Tightly regulated Ca2+ homeostasis is a prerequisite for proper cardiac function. To dissect the regulatory network of cardiac Ca2+ handling, we performed a chemical suppressor screen on zebrafish tremblor embryos, which suffer from Ca2+ extrusion defects. Efsevin was identified based on its potent activity to restore coordinated contractions in tremblor. We show that efsevin binds to VDAC2, potentiates mitochondrial Ca2+ uptake and accelerates the transfer of Ca2+ from intracellular stores into mitochondria. In cardiomyocytes, efsevin restricts the temporal and spatial boundaries of Ca2+ sparks and thereby inhibits Ca2+ overload-induced erratic Ca2+ waves and irregular contractions. We further show that overexpression of VDAC2 recapitulates the suppressive effect of efsevin on tremblor embryos whereas VDAC2 deficiency attenuates efsevin's rescue effect and that VDAC2 functions synergistically with MCU to suppress cardiac fibrillation in tremblor. Together, these findings demonstrate a critical modulatory role for VDAC2-dependent mitochondrial Ca2+ uptake in the regulation of cardiac rhythmicity.DOI: http://dx.doi.org/10.7554/eLife.04801.001

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Compelling reports suggest that intra-mitochondrial [Ca 2þ ] mito changes dramatically with intracellular [Ca 2þ ] i transients (1) while other equally compelling investigations suggest there are little to no changes (2). Here we take advantage of the high temporal and spatial resolution of confocal microscopy imaging to examine this question in rat and rabbit ventricular cardiomyocytes. Using a mitochondrially targeted Ca 2þ -sensitive fluorescent protein ''Mitycam'' (3), we examined the time course of changes in [Ca 2þ ] mito (matrix mitochondrial [Ca 2þ ]) in isolated ventricular myocytes. Examination was performed 48 hours after treatment with adenovirusdependent Mitycam expression in ventricular myocytes in primary culture. Mitycam, which co-localizes with mitochondrial-specific markers (e.g. Mito-Tracker red), responds slowly (minutes) to steep changes of [Ca 2þ ] i in saponin permeabilized myocytes ([Ca 2þ ] i << 10 to 250 nM). Mitycam responds more quickly (~1 s) in intact (i.e. non-permeabilized) myocytes where a large change in [Ca 2þ ] i (from~100 nM to~10 mM) is seen following caffeine (10 mM). However, in these cells, physiologic [Ca 2þ ] i transients produced no significant increase in [Ca 2þ ] mito in a beat-to-beat manner but can modestly affect the timeaveraged [Ca 2þ ] mito following changes in heart rate ([Ca 2þ ] mito responds to changes in [Ca 2þ ] i like a low-pass-filter). In contrast, isolated Mitycamcontaining mitochondria respond to changes in [Ca 2þ ]in milliseconds. We conclude that the cardiac [Ca 2þ ] i transient does not significantly change [Ca 2þ ] mito in cardiac myocytes in a beat-to-beat manner but slowly influences the time-averaged [Ca 2þ ] mito signal.

No abstract

Tightly regulated Ca 2+ homeostasis is a prerequisite for proper cardiac function. To dissect the regulatory network of cardiac Ca 2+ handling, we performed a chemical suppressor screen on zebrafish tremblor embryos, which suffer from Ca 2+ extrusion defects. Efsevin was identified based on its potent activity to restore coordinated contractions in tremblor. We show that efsevin binds to VDAC2, potentiates mitochondrial Ca 2+ uptake and accelerates the transfer of Ca 2+ from intracellular stores into mitochondria. In cardiomyocytes, efsevin restricts the temporal and spatial boundaries of Ca 2+ sparks and thereby inhibits Ca 2+ overload-induced erratic Ca 2+ waves and irregular contractions. We further show that overexpression of VDAC2 recapitulates the suppressive effect of efsevin on tremblor embryos whereas VDAC2 deficiency attenuates efsevin's rescue effect and that VDAC2 functions synergistically with MCU to suppress cardiac fibrillation in tremblor. Together, these findings demonstrate a critical modulatory role for VDAC2-dependent mitochondrial Ca 2+ uptake in the regulation of cardiac rhythmicity.