Properties of single-channel and whole cell CL- currents in Guinea pig detrusor smooth muscle cells

Viktor Yarotskyy, John Malysz, Georgi Petkov

Research output: Contribution to journalArticle

Abstract

Properties of single-channel and whole cell Cl+ currents in guinea pig detrusor smooth muscle cells. Am J Physiol Cell Physiol 316: C698–C710, 2019. First published December 19, 2018; doi:10.1152/ajpcell.00327.2018.—Multi-ple types of Cl- channels regulate smooth muscle excitability and contractility in vascular, gastrointestinal, and airway smooth muscle cells. However, little is known about Cl- channels in detrusor smooth muscle (DSM) cells. Here, we used inside-out single channel and whole cell patch-clamp recordings for detailed biophysical and pharmacological characterizations of Cl- channels in freshly isolated guinea pig DSM cells. The recorded single Cl- channels displayed unique gating with multiple subconductive states, a fully opened single-channel conductance of 164 pS, and a reversal potential of 41.5 mV, which is close to the ECl of 65 mV, confirming preferential permeability to Cl-. The Cl- channel demonstrated strong voltage dependence of activation (half-maximum of mean open probability, V0.5,~20 mV) and robust prolonged openings at depolarizing voltages. The channel displayed similar gating when exposed intracellularly to solutions containing Ca2+-free or 1 mM Ca2+.In whole cell patch-clamp recordings, macroscopic current demonstrated outward rectification, inhibitions by 4,4=-diisothiocyano-2,2=-stilbene-disulfonic acid (DIDS) and niflumic acid, and insensitivity to chlorotoxin. The outward current was reversibly reduced by 94% replacement of extracellular Cl- with I-,Br-, or methanesulfonate (MsO-), resulting in anionic permeability sequence: Cl->Br->I->MsO-. While intracellular Ca2+ levels (0, 300 nM, and 1 mM) did not affect the amplitude of Cl- current and outward rectification, high Ca2+ slowed voltage-step current activation at depolarizing voltages. In conclusion, our data reveal for the first time the presence of a Ca2+-independent DIDS and niflumic acid-sensitive, voltage-dependent Cl- channel in the plasma membrane of DSM cells. This channel may be a key regulator of DSM excitability.

Original languageEnglish (US)
Pages (from-to)C698-C710
JournalAmerican Journal of Physiology - Cell Physiology
Volume316
Issue number5
DOIs
StatePublished - May 1 2019

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Smooth Muscle Myocytes
Guinea Pigs
Niflumic Acid
Stilbenes
Smooth Muscle
Permeability
Acids
Blood Vessels
Cell Membrane
Pharmacology

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cell Biology

Cite this

Properties of single-channel and whole cell CL- currents in Guinea pig detrusor smooth muscle cells. / Yarotskyy, Viktor; Malysz, John; Petkov, Georgi.

In: American Journal of Physiology - Cell Physiology, Vol. 316, No. 5, 01.05.2019, p. C698-C710.

Research output: Contribution to journalArticle

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abstract = "Properties of single-channel and whole cell Cl+ currents in guinea pig detrusor smooth muscle cells. Am J Physiol Cell Physiol 316: C698–C710, 2019. First published December 19, 2018; doi:10.1152/ajpcell.00327.2018.—Multi-ple types of Cl- channels regulate smooth muscle excitability and contractility in vascular, gastrointestinal, and airway smooth muscle cells. However, little is known about Cl- channels in detrusor smooth muscle (DSM) cells. Here, we used inside-out single channel and whole cell patch-clamp recordings for detailed biophysical and pharmacological characterizations of Cl- channels in freshly isolated guinea pig DSM cells. The recorded single Cl- channels displayed unique gating with multiple subconductive states, a fully opened single-channel conductance of 164 pS, and a reversal potential of 41.5 mV, which is close to the ECl of 65 mV, confirming preferential permeability to Cl-. The Cl- channel demonstrated strong voltage dependence of activation (half-maximum of mean open probability, V0.5,~20 mV) and robust prolonged openings at depolarizing voltages. The channel displayed similar gating when exposed intracellularly to solutions containing Ca2+-free or 1 mM Ca2+.In whole cell patch-clamp recordings, macroscopic current demonstrated outward rectification, inhibitions by 4,4=-diisothiocyano-2,2=-stilbene-disulfonic acid (DIDS) and niflumic acid, and insensitivity to chlorotoxin. The outward current was reversibly reduced by 94{\%} replacement of extracellular Cl- with I-,Br-, or methanesulfonate (MsO-), resulting in anionic permeability sequence: Cl->Br->I->MsO-. While intracellular Ca2+ levels (0, 300 nM, and 1 mM) did not affect the amplitude of Cl- current and outward rectification, high Ca2+ slowed voltage-step current activation at depolarizing voltages. In conclusion, our data reveal for the first time the presence of a Ca2+-independent DIDS and niflumic acid-sensitive, voltage-dependent Cl- channel in the plasma membrane of DSM cells. This channel may be a key regulator of DSM excitability.",
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