Several cardiac diseases (e.g. heart failure, muscle dystrophy) are known to be associated with cellular oxidative stress. It is established that SR Ca2+ release channels (a.k.a. ryanodine receptors, RyRs) are susceptible to oxidation. Furthermore, our recent studies suggest that CICR and EC-coupling are sensitized in cardiomyocytes isolated from dystrophic mdx mice due to elevated levels of reactive oxygen species. The aim of this study was to examine the Ca2+ spark activity (as an indicator of RyR Ca2+ sensitivity) in mdx and wild-type (WT) cardiomyocytes at relevant redox potentials. Ventricular myocytes were permeabilized and exposed to solutions containing the Ca2+ indicator fluo-3 (50 μM) and a Ca2+ concentration of 50 nM. Ca2+ sparks were recorded with a laser-scanning confocal microscope in the line-scan mode and analyzed using SparkMaster software. Solutions mimicked intracellular redox potentials (EGSSG/GSH) determined in healthy hearts and in muscle dystrophy or heart failure, e.g. −226 mV and −217 mV. Under corresponding redox conditions the steady-state Ca2+ spark frequency did not show significant difference in mdx and WT cells (24±0.4 vs. 22±0.3 /100μms−1). Therefore, we used stronger reducing and oxidizing conditions to derive a redox/response relationship of spark parameters over a wider range of EGSSG/GSH from −263 mV to −146 mV. Under very oxidative conditions (EGSSG/GSH −146 mV) the spark frequency gradually declined but long-lasting Ca2+ release events appeared (> 70 ms, up to 700 ms) that were more frequent in mdx compared to WT cardiomyocytes (5.6 vs. 0.6 /100μms−1). Taken together, these results indicate that the average and modest change of the cytosolic redox potential may not significantly alter resting Ca2+ spark frequencies, but that stronger oxidative stress, as it has been reported to occur in subcellular regions as superoxide flashes, can lead to dramatic alterations of elementary Ca2+ signaling events.