Alterations in cardiac beta-catenin precede connexin gap junction remodeling in cardiomyocytes exposed to rapid electrical stimulation

Abstract

The Intercalated Disc (ID) contains two complexes, Adhesion Junction (AJ) and Connexin (Cx) Gap Junction (GJ). GJs provide the pathway for intercellular current flow, enabling coordinated action potential propagation and contraction. AJ mediates normal mechanical coupling between cardiomyocytes and plays an important role in the stability of GJ. Purpose: The purpose of this study was to study ID remodeling, especially β-catenin, which is one of the component proteins of AJ, and Cx43 alterations caused by Rapid Electrical Stimulation (RES) in cardiomyocytes. In addition, we investigated the effects of olmesartan, Angiotensin II receptor blockade on ID remodeling. Methods: Cultured neonatal rat ventricular myocytes were subjected to RES at 3.0 Hz for up to 60 min. We used real-time PCR, western blotting, and immunohistochemical methods to investigate expressions. Conduction properties of the cultured ventricular myocytes were examined by extracellular potential mapping by using a specifically designed multi-electrode array system. Results: Cx43 protein expression in cardiomyocytes was significantly increased (x 1.5 vs 0 min. p<0.01) after 60 min RES. Beta-catenin expression in total cell fraction was also significantly increased (x 1.5 vs 0 min. p<0.05) after 30 min. Interestingly, the expression level of β-catenin at nucleus, which functions as T Cell Factor/Lymphocyte Enhancer binding Factor (LEF/TCF) transcriptional activator of Cx43 and its degradation is modulated by glycogen synthase kinase 3β(GSK3β), was dramatically increased (x 1.4 vs 0 min. p<0.05) after 10 min. Conduction velocity was increased significantly by RES for 60min (25% increase). Treatment of cardiomyocytes with olmesartan prevented most these effects of RES. Moreover, we showed that an increase of phospholylated GSK3β, which is phosphorylated by activated mitogen-activated protein kinases due to RES and inhibits β-catenin degradations, was attenuated by olmesartan. Conclusions: The changes in AJ protein β-catenin precede Cx43 GJ remodeling and might play an important role in the formation and stability of GJs during RES exposure to cardiomyocytes. Olmesartan might be a new upstream arrhythmia therapy by modulating both AJ and GJ remodeling through β-catenin signaling pathway.