Pflügers Archiv - European Journal of Physiology

, Volume 460, Issue 3, pp 633–644

Control of volume-sensitive chloride channel inactivation by the coupled action of intracellular chloride and extracellular protons

  • Carmen Y. Hernández-Carballo
  • José A. De Santiago-Castillo
  • Teresa Rosales-Saavedra
  • Patricia Pérez-Cornejo
  • Jorge Arreola
Ion Channels, Receptors and Transporters

Abstract

The volume-sensitive chloride current (IClVol) exhibit a time-dependent decay presumably due to channel inactivation. In this work, we studied the effects of chloride ions (Cl) and H+ ions on IClVol decay recorded in HEK-293 and HL-60 cells using the whole-cell patch clamp technique. Under control conditions ([Cl]e = [Cl]i = 140 mM and pHi = pHe = 7.3), IClVol in HEK cells shows a large decay at positive voltages but in HL-60 cells IClVol remained constant independently of time. In HEK-293 cells, simultaneously raising the [Cl]e and [Cl]i from 25 to 140 mM (with pHe = pHi = 7.3) increased the fraction of inactivated channels (FIC). This effect was reproduced by elevating [Cl]i while keeping the [Cl]e constant. Furthermore, a decrease in pHe from 7.3 to 5.5 accelerated current decay and increased FIC when [Cl] was 140 mM but not 25 mM. In HL-60 cells, a slight IClVol decay was seen when the pHe was reduced from 7.3 to 5.5. Our data show that inactivation of IClVol can be controlled by changing either the Cl or H+ concentration or both. Based on our results and previously published data, we have built a model that explains VRAC inactivation. In the model the H+ binding site is located outside the electrical field near the extracellular entry whilst the Cl binding site is intracellular. The model depicts inactivation as a pore constriction that happens by simultaneous binding of H+ and Cl ions to the channel followed by a voltage-dependent conformational change that ultimately causes inactivation.

Keywords

Volume regulation Cl channels Inactivation pH dependence Chloride 

Supplementary material

424_2010_842_MOESM1_ESM.ppt (153 kb)
Supplemental Fig. 1Hypothetical Vm-dependence of volume-sensitive chloride channels. a Simple barrier model representing the energy landscape along the VRAC pore. This energy profile plus the kinetic model shown in Fig. 7 were used to qualitatively explain the Vm (a) and external Cl-dependence (b) of inactivation. The energy profiles depict the landscape along the pore that the permeant anions (with symmetrical [Cl]i = [Cl]e = 140 mM) experiment at −100, 0, and +100 mV. At each voltage, the pore occupancy changes and thus the probability that the pore is empty (PU) is greater at positive voltages. Moreover, PU changes as a function of the Cl gradient across the membrane. bPU becomes larger as the Cl gradient decreases. PU was calculated using the Vm-dependent rate constants \( {\alpha_{\rm{V}}} = {\left[ {{\hbox{C}}{{\hbox{l}}^{-} }} \right]_{\rm{o}}} \times {k_1} + {\left[ {{\hbox{C}}{{\hbox{l}}^{-} }} \right]_{\rm{i}}} \times {k_{ - {2}}}\;{\hbox{and}}\;{\beta_{\rm{V}}} = {k_{ - {1} + }} \times {k_{ - {2}}} \) recorded in Table 2 (PPT 153 kb)

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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Carmen Y. Hernández-Carballo
    • 1
  • José A. De Santiago-Castillo
    • 2
  • Teresa Rosales-Saavedra
    • 2
  • Patricia Pérez-Cornejo
    • 2
  • Jorge Arreola
    • 1
  1. 1.Instituto de FísicaUniversidad Autónoma de San Luis PotosíSan Luis PotosíMéxico
  2. 2.Facultad de MedicinaUniversidad Autónoma de San Luis PotosíSan Luis PotosíMéxico

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