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Circ Res. 2016 Jan 22;118(2):203-15. doi: 10.1161/CIRCRESAHA.115.307399. Epub 2015 Nov 12.

Intact Heart Loose Patch Photolysis Reveals Ionic Current Kinetics During Ventricular Action Potentials.

Author information

1
From the Department of Molecular Biophysics and Physiology, Rush University Medical Center, Chicago, IL (J.R.-F.); and Quantitative Systems Biology Program, School of Natural Sciences (Y.A.-S.) and Biological Engineering and Small Scale Technologies Program, School of Engineering (A.L.E.), University of California, Merced, CA.
2
From the Department of Molecular Biophysics and Physiology, Rush University Medical Center, Chicago, IL (J.R.-F.); and Quantitative Systems Biology Program, School of Natural Sciences (Y.A.-S.) and Biological Engineering and Small Scale Technologies Program, School of Engineering (A.L.E.), University of California, Merced, CA. aescobar4@ucmerced.edu.

Abstract

RATIONALE:

Assessing the underlying ionic currents during a triggered action potential (AP) in intact perfused hearts offers the opportunity to link molecular mechanisms with pathophysiological problems in cardiovascular research. The developed loose patch photolysis technique can provide striking new insights into cardiac function at the whole heart level during health and disease.

OBJECTIVE:

To measure transmembrane ionic currents during an AP to determine how and when surface Ca(2+) influx that triggers Ca(2+)-induced Ca(2+) release occurs and how Ca(2+)-activated conductances can contribute to the genesis of AP phase 2.

METHODS AND RESULTS:

Loose patch photolysis allows the measurement of transmembrane ionic currents in intact hearts. During a triggered AP, a voltage-dependent Ca(2+) conductance was fractionally activated (dis-inhibited) by rapidly photo-degrading nifedipine, the Ca(2+) channel blocker. The ionic currents during a mouse ventricular AP showed a fast early component and a slower late component. Pharmacological studies established that the molecular basis underlying the early component was driven by an influx of Ca(2+) through the L-type channel, CaV 1.2. The late component was identified as an Na(+)-Ca(2+) exchanger current mediated by Ca(2+) released from the sarcoplasmic reticulum.

CONCLUSIONS:

The novel loose patch photolysis technique allowed the dissection of transmembrane ionic currents in the intact heart. We were able to determine that during an AP, L-type Ca(2+) current contributes to phase 1, whereas Na(+)-Ca(2+) exchanger contributes to phase 2. In addition, loose patch photolysis revealed that the influx of Ca(2+) through L-type Ca(2+) channels terminates because of voltage-dependent deactivation and not by Ca(2+)-dependent inactivation, as commonly believed.

KEYWORDS:

action potentials; calcium signaling; excitation contraction coupling; ionic currents; nifedipine; sarcoplasmic reticulum

PMID:
26565013
PMCID:
PMC4851170
DOI:
10.1161/CIRCRESAHA.115.307399
[Indexed for MEDLINE]
Free PMC Article

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