Decreased NCX1 expression reduces proarrhythmogenic spontaneous Ca2+ releases in atrial cardiomyocytes from CREM-IbΔC-X transgenic mice, a model of atrial fibrillation

Introduction: Atrial fibrillation (AF) is the most common cardiac arrhythmia and is associated with extensive arrhythmogenic functional and structural atrial remodeling. Impaired Ca²⁺ dynamics are thought to promote AF through increased incidence of arrhythmogenic spontaneous Ca²⁺ releases. The cardiac Na⁺/Ca²⁺-Exchanger NCX1 generates a depolarising inward current during Ca²⁺ extrusion that may increase triggered activity during extrusion of spontaneously released Ca²⁺. Studies in genetically modified mice have shown that the NCX1-expression level affects triggered activity in atrial myocytes. Reportedly, NCX1 is upregulated in human AF. Here, we examined for the first time the effects of altered NCX1 expression in a disease model and tested whether the NCX1-expression level affects functional and structural remodeling in CREM-IbΔC-X transgenic mice, a model of atrial fibrillation.

Material & Methods: Mice with cardiomyocyte-specific expression of CREM-IbΔC-X (TG) were mated with mice with either increased (NCX TG) or reduced (NCX hetKO) NCX1 expression resulting in TG x NCX TG (DTG) and TG x NCX hetKO (TGxhetKO) mice, which were investigated at 12 weeks of age. Regular Ca²⁺ transients and spontaneous Ca²⁺ releases (sCaR) were measured with Fluo-4/AM in atrial cardiomyocytes (ACMs) using a specific stress protocol (FIG. A). Based on their characteristics, the sCaR were divided into 3 groups: waves, oscillations and transient-like sCaR (FIG. B). Morphometry of ACMs was optically measured. In addition, atrial weights were determined. Atrial tissue mRNA and proteins were analyzed by qPCR and western blotting.

Results: As shown in figure C, NCX TG and DTG ACMs showed lower rhythmic stability due to an increased occurrence of sCaR versus WT and TG, respectively (Proportion of ACMs without sCaR in %: NCX TG 52% vs WT 63%; DTG 46% vs TG 57%). In contrast, atrial myocytes of NCX hetKO and TGxhetKO were more stable (Proportion of ACMs without sCaR in %: NCX hetKO 69% vs WT 63%; TGxhetKO 66% vs TG 57%) and were protected from supra-threshold transient-like sCaR versus respective controls (Proportion of transient-positive ACMs in %: hetKO 0% vs WT 18%; TGxhetKO 2% vs TG 7%) (mice/ACMs=8-12/58-87). On the other hand, the kinetics of regular Ca²⁺ transients (mice/ACMs=8-12/56-87), the expression of key proteins related to Ca²⁺ dynamics (SERCA2, Ca_V1.2, Phospholamban, Calsequestrin) and their genes (n=7-8) were not different between groups. ACMs dimensions (mice/ACMs=8/292-320) and atrial weights (mice=42-51/group) as a measure for the structural remodeling were not affected by the altered NCX1 expression level.

Conclusion: The structural remodeling was not affected by the NCX1-expression level. Increased expression of NCX1 promoted the proarrhythmic phenotype in the AF model of CREM-IbΔC-X mice. In contrast, the heterozygous inactivation of NCX1 had a protective effect on ACMs by increasing their stability even when CREM-IbΔC-X was overexpressed. Thus, for the treatment of atrial fibrillation, the inhibition of NCX1 may be a promising option.

Supported by the DFG