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Time-dependent Interactions of Glibenclamide with CFTR: Kinetically Complex Block of Macroscopic Currents

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Abstract

Blockade of the CFTR chloride channel by glibenclamide was studied in Xenopus oocytes using two-electrode voltage-clamp recordings, macropatch recordings, and summations of single-channel currents, in order to test a kinetic model recently developed by us from single-channel experiments. Both the forward and reverse macroscopic reactions, at negative and positive membrane potential VM, respectively, were slow in comparison to those reactions for other CFTR pore blockers such as DPC and NPPB, resulting in prominent relaxations on the order of tens of milliseconds. The rate of the reverse reaction was voltage-dependent, and dependent on the Cl− driving force, while that of the forward reaction was not. In inside-out macropatches, block and relief from block occurred in two distinct phases that differed in apparent affinity. The results are consistent with the presence of multiple glibenclamide binding sites in CFTR, with varying affinity and voltage dependence; they support the kinetic model and suggest experimental approaches for identification of those sites by mutagenesis.

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Acknowledgements

This work was supported by the American Heart Association (9820032SE), the Cystic Fibrosis Foundation (MCCART00P0), and the NIH (DK056481). S. Zeltwanger was supported by an NIH postdoctoral training grant (DK07656). N.A.M. is an Established Investigator of the American Heart Association.

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Authors and Affiliations

  1. School of Biology, Georgia Institute of Technology, Atlanta, GA, 30332-0230, USA

    Z.-R. Zhang, G. Cui & N.A. McCarty

  2. Department of Physiology, Emory University School of Medicine, Atlanta, GA, 30322, USA

    S. Zeltwanger

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  1. Z.-R. Zhang
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  2. G. Cui
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  3. S. Zeltwanger
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  4. N.A. McCarty
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Correspondence to N.A. McCarty.

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Zhang, ZR., Cui, G., Zeltwanger, S. et al. Time-dependent Interactions of Glibenclamide with CFTR: Kinetically Complex Block of Macroscopic Currents. J Membrane Biol 201, 139–155 (2004). https://doi.org/10.1007/s00232-004-0712-9

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  • DOI: https://doi.org/10.1007/s00232-004-0712-9

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