Phospholamban ablation rescues the enhanced propensity to arrhythmias of mice with CaMKII‐constitutive phosphorylation of RyR2 at site S2814. (2nd February 2016)

• Record Type: Journal Article
• Authors: Mazzocchi, G.; Sommese, L.; Palomeque, J.; Felice, J. I.; Di Carlo, M. N.; Fainstein, D.; Gonzalez, P.; Contreras, P.; Skapura, D.; McCauley, M. D.; Lascano, E. C.; Negroni, J. A.; Kranias, E. G.; Wehrens, X. H. T.; Valverde, C. A.; Mattiazzi, A.
• Is Part Of: Journal of physiology. Volume 594:Number 11(2016:Jun.)
• Page Start: 3005
• Page End: 3030
• Publication Date: 2016-02-02
• Subjects: Physiology -- Periodicals; 612.005
• DOI: 10.1113/JP271622
• Languages: English
• ISSNs: 0022-3751

Abstract

Abstract : Key points: Mice with Ca 2+ –calmodulin‐dependent protein kinase (CaMKII) constitutive pseudo‐phosphorylation of the ryanodine receptor RyR2 at Ser2814 (S2814D +/+ mice) exhibit a higher open probability of RyR2, higher sarcoplasmic reticulum (SR) Ca 2+ leak in diastole and increased propensity to arrhythmias under stress conditions. We generated phospholamban (PLN)‐deficient S2814D +/+ knock‐in mice by crossing two colonies, S2814D +/+ and PLNKO mice, to test the hypothesis that PLN ablation can prevent the propensity to arrhythmias of S2814D +/+ mice. PLN ablation partially rescues the altered intracellular Ca 2+ dynamics of S2814D +/+ hearts and myocytes, but enhances SR Ca 2+ sparks and leak on confocal microscopy. PLN ablation diminishes ventricular arrhythmias promoted by CaMKII phosphorylation of S2814 on RyR2. PLN ablation aborts the arrhythmogenic SR Ca 2+ waves of S2814D +/+ and transforms them into non‐propagating events. A mathematical human myocyte model replicates these results and predicts the increase in SR Ca 2+ uptake required to prevent the arrhythmias induced by a CaMKII‐dependent leaky RyR2. Abstract: Mice with constitutive pseudo‐phosphorylation at Ser2814‐RyR2 (S2814D +/+ ) have increased propensity to arrhythmias under β‐adrenergic stress conditions. Although abnormal Ca 2+ release from the sarcoplasmic reticulum (SR) has been linked to arrhythmogenesis, the role played by SR Ca 2+ uptake remains controversial. We tested the hypothesis that an increase in SR Ca 2+ uptake is able to rescue the increased arrhythmia propensity of S2814D +/+ mice. We generated phospholamban (PLN)‐deficient/S2814D +/+ knock‐in mice by crossing two colonies, S2814D +/+ and PLNKO mice (SD +/+ /KO). SD +/+ /KO myocytes exhibited both increased SR Ca 2+ uptake seen in PLN knock‐out (PLNKO) myocytes and diminished SR Ca 2+ load (relative to PLNKO), a characteristic of S2814D +/+ myocytes. Ventricular arrhythmias evoked by catecholaminergic challenge (caffeine/adrenaline) in S2814D +/+ mice in vivo or programmed electric stimulation and high extracellular Ca 2+ in S2814D + / − hearts ex vivo were significantly diminished by PLN ablation. At the myocyte level, PLN ablation converted the arrhythmogenic Ca 2+ waves evoked by high extracellular Ca 2+ provocation in S2814D +/+ mice into non‐propagated Ca 2+ mini‐waves on confocal microscopy. Myocyte Ca 2+ waves, typical of S2814D +/+ mice, could be evoked in SD +/+ /KO cells by partially inhibiting SERCA2a. A mathematical human myocyte model replicated these results and allowed for predicting the increase in SR Ca 2+ uptake required to prevent the arrhythmias induced by a Ca 2+ –calmodulin‐dependent protein kinase (CaMKII)‐dependent leaky RyR2. Our results demonstrate that increasing SR Ca 2+ uptake by PLN ablation can prevent the arrhythmic events triggered by SR Ca 2+ leak due to CaMKII‐dependent phosphorylation of the RyR2‐S2814 site and underscore the benefits of increasing SERCA2a activity on SR Ca 2+ ‐triggered arrhythmias. Key points: Mice with Ca 2+ –calmodulin‐dependent protein kinase (CaMKII) constitutive pseudo‐phosphorylation of the ryanodine receptor RyR2 at Ser2814 (S2814D +/+ mice) exhibit a higher open probability of RyR2, higher sarcoplasmic reticulum (SR) Ca 2+ leak in diastole and increased propensity to arrhythmias under stress conditions. We generated phospholamban (PLN)‐deficient S2814D +/+ knock‐in mice by crossing two colonies, S2814D +/+ and PLNKO mice, to test the hypothesis that PLN ablation can prevent the propensity to arrhythmias of S2814D +/+ mice. PLN ablation partially rescues the altered intracellular Ca 2+ dynamics of S2814D +/+ hearts and myocytes, but enhances SR Ca 2+ sparks and leak on confocal microscopy. PLN ablation diminishes ventricular arrhythmias promoted by CaMKII phosphorylation of S2814 on RyR2. PLN ablation aborts the arrhythmogenic SR Ca 2+ waves of S2814D +/+ and transforms them into non‐propagating events. A mathematical human myocyte model replicates these results and predicts the increase in SR Ca 2+ uptake required to prevent the arrhythmias induced by a CaMKII‐dependent leaky RyR2.