Abstract 18314: Dysfunction of Junctional NCX Following JPH2 Knockdown Alters Spatial Properties of Ca 2+ Sparks and Contributes to Arrhythmogenesis in Mice

Abstract

Background. Na + /Ca 2+ exchanger (NCX) is essential in tuning intracellular Ca 2+ and the development of heart failure (HF). We reported that loss of Junctophilin 2 (JPH2) in adult mouse hearts leads to reduced NCX activity in addition to hyperactive RyR2s and more arrhythmogenic Ca 2+ waves. NCX has been found to co-localize with RyR2 recently. However, what is the physiological significance of junctional NCX and whether the reduced junctional NCX activity following JPH2 knockdown contributes to the arrhythmogenesis is still unknown. Hypothesis. Dysfunction of Junctional NCX in response to JPH2 knockdown alters spatial properties of Ca 2+ sparks and contributes to arrhythmogenesis. Methods. Tamoxifen-treated MCM and MCM-shJPH2 (KD) mice were studied. Contractile function was measured by echocardiography. Open probability of single RyR2 was evaluated by lipid bilayer. Ca 2+ spark were assessed with confocal microscopy. NCX activity was measured by exponential fitting of the declining phase of caffeine-induced Ca 2+ transient. Results. JPH2 knockdown increased spark size (MCM: 2.55 ± 0.14; KD: 4.25 ± 0.16, P < 0.001), suggesting an increased occurrence of proarrhythmic macrosparks. Cardiac Contractile function (EF% ) is negatively correlated with Ca 2+ spark size (n = 8, P < 0.05). Increased Open probability of RyR2 (MCM: 0.038 ± 0.018 KD: 0.274 ± 0.142, P < 0.01) is unlikely the reason for the ~70% increase in spark size, since it is not observed in 3 other mouse models with hyperactive RyR2s but normal JPH2 levels (RyR2:S2814D, mdx and FKBP12.6 -/- ). Loss of JPH2 also slows Ca 2+ efflux via NCX by ~50% (MCM: 2.70 ± 0.23 vs. KD: 5.11 ± 0.61 s, P < 0.01) without altering NCX expression level. Spark size and NCX activity are negatively correlated (n = 8 mice, P < 0.05), and partial inhibition (~30%) of NCX with 1 mM Cd 2+ significantly increased spark size by 14.0 ± 2.8% (n = 8, P <0.05) in wt mice. Conclusion. For the first time we showed that reduced NCX activity increases spatial properties of Ca 2+ sparks and thus arrhythmogenesis. Since reduced JPH2 is associated with HF and NCX plays more important role in human than in murine, reduced junctional NCX activity following loss of JPH2 could be a novel mechanism underlying the development of HF and associated ventricular arrhythmias.