Abstract 2810: Increased Diastolic Calcium Levels Due to CaMKII-Dependent Sarcoplasmic Reticulum Calcium Leak in Patients With Atrial Fibrillation
While diastolic Ca leak from the sarcoplasmic reticulum (SR), a possible potent arrhythmogenic mechanism, has been observed in atrial fibrillation (AF), there is lack of knowledge about its cause and consequences. Ca/calmodulin protein kinase II (CaMKII)-dependent phosphorylation of the ryanodine receptor (RyR2) is believed to act as a critical mediator of SR Ca leak. Right atrial samples from 18 patients with sinus rhythm and 12 patients with AF were obtained during surgery. They were used for Western blot experiments and for the isolation of single atrial myocytes. Both under control conditions (KN-92) and CaMKII-inhibition (KN-93), SR Ca leak (Ca sparks) as well as diastolic [Ca] were measured in these cells. Tetracaine was used to investigate SR Ca leak consequences for diastolic [Ca]. CaMKII expression was increased by 34±10% in AF compared to sinus rhythm (n=11 vs. n=9, p<0.05), accompanied by increased RyR2 phosphorylation at the CaMKII site by 134±58% (n=11 vs. n=9, p<0.05). Furthermore, both frequency (n=19 vs. n=32 cells, p<0.05) and size of Ca sparks (n=96 vs. n=72 sparks, p<0.05) were significantly increased, leading to a marked increase in total SR Ca leak in AF (723±63 nM vs. 186±24 nM,p<0.05). This was associated with elevated diastolic [Ca] in AF cells (230±20 vs. 164±8 nM, n=33 vs. n=43, p<0.05). Tetracaine experiments confirmed that in fact SR Ca leak caused the elevated diastolic [Ca] observed. To investigate the role of CaMKII, measurements from the same samples were also performed in the presence of CaMKII-inhibition, which completely normalized SR Ca leak in AF cells and also lead to complete normalization of diastolic [Ca]. To evaluate possible antiarrhythmic effects of CaMKII-inhibition beyond the single cell level, isolated muscle strips were investigated. Preliminary data show arrhythmias in 4 out of 9 experiments in AF muscle strips under control conditions, but only in 1 out of 7 in the presence of CaMKII-inhibition. Our experiments show elevated diastolic [Ca] in human AF. We furthermore show that this is due to hyperphosphorylation of the RyR2 at the CaMKII site, which induces a strong SR Ca leak in AF, and that this can be completely normalized by CaMKII-inhibition. CaMKII-dependent SR Ca leak could be a novel therapeutic target in AF.