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Resurgent-like currents in mouse vas deferens myocytes are mediated by NaV1.6 voltage-gated sodium channels

  • Muscle physiology
  • Published:
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Abstract

Patch-clamp experiments were performed to investigate the molecular properties of resurgent-like currents in single smooth muscle cells dispersed from mouse vas deferens, utilizing both NaV1.6-null mice (NaV1.6−/−), lacking the expression of the Scn8a Na+ channel gene, and their wild-type littermates (NaV1.6+/+). NaV1.6 immunoreactivity was clearly visible in dispersed smooth muscle cells obtained from NaV1.6+/+, but not NaV1.6−/−, vas deferens. Following a depolarization to +30 mV from a holding potential of −70 mV (to produce maximal inactivation of the Na+ current), repolarization to voltages between −60 and +20 mV elicited a tetrodotoxin (TTX)-sensitive inward current in NaV1.6+/+, but not NaV1.6−/−, vas deferens myocytes. The resurgent-like current in NaV1.6+/+ vas deferens myocytes peaked at approximately −20 mV in the current–voltage relationship. The peak amplitude of the resurgent-like current remained at a constant level when the membrane potential was repolarized to −20 mV following the application of depolarizing rectangular pulses to more positive potentials than +20 mV. 4,9-Anhydrotetrodotoxin (4,9-anhydroTTX), a selective NaV1.6 blocking toxin, purified from a crude mixture of TTX analogues by LC-FLD techniques, reversibly suppressed the resurgent-like currents. β-Pompilidotoxin, a voltage-gated Na+ channel activator, evoked sustained resurgent-like currents in NaV1.6+/+ but not NaV1.6−/− murine vas deferens myocytes. These results strongly indicate that, primarily, resurgent-like currents are generated as a result of NaV1.6 channel activity.

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Abbreviations

4,9-anhydroTTX:

4,9-Anhydrotetrodotoxin

BSA:

Bovine serum albumin

DAPI:

4′,6-Diamidino-2-phenylindole

EJPs:

Excitatory junction potentials

HEPES:

N-[2-Hydroxyethyl]piperazine-N′-[2-ethanesulfonic acid]

I Na :

Transient voltage-gated Na+ currents

LC-FLD:

Liquid chromatography-fluorescent detection

med :

Motor end-plate disease

NaV channel:

Voltage-gated Na+ channel

NMDG+ :

N-Methyl- d-glucamine

PBS:

Phosphate-buffered saline

PSS:

Physiological salt solution

TEA:

Tetraethylammonium

Tris:

Tris(hydroxymethyl)methylammonium chloride

Triton-X:

Polyoxyethylene-p-isooctylphenol

TTX:

Tetrodotoxin

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Acknowledgments

This work was supported by a Funding Program for Next Generation World-Leading Researcher (Noriyoshi Teramoto, Grant Number LS096; Mari Yotsu-Yamashita, Grant Number LS012) from the Japanese Society for the Promotion of Science. We thank Dr. Stevan Rakovic (OxProfs, Oxford, UK) for his helpful discussion and critical reading of the manuscript.

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Authors and Affiliations

  1. Department of Pharmacology, Faculty of Medicine, Saga University, Saga, 849-8501, Japan

    Noriyoshi Teramoto & Hai-Lei Zhu

  2. Laboratory of Bioorganic Chemistry of Natural Products, Graduate School of Agricultural Science, Tohoku University, Sendai, 981-8555, Japan

    Mari Yotsu-Yamashita

  3. Department of Morphological Biology, Division of Biomedical Sciences, Fukuoka Dental College, Fukuoka, 814-0193, Japan

    Tetsuichiro Inai

  4. Department of Pharmacology, University of Oxford, Oxford, OX1 3QT, UK

    Thomas C. Cunnane

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  1. Noriyoshi Teramoto
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  2. Hai-Lei Zhu
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  3. Mari Yotsu-Yamashita
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  4. Tetsuichiro Inai
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  5. Thomas C. Cunnane
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Correspondence to Noriyoshi Teramoto.

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Teramoto, N., Zhu, HL., Yotsu-Yamashita, M. et al. Resurgent-like currents in mouse vas deferens myocytes are mediated by NaV1.6 voltage-gated sodium channels. Pflugers Arch - Eur J Physiol 464, 493–502 (2012). https://doi.org/10.1007/s00424-012-1153-4

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  • DOI: https://doi.org/10.1007/s00424-012-1153-4

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