Rationale

The mechanisms underlying atrial fibrillation (AF), the most common clinical arrhythmia, are poorly understood. Nucleoplasmic Ca²⁺ regulates gene expression, but the nature and significance of nuclear Ca²⁺-changes in AF are largely unknown.

Objective

To elucidate mechanisms by which AF alters atrial-cardiomyocyte nuclear Ca²⁺ ([Ca²⁺]_Nuc) and CaMKII (Ca²⁺/calmodulin-dependent protein kinase-II)-related signaling.

Methods and results

Atrial cardiomyocytes were isolated from control and AF dogs (kept in AF by atrial tachypacing [600 bpm × 1 week]). [Ca²⁺]_Nuc and cytosolic [Ca²⁺] ([Ca²⁺]_Cyto) were recorded via confocal microscopy. Diastolic [Ca²⁺]_Nuc was greater than [Ca²⁺]_Cyto under control conditions, while resting [Ca²⁺]_Nuc was similar to [Ca²⁺]_Cyto; both diastolic and resting [Ca²⁺]_Nuc increased with AF. IP₃R (Inositol-trisphosphate receptor) stimulation produced larger [Ca²⁺]_Nuc increases in AF versus control cardiomyocytes, and IP₃R-blockade suppressed the AF-related [Ca²⁺]_Nuc differences. AF upregulated nuclear protein expression of IP₃R1 (IP₃R-type 1) and of phosphorylated CaMKII (immunohistochemistry and immunoblot) while decreasing the nuclear/cytosolic expression ratio for HDAC4 (histone deacetylase type-4). Isolated atrial cardiomyocytes tachypaced at 3 Hz for 24 hours mimicked AF-type [Ca²⁺]_Nuc changes and L-type calcium current decreases versus 1-Hz-paced cardiomyocytes; these changes were prevented by IP₃R knockdown with short-interfering RNA directed against IP₃R1. Nuclear/cytosolic HDAC4 expression ratio was decreased by 3-Hz pacing, while nuclear CaMKII phosphorylation was increased. Either CaMKII-inhibition (by autocamtide-2-related peptide) or IP₃R-knockdown prevented the CaMKII-hyperphosphorylation and nuclear-to-cytosolic HDAC4 shift caused by 3-Hz pacing. In human atrial cardiomyocytes from AF patients, nuclear IP₃R1-expression was significantly increased, with decreased nuclear/nonnuclear HDAC4 ratio. MicroRNA-26a was predicted to target ITPR1 (confirmed by luciferase assay) and was downregulated in AF atrial cardiomyocytes; microRNA-26a silencing reproduced AF-induced IP₃R1 upregulation and nuclear diastolic Ca²⁺-loading.

Conclusions

AF increases atrial-cardiomyocyte nucleoplasmic [Ca²⁺] by IP₃R1-upregulation involving miR-26a, leading to enhanced IP₃R1-CaMKII-HDAC4 signaling and L-type calcium current downregulation. Graphic Abstract: A graphic abstract is available for this article.