Ventricular remodeling in a murine model of cardiac selective and inducible deletion of HCN4 gene

Purpose- Heart rate disorders are responsible for ventricular remodeling and enhanced susceptibility to cardiac arrhythmias. In experimental bradycardia electrical remodeling of cardiac myocytes (CM) includes a reduction of the delayed rectifier K+ currents, which are responsible for action potential repolarization delay and increased propensity to develop potentially lethal arrhythmias. The aim of this study was to evaluate ventricular remodeling occurring in a novel model of bradycardia (Baruscotti M., PNAS 2011), based on inducible and cardiac specific ablation of HCN4 gene. It encodes for the f current in sinoatrial node cells and contributes fundamentally to pacemaker activity. Methods- Cardiac HCN4 knockout was induced by i.p. injections of tamoxifen (200 mg/Kg) for 5 consecutive days in adult HCN4lox/lox;Cre mice. Control mice underwent the same treatment. Heart rate was monitored daily. Electrophysiological study was performed using the single-cell patch-clamp technique. Gene expression was studied by quantitative RT-PCR on ventricular tissue. Results- Tamoxifen injections produced a progressive development of bradycardia reducing heart rate by 25% in the HCN4lox/lox;Cre mice but not in control animals. Bradycardia was associated to 75% reduction of cardiac HCN4 mRNA level. Patch-clamp recordings on ventricular CM showed a marked prolongation of action potential duration in HCN4lox/lox;Cre bradycardic mice with respect to control animals (80±0.4 vs 30±0.1 ms at 90% of repolarization). Quantitative mRNA expression of subunits underlying repolarizing currents showed downregulation of the inward-rectifier K+ channel subunit Kir2.1 (-60%), the slow delayed-rectifier K+ channel subunit Kcnq1 (-50%), and the transient outward K+ channel subunits Kv4.2 (-80%),Kv4.3 (-50%) and KChIP2 (-90%) in HCN4lox/lox;Cre bradycardic mice with respect to control animals. The rapid delayed-rectifier K+ channel subunit erg and the regulatory subunit MinK were unchanged, while the regulatory subunit Mirp-1 was upregulated. Conclusions- The HCN4lox/lox;Cre transgenic mouse represents an innovative and reliable model of inducible bradycardia useful to study rate-related electrical remodeling of CM. In accordance to Gross’s model, we identified a prominent repolarization delay of ventricular action potential as major electrophysiological modification in bradycardic HCN4lox/lox;Cre mice. Expression studies point to a crucial role for the downregulation of the inward-rectifier, the slow delayed-rectifier, and the transient outward K+ channels to underlie electrical remodeling in braycardic HCN4lox/lox;Cre mice.