title: Cystic fibrosis transmembrane conductance regulator mediates sulphonylurea block of the inwardly rectifying K+ channel Kir6.1.
abstract: 1. Recombinant ATP-sensitive K+ channels (KATP channels) were heterologously expressed in the NIH3T3 mouse cell line, and the electrophysiological properties were studied using patch-clamp techniques. 2. The NIH3T3 cell lines transfected with the inwardly rectifying K+ channel Kir6.1 alone or with both Kir6.1 and cystic fibrosis transmembrane conductance regulator (CFTR) exhibited time-independent K+ currents with weak inward rectification. In contrast, no measurable K+ conductance was observed in mock-transfected cells or in cells transfected with CFTR alone. Regardless of co-transfection with Kir6.1, the transfection with CFTR produced a Cl- conductance that was activated by cell dialysis with cAMP (1 mM). The conductance was reversibly suppressed by glibenclamide (30 microM). 3. Whole-cell currents at +60 mV were blocked in a concentration-dependent manner by Ba2+ ions with similar IC50 values: 89.3 +/- 23.3 microM (Kir6.1 alone) and 67.3 +/- 24.9 microM (Kir6.1-CFTR). 4. The currents recorded from Kir6. 1-transfected cells were not affected by glibenclamide, whereas glibenclamide did inhibit the conductance expressed in cells co-transfected with CFTR (IC50 = 35.9 +/- 6.6 microM). 5. In the cell-attached mode with a 150 mM K+ pipette solution, both Kir6.1- and Kir6.1-CFTR-transfected cells displayed a class of K+ channels showing weak inward rectification and a slope conductance of 50.7 +/- 1.0 and 52.4 +/- 4.9 pS, respectively. 6. In the inside-out mode, the single-channel currents recorded from both types of cells were not inhibited by intracellular ATP (1 mM). However, glibenclamide was found to block the single-channel activities in the co-transfected cells.
authors.affiliation: Department of Cardiovascular Medicine, Kyoto University Graduate School of Medicine, Kyoto 606-01, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Ishida-Takahashi
authors.forename: H
authors.initials: H
authors.lastname: Otani
authors.forename: C
authors.initials: C
authors.lastname: Takahashi
authors.forename: T
authors.initials: T
authors.lastname: Washizuka
authors.forename: K
authors.initials: K
authors.lastname: Tsuji
authors.forename: M
authors.initials: M
authors.lastname: Noda
authors.forename: M
authors.initials: M
authors.lastname: Horie
authors.forename: S
authors.initials: S
authors.lastname: Sasayama
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001464:Barium / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000096:biosynthesis / Q000502:physiology*; D005905:Glyburide / Q000494:pharmacology*; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000096:biosynthesis / Q000502:physiology*; D024661:Potassium Channels, Inwardly Rectifying; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D013997:Time Factors; D014162:Transfection
chemical_list: D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D001464:Barium; D000242:Cyclic AMP; D005905:Glyburide