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Related Experiment Videos

Visualization of Ca2+ entry through single stretch-activated cation channels.

Hui Zou1, Lawrence M Lifshitz, Richard A Tuft

  • 1Department of Physiology and Biomedical Imaging Group, University of Massachusetts Medical School, Worcester, MA 01655, USA. imaging.ionchannels@umassmed.edu

Proceedings of the National Academy of Sciences of the United States of America
|May 2, 2002
PubMed
Summary

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Stretch-activated channels (SACs) allow significant calcium (Ca2+) entry into smooth muscle cells. This Ca2+ influx through SACs can activate other channels and intracellular stores.

Area of Science:

  • Cell Physiology
  • Biophysics
  • Smooth Muscle Biology

Background:

  • Stretch-activated channels (SACs) are present in smooth muscle and implicated in myogenic responses.
  • Previous studies showed SACs are Ca2+ permeable only when Ca2+ is the sole charge carrier.
  • Direct evidence for significant Ca2+ passage through SACs in physiological conditions was lacking.

Purpose of the Study:

  • To provide direct evidence of significant Ca2+ influx through SACs in physiological solutions.
  • To quantify the contribution of Ca2+ to SAC current in smooth muscle cells.

Main Methods:

  • Utilized high-resolution imaging of single-channel Ca2+ fluorescence transients (SCCaFT).
  • Simultaneously recorded SAC currents from cell-attached patches on smooth muscle cells.

Related Experiment Videos

  • Loaded cells with fluo-3 acetoxymethyl ester and recorded fluorescence with a wide-field digital imaging microscope.

    • Main Results:

    • Directly visualized Ca2+ entry through single SAC openings, increasing local Ca2+ concentration.
    • Observed fluorescence increases preceding global Ca2+ increases mediated by voltage-gated Ca2+ channels.
    • Determined that approximately 18% of SAC current is carried by Ca2+ at negative membrane potentials.

    Conclusions:

    • Demonstrated significant Ca2+ permeability of SACs under physiological conditions.
    • Quantified unitary Ca2+ currents through SACs, sufficient to activate other channels and intracellular stores.
    • Established a novel method for studying Ca2+ flux through single ion channels in real-time.