Decomposition of a Calcium Spark in Cardiac Myocytes

Abstract

Ca2+ sparks are the elementary SR Ca2+ release events in heart muscle. Spontaneous and triggered Ca2+ sparks are usually centered on a junctional sarcoplasmic reticulum (jSR) across a 15 nm subspace gap from a transverse tubule (TT) at the Z-disk. The jSR-TT junction contains ryanodine receptor (RyR2) clusters of a variable number of RyR2s and diverse organizations. To determine if Ca2+ sparks may reveal a substructure that reflects the jSR-RyR2 organizational variability, we have examined rabbit ventricular myocytes and imaged [Ca2+] in living myocytes using a confocal microscope. The cytosol was loaded with the [Ca2+]i indicator rhod-2 and the SR loaded with the low affinity [Ca2+]SR indicator fluo-5N. This method has allowed the detection of Ca2+ blinks (the local SR Ca2+ depletion during a Ca2+ spark) and Ca2+ sparks simultaneously. Furthermore, this method makes it possible to image the recently described set of small, sub-spark events, the quarky SR Ca2+ release or QCR. The spatial profiles of Ca2+ sparks and Ca2+ blinks has thus revealed a sub-structure within a single Ca2+ spark. The images suggest that Ca2+ sparks could be composed of a central Ca2+ release site with surrounding small release site elements (RS-elements). These RS-elements may be asymmetrically distributed around the central site. That high concentrations of EGTA (30 μM) prevented the activation of the RS-elements suggests that these RS-elements or QCRs are activated by Ca2+-induced Ca2+-release (CICR) triggered from the central site. How the underlying structures may change in diverse diseases may be important in broadening our understanding of both arrhythmogenesis and heart failure, diseases widely associated with altered Ca2+ signaling, [Ca2+]i stability and altered cellular ultrastructure.