Ca2+ spark-dependent and -independent sarcoplasmic reticulum Ca2+ leak in normal and failing rabbit ventricular myocytes

Authors


Corresponding author A. V. Zima: Department of Cell and Molecular Physiology, Loyola University Chicago, Stritch School of Medicine, 2160 South First Avenue, Maywood, IL 60153, USA.  Email: azima@lumc.edu

Abstract

Sarcoplasmic reticulum (SR) Ca2+ leak is an important component of cardiac Ca2+ signalling. Together with the SR Ca2+-ATPase (SERCA)-mediated Ca2+ uptake, diastolic Ca2+ leak determines SR Ca2+ load and, therefore, the amplitude of Ca2+ transients that initiate contraction. Spontaneous Ca2+ sparks are thought to play a major role in SR Ca2+ leak. In this study, we determined the quantitative contribution of sparks to SR Ca2+ leak and tested the hypothesis that non-spark mediated Ca2+ release also contributes to SR Ca2+ leak. We simultaneously measured spark properties and intra-SR free Ca2+ ([Ca2+]SR) after complete inhibition of SERCA with thapsigargin in permeabilized rabbit ventricular myocytes. When [Ca2+]SR declined to 279 ± 10 μm, spark activity ceased completely; however SR Ca2+ leak continued, albeit at a slower rate. Analysis of sparks and [Ca2+]SR revealed, that SR Ca2+ leak increased as a function of [Ca2+]SR, with a particularly steep increase at higher [Ca2+]SR (>600 μm) where sparks become a major pathway of SR Ca2+ leak. At low [Ca2+]SR (<300 μm), however, Ca2+ leak occurred mostly as non-spark-mediated leak. Sensitization of ryanodine receptors (RyRs) with low doses of caffeine increased spark frequency and SR Ca2+ leak. Complete inhibition of RyR abolished sparks and significantly decreased SR Ca2+ leak, but did not prevent it entirely, suggesting the existence of RyR-independent Ca2+ leak. Finally, we found that RyR-mediated Ca2+ leak was enhanced in myocytes from failing rabbit hearts. These results show that RyRs are the main, but not sole contributor to SR Ca2+ leak. RyR-mediated leak occurs in part as Ca2+ sparks, but there is clearly RyR-mediated but Ca2+ sparks independent leak.

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