title: Deep bradycardia and heart block caused by inducible cardiac-specific knockout of the pacemaker channel gene Hcn4.
abstract: Cardiac pacemaking generation and modulation rely on the coordinated activity of several processes. Although a wealth of evidence indicates a relevant role of the I(f) ("funny," or pacemaker) current, whose molecular constituents are the hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels and particularly HCN4, work with mice where Hcn genes were knocked out, or functionally modified, has challenged this view. However, no previous studies used a cardiac-specific promoter to induce HCN4 ablation in adult mice. We report here that, in an inducible and cardiac-specific HCN4 knockout (ciHCN4-KO) mouse model, ablation of HCN4 consistently leads to progressive development of severe bradycardia (∼50% reduction of original rate) and AV block, eventually leading to heart arrest and death in about 5 d. In vitro analysis of sinoatrial node (SAN) myocytes isolated from ciHCN4-KO mice at the mean time of death revealed a strong reduction of both the I(f) current (by ∼70%) and of the spontaneous rate (by ∼60%). In agreement with functional results, immunofluorescence and Western blot analysis showed reduced expression of HCN4 protein in SAN tissue and cells. In ciHCN4-KO animals, the residual I(f) was normally sensitive to β-adrenergic receptor (β-AR) modulation, and the permanence of rate response to β-AR stimulation was observed both in vivo and in vitro. Our data show that cardiac HCN4 channels are essential for normal heart impulse generation and conduction in adult mice and support the notion that dysfunctional HCN4 channels can be a direct cause of rhythm disorders. This work contributes to identifying the molecular mechanism responsible for cardiac pacemaking.
authors.affiliation: Department of Biomolecular Sciences and Biotechnology, PaceLab and Centro Interuniversitario di Medicina Molecolare e Biofisica Applicata, Milano, Università degli Studi di Milano, 20133 Milan, Italy.
authors.forename: Mirko
authors.initials: M
authors.lastname: Baruscotti
authors.forename: Annalisa
authors.initials: A
authors.lastname: Bucchi
authors.forename: Carlo
authors.initials: C
authors.lastname: Viscomi
authors.forename: Giacomo
authors.initials: G
authors.lastname: Mandelli
authors.forename: Giacomo
authors.initials: G
authors.lastname: Consalez
authors.forename: Tomaso
authors.initials: T
authors.lastname: Gnecchi-Rusconi
authors.forename: Nicola
authors.initials: N
authors.lastname: Montano
authors.forename: Karina Rabello
authors.initials: KR
authors.lastname: Casali
authors.forename: Stefano
authors.initials: S
authors.lastname: Micheloni
authors.forename: Andrea
authors.initials: A
authors.lastname: Barbuti
authors.forename: Dario
authors.initials: D
authors.lastname: DiFrancesco
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D015153:Blotting, Western; D050071:Bone Density Conservation Agents / Q000494:pharmacology; D001919:Bradycardia / Q000235:genetics / Q000503:physiopathology*; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000235:genetics / Q000502:physiology*; D004562:Electrocardiography; D005260:Female; D005455:Fluorescent Antibody Technique; D006321:Heart / Q000187:drug effects / Q000503:physiopathology*; D006327:Heart Block / Q000235:genetics / Q000503:physiopathology*; D006339:Heart Rate / Q000187:drug effects; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D008297:Male; D051379:Mice; D057507:Mice, 129 Strain; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009206:Myocardium / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D011943:Receptors, Adrenergic, beta / Q000378:metabolism; D012849:Sinoatrial Node / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D013629:Tamoxifen / Q000494:pharmacology
chemical_list: D050071:Bone Density Conservation Agents; D054815:Cyclic Nucleotide-Gated Cation Channels; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D011943:Receptors, Adrenergic, beta; D013629:Tamoxifen