Deep bradycardia and heart block caused by inducible cardiac-specific knockout of the pacemaker channel gene Hcn4
Deep bradycardia and heart block caused by inducible cardiac-specific knockout of the pacemaker channel gene Hcn4
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DOI：
10.1073/pnas.1010122108
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发表时间：
2011-01-25
影响因子：
11.1
通讯作者：
DiFrancesco, Dario
中科院分区：
文献类型：
--
作者：
Baruscotti, Mirko;Bucchi, Annalisa;DiFrancesco, Dario
心脏起搏的产生和调节依赖于几个过程的协调活动。虽然大量证据表明If（“有趣”或起搏器）电流的相关作用，其分子成分是超极化激活的环核苷酸门控（HCN）通道，特别是HCN 4，但在Hcn基因被敲除或功能修饰的小鼠中工作，挑战了这一观点。然而，以前的研究没有使用心脏特异性启动子来诱导成年小鼠的HCN 4消融。我们在此报告，在诱导型和心脏特异性HCN 4敲除（ciHCN 4-KO）小鼠模型中，HCN 4消融持续导致严重心动过缓（类似于原始心率降低50%）和AV阻滞的进展，最终导致心脏骤停和约5 d内死亡。在平均死亡时间从ciHCN 4-KO小鼠分离的窦房结（SAN）肌细胞的体外分析显示If电流（类似于70%）和自发速率（类似于60%）两者的强烈降低。与功能结果一致，免疫荧光和Western印迹分析显示HCN 4蛋白在SAN组织和细胞中的表达减少。在ciHCN 4-KO动物中，残余I-f通常对β-肾上腺素能受体（β-AR）调节敏感，并且在体内和体外均观察到对β-AR刺激的频率响应的持久性。我们的数据表明，心脏HCN 4通道是必不可少的正常心脏冲动的产生和传导在成年小鼠和支持的概念，功能失调的HCN 4通道可能是一个直接原因的节律紊乱。这项工作有助于确定负责心脏起搏的分子机制。
Cardiac pacemaking generation and modulation rely on the coordinated activity of several processes. Although a wealth of evidence indicates a relevant role of the If ("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 (similar to 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 If current (by similar to 70%) and of the spontaneous rate (by similar to 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 beta-adrenergic receptor (beta-AR) modulation, and the permanence of rate response to beta-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.