Abstract
Rationale: Diabetic hyperglycemia is associated with cardiac dysfunction and increased arrhythmia risk, and CaMKII (calcium/calmodulin-dependent protein kinase II) function has been implicated. CaMKII activity is promoted by both oxidation and O -linked β-N-acetylglucosamine ( O -GlcNAc) of known CaMKII sites. Objective: To investigate which posttranslational modifications occur in human diabetic hearts and how they alter electrophysiological and Ca 2+ handling properties in hyperglycemia. Methods and Results: We assessed echocardiography, electrophysiology, Ca 2+ -handling, and protein expression in site-specific CaMKII mutant mice ( O -GlcNAc-resistant S280A and oxidation-resistant MM281/2VV knock-ins, and global and cardiac-specific knockouts), in myocytes subjected to acute hyperglycemia and Ang II (angiotensin II) and mice after streptozotocin injections (to induce diabetes). Human patients with diabetes exhibit elevated CaMKII O -GlcNAcylation but not oxidation. In mice, acute hyperglycemia increased spontaneous diastolic Ca 2+ sparks and waves and arrhythmogenic action potential changes (prolongation, alternans, and delayed afterdepolarizations), all of which required CaMKII-S280 O -GlcNAcylation. Ang II effects were dependent on NOX2 (NADPH oxidase 2)-mediated CaMKII MM281/2 oxidation. Diabetes led to much greater Ca 2+ leak, RyR2 S2814 phosphorylation, electrophysiological remodeling, and increased susceptibility to in vivo arrhythmias, requiring CaMKII activation, predominantly via S280 O -GlcNAcylation and less via MM281/2 oxidation. These effects were present in myocytes at normal glucose but were exacerbated with the in vivo high circulating glucose. PLB (phospholamban) O -GlcNAcylation was increased and coincided with reduced PLB S16 phosphorylation in diabetes. Dantrolene, which reverses CaMKII-dependent proarrhythmic RyR-mediated Ca 2+ leak, also prevented hyperglycemia-induced APD prolongation and delayed afterdepolarizations. Conclusions: We found that CaMKII-S280 O -GlcNAcylation is required for increased arrhythmia susceptibility in diabetic hyperglycemia, which can be worsened by an additional Ang II-NOX2-CaMKII MM281/2 oxidation pathway. CaMKII-dependent RyR2 S2814 phosphorylation markedly increases proarrhythmic Ca 2+ leak and PLB O -GlcNAcylation may limit sarcoplasmic reticulum Ca 2+ reuptake, leading to impaired excitation-contraction coupling and arrhythmogenesis in diabetic hyperglycemia.