A Simple Regulation of Cardiomyocyte Excitability

Abstract

Introduction: Recently, genome wide association studies (GWAS) identified LITAF (lipopolysaccharide-induced TNF factor) as one of the novel loci associated with prolongation of QT interval. Our goal is to investigate the role of LITAF in regulating cardiomyocyte excitability and action potential duration (APD). Methods: Optical mapping was performed in zebrafish hearts to determine Ca2+ transients and transmembrane (TM) potentials. Propagation velocities of depolarizing waves were determined. Action potential (AP) was digitally imaged in adenoviral infected neonatal rabbit cardiomyocytes (NRbCM). Live cell confocal calcium imaging was performed on adult rabbit ventricular myocytes (ARbCM) to determine intracellular Ca2+ handling. A current-voltage (I-V) relationship for the L-type calcium current (ICa,L ) was determined using whole cell patch clamping. To study the effect of LITAF on Cav1.2 channel expression, surface biotinylation and western blot assays were performed using tSA201 cells. Results: Our studies demonstrate that loss-of-function of LITAF in vivo (zebrafish) increases the Ca2+ transient amplitude (which primarily depends on calcium influx), slows atrial-ventricular conduction, and prolongs the APD. In contrast, in vitro gain-of-function experiments in rabbit CM established that overexpression of LITAF downregulates the total and surface pools of Cav1.2 channel polypeptides, decreases Ca2+ transients and L-type Ca2+ conductance, and shorten the APD. Studies in heterologous expression systems revealed that one of the auxiliary subunits of Cav1.2 channels, Cavβ2b, is likely involved in LITAF-mediated inhibition of recombinant Cav1.2 channels. Conclusion: LITAF regulates the expression and function of Cav1.2 channels, thereby determining APD and cardiac cell excitation.