off rate (koff = 16 s-1), and the dissociation constant (KD =
14 [mu]M). The difference between the Ki (30 [mu]M) and the KD (14
[mu]M) 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.
Received 11 December 1995; accepted in final form 19 March 1996.
APS Manuscript Number L360-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 16 April 96