Diabetes increases oxidant stress and doubles the risk of dying after myocardial infarction, but the mechanisms underlying increased mortality are unknown. Mice with streptozotocin-induced diabetes developed profound heart rate slowing and doubled mortality compared with controls after myocardial infarction. Oxidized Ca 2+ /calmodulin-dependent protein kinase II (ox-CaMKII) was significantly increased in pacemaker tissues from diabetic patients compared with that in nondiabetic patients after myocardial infarction. Streptozotocin-treated mice had increased pacemaker cell ox-CaMKII and apoptosis, which were further enhanced by myocardial infarction. We developed a knockin mouse model of oxidation-resistant CaMKIIδ (MM-VV), the isoform associated with cardiovascular disease. Streptozotocin-treated MM-VV mice and WT mice infused with MitoTEMPO, a mitochondrial targeted antioxidant, expressed significantly less ox-CaMKII, exhibited increased pacemaker cell survival, maintained normal heart rates, and were resistant to diabetes-attributable mortality after myocardial infarction. Our findings suggest that activation of a mitochondrial/ox-CaMKII pathway contributes to increased sudden death in diabetic patients after myocardial infarction.

Background Voltage-gated Na+ channels (Nav) are essential for myocyte membrane excitability and cardiac function. Nav current (INa) is a large amplitude, short duration “spike” generated by rapid channel activation followed immediately by inactivation. However, even under normal conditions, a small “late” component of INa (INa,L) persists due to incomplete/failed inactivation of a subpopulation of channels. Notably, INa,L is directly linked with both congenital and acquired disease states. The multifunctional Ca2+/calmodulin-dependent kinase II (CaMKII) has been identified as an important activator of INa,L in disease. Several potential CaMKII phosphorylation sites have been discovered, including Ser571 in the Nav1.5 DI-DII linker, but the molecular mechanism underlying CaMKII-dependent regulation of INa,L in vivo remains unknown. Methods and Results To determine the in vivo role of Ser571, two Scn5a knock-in mouse models were generated expressing either: 1) Nav1.5 with a phosphomimetic mutation at Ser571 (S571E), or 2) Nav1.5 with the phosphorylation site ablated (S571A). Electrophysiology studies revealed that Ser571 regulates INa,L but not other channel properties previously linked to CaMKII. Ser571-mediated increases in INa,L promote abnormal repolarization and intracellular Ca2+ handling, and increase susceptibility to arrhythmia at the cellular and animal level. Importantly, Ser571 is required for maladaptive remodeling and arrhythmias in response to pressure overload. Conclusions Our data provide the first in vivo evidence for the molecular mechanism underlying CaMKII activation of the pathogenic INa,L. Relevant for improved rational design of potential therapies, our findings demonstrate that Ser571-dependent regulation of Nav1.5 specifically tunes INa,L without altering critical physiological components of the current.

Background Human gene variants affecting ion channel biophysical activity and/or membrane localization are linked with potentially fatal cardiac arrhythmias. However, the mechanism for many human arrhythmia variants remains undefined despite over a decade of investigation. Post-translational modulation of membrane proteins is essential for normal cardiac function. Importantly, aberrant myocyte signaling has been linked to defects in cardiac ion channel post-translational modifications and disease. We recently identified a novel pathway for post-translational regulation of the primary cardiac voltage-gated Na+ channel (Nav1.5) by CaMKII. However, a role for this pathway in cardiac disease has not been evaluated. Methods and Results We evaluated the role of CaMKII-dependent phosphorylation in human genetic and acquired disease. We report an unexpected link between a short motif in the Nav1.5 DI-DII loop, recently shown to be critical for CaMKII-dependent phosphorylation, and Nav1.5 function in monogenic arrhythmia and common heart disease. Experiments in heterologous cells and primary ventricular cardiomyocytes demonstrate that human arrhythmia susceptibility variants (A572D and Q573E) alter CaMKII-dependent regulation of Nav1.5 resulting in abnormal channel activity and cell excitability. In silico analysis reveals that these variants functionally mimic the phosphorylated channel resulting in increased susceptibility to arrhythmia-triggering afterdepolarizations. Finally, we report that this same motif is aberrantly regulated in a large animal model of acquired heart disease and in failing human myocardium. Conclusions We identify the mechanism for two human arrhythmia variants that affect Nav1.5 channel activity through direct effects on channel post-translational modification. We propose that the CaMKII phosphorylation motif in the Nav1.5 DI-DII cytoplasmic loop is a critical nodal point for pro-arrhythmic changes to Nav1.5 in congenital and acquired cardiac disease.

These data provide new insight into membrane targeting of TREK-1 in the heart and establish a broader role for β(IV)-spectrin in organizing functional membrane domains critical for normal heart function.