Human Atrial Fibrillation Is Not Associated With Remodeling of Ryanodine Receptor Clusters

Abstract

The release of Ca2+ by ryanodine receptor (RyR2) channels is critical for cardiac function. However, abnormal RyR2 activity has been linked to the development of arrhythmias, including increased spontaneous Ca2+ release in human atrial fibrillation (AF). Clustering properties of RyR2 have been suggested to alter the activity of the channel, with remodeling of RyR2 clusters identified in pre-clinical models of AF and heart failure. Whether such remodeling occurs in human cardiac disease remains unclear. This study aimed to investigate the nanoscale organization of RyR2 clusters in AF patients – the first known study to examine this potential remodeling in diseased human cardiomyocytes. Right atrial appendage from cardiac surgery patients with paroxysmal or persistent AF, or without AF (non-AF) were examined using super-resolution (dSTORM) imaging. Significant atrial dilation and cardiomyocyte hypertrophy was observed in persistent AF patients compared to non-AF, with these two parameters significantly correlated. Interestingly, the clustering properties of RyR2 were remarkably unaltered in the AF patients. No significant differences were identified in cluster size (mean ∼18 RyR2 channels), density or channel packing within clusters between patient groups. The spatial organization of clusters throughout the cardiomyocyte was also unchanged across the groups. RyR2 clustering properties did not significantly correlate with patient characteristics. In this first study to examine nanoscale RyR2 organization in human cardiac disease, these findings indicate that RyR2 cluster remodeling is not an underlying mechanism contributing to altered channel function and subsequent arrhythmogenesis in human AF.

Molecular, Subcellular, and Arrhythmogenic Mechanisms in Genetic RyR2 Disease

“…Yin et al found a reduced association between junctophilin-2 and RyR2 in RyR2-R420Q +/− knock-in mice measured by co-immunoprecipitation, suggesting this could contribute to the redistribution of cluster size that results from this mutation [ 77 ]. In contrast to work in the experimental sheep heart failure [ 89 ], Munro et al found RyR2 cluster and CRU properties to be unaltered in atrial myocytes from human patients with persistent AF versus patients without AF [ 90 ]. Here, the non-AF samples came from patients requiring coronary artery bypass graft surgery and may have experienced some degree of prior cardiac remodeling, which could contribute to the lack of CRU disruption observed.…”

“…Recent advances in super-resolution optical microscopy have further enabled the detection and characterization of RYR2 organization at near single molecule level in both 2D [ 86 , 87 ] and 3D [ 88 ], which was previously not possible due to diffraction limitations of conventional confocal microscopes. These recent investigations in rodent, sheep, and human myocytes have shown that RyR2 clusters typically consist of ~15–20 channels that can be sparsely packed [ 86 , 88 , 89 , 90 ]. This is much smaller than previous estimates of ~100 RyR2 per cluster, calculated from thin sectioning that relied on assumptions of complete filling and circular geometry [ 91 ].…”

Nanoscale Organization, Regulation, and Dynamic Reorganization of Cardiac Calcium Channels

“…AFib CRUs were more fragmented and contained more RyRs, dispersed over a larger area than healthy cell controls ( MacQuaide et al, 2015 ). However, in a recent study of RyR2 clustering in diseased atrial myocytes from human AFib patients, no dispersion or reorganization of RyR2 was observed suggesting that RyR2 reorganization is not a necessary component of the Ca 2+ leak and arrhythmogenicity associated with human AFib ( Munro et al, 2021 ).…”

“…The rat and sheep studies discussed above suggest that altered nanoscale organization of RyR2 in cardiomyopathies can contribute to the altered kinetics of Ca 2+ release and the disease phenotype in these species. Further investigations are needed to determine whether the same can be said for multi-factorial human cardiomyopathies given that human AFib does not appear to exhibit this phenomenon ( Munro et al, 2021 ).…”

Magnesium Ions Moderate Calcium-Induced Calcium Release in Cardiac Calcium Release Sites by Binding to Ryanodine Receptor Activation and Inhibition Sites

“…In heart failure (HF), increased non-spark leak, increased spark size, and decreased spark frequency were observed ( Kolstad et al, 2018 ); in atrial fibrillation (AF), increased duration, frequency, and time to peak of sparks were observed ( Macquaide et al, 2015 ). The changes observed at the cellular level might stem not only from changes in RyR gating but also from a changed structure of the calcium release sites that occurred in some ( Macquaide et al, 2015 ; Kolstad et al, 2018 ) but not all models of cardiac disease ( Munro et al, 2021 ). It is not known whether the changes observed in diseased cardiac myocytes are due to changes in the action of Mg 2+ , but the RyR inhibitor dantrolene, which requires Mg 2+ to exert its effect ( Choi et al, 2017 ), was shown to have beneficial effects in AF ( Hartmann et al, 2017 ).…”