Loop diuretics are open-channel blockers of the cystic fibrosis transmembrane conductance regulator with distinct kinetics

Min Ju, Toby S. Scott-Ward, J Liu, Pissared Khuituan, Hongyu Li, Zhiwei Cai, Stephen M. Husbands, David N. Sheppard*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

4 Citations (Scopus)

Abstract

Background and PurposeLoop diuretics are widely used to inhibit the Na+, K+, 2Cl(-) co-transporter, but they also inhibit the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel. Here, we investigated the mechanism of CFTR inhibition by loop diuretics and explored the effects of chemical structure on channel blockade.

Experimental ApproachUsing the patch-clamp technique, we tested the effects of bumetanide, furosemide, piretanide and xipamide on recombinant wild-type human CFTR.

Key ResultsWhen added to the intracellular solution, loop diuretics inhibited CFTR Cl- currents with potency approaching that of glibenclamide, a widely used CFTR blocker with some structural similarity to loop diuretics. To begin to study the kinetics of channel blockade, we examined the time dependence of macroscopic current inhibition following a hyperpolarizing voltage step. Like glibenclamide, piretanide blockade of CFTR was time and voltage dependent. By contrast, furosemide blockade was voltage dependent, but time independent. Consistent with these data, furosemide blocked individual CFTR Cl- channels with very fast' speed and drug-induced blocking events overlapped brief channel closures, whereas piretanide inhibited individual channels with intermediate' speed and drug-induced blocking events were distinct from channel closures.

Conclusions and ImplicationsStructure-activity analysis of the loop diuretics suggests that the phenoxy group present in bumetanide and piretanide, but absent in furosemide and xipamide, might account for the different kinetics of channel block by locking loop diuretics within the intracellular vestibule of the CFTR pore. We conclude that loop diuretics are open-channel blockers of CFTR with distinct kinetics, affected by molecular dimensions and lipophilicity.

Original languageEnglish
Pages (from-to)265-278
Number of pages14
JournalBritish Journal of Pharmacology
Volume171
Issue number1
DOIs
Publication statusPublished - Jan 2014

Keywords

  • ATP-binding cassette transporter
  • CFTR
  • chloride ion channel
  • open-channel blockade
  • epithelial ion transport
  • loop diuretics
  • NKCC
  • bumetanide
  • furosemide
  • glibenclamide
  • K-CL COTRANSPORTER
  • CFTR CHLORIDE CHANNEL
  • BUMETANIDE BINDING
  • NA+-K+-2CL(-) COTRANSPORTER
  • MULTIPLE SITES
  • CO-TRANSPORT
  • PORE
  • INHIBITION
  • GLIBENCLAMIDE
  • MECHANISM

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