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Recording of Ionic Currents Under Physiological Conditions: Action Potential-Clamp and ‘Onion-Peeling’ Techniques

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Modern Tools of Biophysics

Part of the book series: Handbook of Modern Biophysics ((HBBT,volume 5))

Abstract

Upon stimulation, excitable cells generate a transient change in the membrane potential called Action Potential (AP). The AP is governed by numerous ionic currents that flow in or out of the cell membrane. The goal of cellular electrophysiology is to understand the role of individual ionic currents and the interplay between currents in determining the profile and time course of AP. A critically important question of the field is how the ionic currents behave individually and interact collectively during the AP cycle of an excitable cell? To answer this question we need to know the dynamic behavior of ionic currents during AP and how these currents work in concert to determine the cell’s membrane potential at every moment.

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Author information

Authors and Affiliations

  1. Departments of Pharmacology, Biomedical Engineering, Internal Medicine/Cardiology, University of California, Davis, Davis, CA, 95616, USA

    Ye Chen-Izu Ph.D. & Tamás Bányász M.D., Ph.D.

  2. Department of Pharmacology, University of California, Davis, Davis, CA, 95616, USA

    Leighton T. Izu Ph.D. & Bence Hegyi M.D., Ph.D.

  3. Department of Physiology, University of Debrecen, Debrecen, 4012, Hungary

    Tamás Bányász M.D., Ph.D.

Authors
  1. Ye Chen-Izu Ph.D.
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  2. Leighton T. Izu Ph.D.
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  3. Bence Hegyi M.D., Ph.D.
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  4. Tamás Bányász M.D., Ph.D.
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Corresponding author

Correspondence to Ye Chen-Izu Ph.D. .

Editor information

Editors and Affiliations

  1. Department of Biochemistry and Molecular Medicine, University of California, Davis, Davis, California, USA

    Thomas Jue

Appendices

Problems

  1. 2.1.

    How is the cell’s action potential shaped by the inward currents and the outward currents?

  2. 2.2.

    What are the advantages of using AP-clamp and ‘Onion-Peeling’ technique, instead of conventional voltage-clamp, to record ionic currents?

  3. 2.3.

    Does Ca2+ transient during the action potential cycle affect ionic currents and the action potential? How to preserve the Ca2+ cycling during action potential?

  4. 2.4.

    What are the basic requirements for performing successful AP-clamp experiments?

Solutions

  1. 2.1

    The inward currents provide the Depolarization and the outward currents provide the Repolarization; these opposing currents counterbalance to shape the cell’s action.

  2. 2.2

    AP-clamp recording provides a different type of data from conventional voltage clamp: (a) the ionic currents are directly recorded during AP with Ca2+ cycling in a physiological milieu, mimicking in vivo condition; (b) the unprecedented ability to measure multiple currents in the same cell allows deciphering their relationships in the same cell, which is necessary for studying how multiple currents interact and integrate in the single cell to shape APs.

  3. 2.3
    1. 1.

      Not adding any exogenous Ca2+ buffer;

    2. 2.

      Using the cell’s own steady-state AP as the voltage-clamp command waveform;

    3. 3.

      Using internal and external solutions having physiological ionic composition;

    4. 4.

      Using physiological stimulation frequency and body temperature.

  4. 2.4
    1. 1.

      High-quality isolated single cells that can reach steady-state AP;

    2. 2.

      Using the internal and external solutions with physiological ionic composition, pH, and osmolarity;

    3. 3.

      Making the whole-cell seal with low access resistance (<5 MΩ) and seal condition kept constant throughout the entire experiment;

    4. 4.

      Using highly specific channel blockers to obtain drug-sensitive current.

Further Study

  • The recent advances in patch-clamp techniques and their application in cardiac electrophysiology are summarized by Bébarová [67].

  • How AP‐clamp can be used to test the selectivity of drugs acting on cardiac ion channels are reviewed by Szentandrássy et al. [68].

  • AP-clamp studies can reveal the exact frequency-dependent properties of ionic currents under AP [69].

  • Dynamic clamp can be used to assess ionic current properties during action potentials with beat-to-beat variability [70].

  • Dynamic clamp can be used to study the consequences of cardiac ion channel mutations in causing inherited arrhythmias [22, 71].

  • Dynamic clamp used in the field of cardiac stem cell research [72].

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Chen-Izu, Y., Izu, L.T., Hegyi, B., Bányász, T. (2017). Recording of Ionic Currents Under Physiological Conditions: Action Potential-Clamp and ‘Onion-Peeling’ Techniques. In: Jue, T. (eds) Modern Tools of Biophysics. Handbook of Modern Biophysics, vol 5. Springer, New York, NY. https://doi.org/10.1007/978-1-4939-6713-1_2

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