Unitary Ca2+ Current through Cardiac Ryanodine Receptor Channels under Quasi-Physiological Ionic Conditions |
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Authors: | Rafael Mejía-Alvarez Claudia Kettlun Eduardo Ríos Michael Stern Michael Fill |
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Institution: | From the *Department of Physiology, Stritch School of Medicine, Loyola University Chicago, Maywood, Illinois 60153; ‡Department of Molecular Biophysics and Physiology, Rush University School of Medicine, Chicago, Illinois 60612; and §Laboratory of Cardiovascular Science, Gerontology, Research Center, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21214 |
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Abstract: | Single canine cardiac ryanodine receptor channels were incorporated into planar lipid bilayers. Single-channel currents were sampled at 1–5 kHz and filtered at 0.2–1.0 kHz. Channel incorporations were obtained in symmetrical solutions (20 mM HEPES-Tris, pH 7.4, and pCa 5). Unitary Ca2+ currents were monitored when 2–30 mM Ca2+ was added to the lumenal side of the channel. The relationship between the amplitude of unitary Ca2+ current (at 0 mV holding potential) and lumenal Ca2+] was hyperbolic and saturated at ∼4 pA. This relationship was then defined in the presence of different symmetrical CsCH3SO3 concentrations (5, 50, and 150 mM). Under these conditions, unitary current amplitude was 1.2 ± 0.1, 0.65 ± 0.1, and 0.35 ± 0.1 pA in 2 mM lumenal Ca2+; and 3.3 ± 0.4, 2.4 ± 0.2, and 1.63 ± 0.2 pA in 10 mM lumenal Ca2+ (n > 6). Unitary Ca2+ current was also defined in the presence of symmetrical Mg2+] (1 mM) and low Cs+] (5 mM). Under these conditions, unitary Ca2+ current in 2 and 10 mM lumenal Ca2+ was 0.66 ± 0.1 and 1.52 ± 0.06 pA, respectively. In the presence of higher symmetrical Cs+] (50 mM), Mg2+ (1 mM), and lumenal Ca2+] (10 mM), unitary Ca2+ current exhibited an amplitude of 0.9 ± 0.2 pA (n = 3). This result indicates that the actions of Cs+ and Mg2+ on unitary Ca2+ current were additive. These data demonstrate that physiological levels of monovalent cation and Mg2+ effectively compete with Ca2+ as charge carrier in cardiac ryanodine receptor channels. If lumenal free Ca2+ is 2 mM, then our results indicate that unitary Ca2+ current under physiological conditions should be <0.6 pA. |
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Keywords: | Ca2+ release sarcoplasmic reticulum Ca2+ spark excitation– contraction coupling planar bilayers |
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