Abstract P1101: Mitochondrial Ca Uniporter Complex, ROS And CaT Alternans

Abstract

Background: Atrial calcium transient (CaT) alternans is defined as beat-to-beat alternation in CaT amplitude and is causally linked to atrial fibrillation (AF). Inhibition of mitochondrial Ca uptake via the mitochondrial Ca uniporter complex (MCUC) facilitates CaT alternans and can lead to an increase in mitochondrial reactive oxygen species(ROS) production that compromises the cellular antioxidative capacity. The objective was to determine the mechanisms through which mitochondrial Ca uptake and ROS production determine the susceptibility of atrial cells to develop CaT alternans. Methods: CaT alternans was measured in single rabbit left atrial myocytes with fluorescence microscopy using the fluorescent Ca indicator Cal-520/AM, and for simultaneous measurements of ROS and alternans, cells were loaded with CM-DCFDA and Cal-590/AM. CaT alternans was induced by increasing the pacing frequency until stable CaT alternans was observed. The degree of alternans was quantified as the alternans ratio (AR = 1 – S/L, where S/L is the ratio of the small to the large amplitude of a pair of alternating CaTs). Results: Exposure of LA myocytes to the specific MCUC inhibitor Ru360 increased the AR by 86%. However, the subsequent stimulation of mitochondrial Ca uptake with the MCUC activator spermine (Spm) decreased AR by 79% (p=0.028). Quercetin (a O2 .- scavenger) and Tempol (a superoxide dismutase mimetic) lowered CaT AR by 96% (p=0.0007) and 79% (p=0.0029), respectively. An oxidative challenge with tert-butylhydroperoxide (tBHP) increased AR by 140% (p=<0.0001), and the reducing agent dithiothreitol (DTT) decreased AR by 39% (p=0.0194). Moreover, DTT treatment rescued CaT alternans previously enhanced by tBHP. During CaT alternans the rate of ROS production increased by 23% (p=0.026), however Spm treatment attenuated the increased ROS production. Conclusion: An oxidizing environment facilitates the development of CaT alternans. During CaT alternans mitochondrial ROS formation is increased and involves MCUC activity.