Role of Potassium Channels in Fibrillatory Conduction

描述(申请人提供)：这项工作的重点是纤颤的分子机制。最近发现了两种不同的通道病，与KCNJ2基因的功能获得突变有关，KCNJ2基因编码了导致IK1的内向整流通道(Kir2.1)蛋白：i)短QT综合征的变体SQTS3，由增加心源性猝死风险的突变(D172)引起；以及ii)一种新型的家族性心房颤动(AF)，由KCNJ2的另一种突变(V93I)引起。我们的具体目标是：1.检验IK1的外向分量在控制导致纤颤的转子的频率和稳定性方面起关键作用的假设。我们将在D172N和V93I突变小鼠、过度表达野生型Kir2.1通道的小鼠和Kir2.1-AAA小鼠的成年心脏和新生心肌细胞单层中使用计算机模拟2D繁殖和实验。我们还将确定改变[K+]o对再入和纤颤的频率和稳定性的影响。对人体心脏兴奋的模拟将帮助我们检查结果的临床相关性。2.比较K1增大和减小在汇-源失配导致破波和再入机制中的作用。模拟和单层实验将比较D172N和V93I突变与Kir2.1通道表达增加或减少对兴奋性、曲率-速度关系和涡旋脱落的影响。我们将检验这一假设，即Kir2.1的函数增益增加了成功传播的临界曲率半径，从而增加了波破裂的发生率，而函数损失具有相反的效果。3.确定空间梯度在波折形成和再入过程中野生型和突变型Kir2.1蛋白表达中的作用。我们假设，IK1密度的梯度有助于难治性的分散，而Kir2.1的过度表达放大了弥散性的心律失常效应。在野生型和遗传改变的具有IK1密度分散特定模式的心肌细胞单层中进行的数值和生物学实验将确定Kir2.1通道梯度在再入中的作用。此外，在成人心脏中，免疫组织化学、膜片钳和光学标测将确定Kir2.1梯度是否有助于完整小鼠心脏的波形破裂和再入。总之，这些研究应该为SQTS3和房颤患者的心律失常机制提供洞察力，也可能为许多受特发性室颤影响的患者提供见解。

DESCRIPTION (provided by applicant): This work focuses on the molecular mechanisms of fibrillation. It is motivated by the recent discovery of two different channelopathies associated with gain-of-function mutations in the KCNJ2 gene that codes for the inward rectifier channel (Kir2.1) protein responsible for IK1: i) a variant of short QT syndrome, SQTS3, resulting from a mutation (D172) that increases risk of sudden cardiac death; and ii) a new type of familial atrial fibrillation (AF) that results from a different mutation (V93I) in KCNJ2. Our Specific Aims are: 1. To test the hypothesis that the outward component of IK1 is critical in controlling frequency and stability of rotors responsible for fibrillation. We will use computer simulations of 2D propagation and experiments in adult hearts and neonatal myocyte monolayers from D172N and V93I mutant mice, mice overexpressing wild-type Kir2.1 channels, and Kir2.1-AAA mice. We will also determine the effects of changing [K+]o on frequency and stability of reentry and fibrillation. Simulations of human cardiac excitation will help us check the clinical relevance of our results. 2. To compare the effects of increasing versus decreasing IK1 in the mechanism of sink-to-source mismatch leading to wavebreak and reentry. Simulations and experiments in monolayers will compare the effects of D172N and V93I mutations with those of increasing or decreasing the expression of Kir2.1 channels on excitability, curvature-velocity relationships and vortex-shedding. We will test the hypothesis that gain-of-function of Kir2.1 increases the critical radius of curvature for successful propagation and thus the incidence of wavebreaks, whereas loss-of function has the opposite effects. 3. To determine the role of spatial gradients in the expression of wildtype and mutant Kir2.1 proteins on wavebreak formation and reentry. We hypothesize that gradients in IK1 density contribute to dispersion of refractoriness, and Kir2.1 overexpression amplifies the arrhythmogenic effect of dispersion. Numerical and biological experiments in wildtype and genetically altered myocyte monolayers having specific patterns of IK1 density dispersion will ascertain the role of Kir2.1 channel gradients in reentry. Also, immunohistochemistry, patch clamping and optical mapping in adult hearts will determine whether Kir2.1 gradients contribute to wavebreak and reentry in the intact mouse heart. Altogether, these studies should provide insights into mechanisms of arrhythmias in SQTS3 and AF patients, and possibly also in many patients affected by idiopathic VF.