Cardiac chloride and pH regulation in health and disease

健康和疾病中的心氯和 pH 调节

• 批准号: 10586799
• 负责人: Xiao-Dong Zhang
• 金额: $ 39.95万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10586799
• 关键词: Ablation; Acids; Action Potentials; Affect; Anions; Arrhythmia; Bicarbonates; Biochemical; Buffers; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cations; Cell physiology; Cells; Cessation of life; Chlorides; Conscious; Coronary heart disease; Coupled; Data; Disease; EKG QRS Complex; Electrocardiogram; Electrophysiology (science); Family; Frequencies; Generations; Goals; Health; Heart; Heart Atrium; Heart Diseases; Homeostasis; Human; Human Cloning; Image; Impairment; Ischemia; Knockout Mice; Link; Malignant Neoplasms; Mechanics; Mediating; Microscopy; Molecular; Motivation; Mus; Muscle Cells; Myocardial Infarction; Myocardial Ischemia; Myocardium; Pacemakers; Pathologic; Physiological; Play; Protein Isoforms; Reagent; Regulation; Reperfusion Injury; Reperfusion Therapy; Reporting; Role; Signal Transduction; Sinus Bradycardia; Skeletal Muscle; System; Techniques; Telemetry; Testing; Therapeutic; Transcript; United States; Ventricular; antiporter; base; cardioprotection; confocal imaging; design; dynamic system; experimental study; genetic approach; heart function; in vivo; insight; interdisciplinary approach; member; mortality; mouse model; multimodality; new therapeutic target; novel; second harmonic; solute; symporter

Heart disease is the leading cause of mortality in the United States and causes more deaths than all cancers combined. Coronary heart disease (or ischemic heart disease, IHD), the most common type of heart disease, is accompanied by a major decline of local pH in myocardium. However, the mechanisms of pH regulation and the homeostasis of H+ neutralizing buffers, such as HCO3- and Cl- in cardiomyocytes remain incompletely understood, making it difficult to design therapeutic strategies targeting pH regulation. Recently, we have identified and cloned different isoforms of a solute carrier, Slc26a6, from cardiac myocytes. Slc26a6 is the predominant Cl-/HCO3- exchanger in the heart. We demonstrated that Slc26a6 mediates electrogenic Cl-/HCO3- exchange activities in both atrial and ventricular myocytes. Our findings raise the possibility that Slc26a6 may represent the predominant Cl-/HCO3- regulatory mechanism in the heart. We have obtained exciting data to support the critical roles of Slc26a6 in cardiac excitability and contractility. We documented that null deletion of Slc26a6 in mice results in shortened action potentials (APs), sinus bradycardia, fragmented QRS complexes and impaired cardiac function compared to wild type littermates. We have identified and characterized two isoforms of human SLC26A6 in human heart, which are also electrogenic, akin to mouse cardiac Slc26a6. In addition, we recently identified and reported a dynamic beat-to-beat intracellular pH (pHi) regulation system, termed “pHi transients”, which dovetails with the prevailing three known dynamic systems, namely electrical, Ca2+, and mechanical systems. However, critical questions remain unanswered. How do Slc26a6 activities affect not only pHi, but also cardiac AP and contractility? The goal of study is to determine the mechanistic links between the Slc26a6 activities and cardiac AP and contractility. Contributions of Slc26a6-mediated Cl-/HCO3- towards the pHi transients will also be tested. Taken together, we hypothesize that the activities of Slc26a6 on pHi will directly contribute towards intracellular Na+ homeostasis, through Na+/HCO3- cotransporter (NBCe) and Na+/H+ exchanger (NHE), and subsequently regulate intracellular Ca2+ concentration through sarcolemmal Na+- Ca2+ exchanger (NCX). Therefore, ablation of Slc26a6 will result in a reduction in intracellular Na+ and Ca2+ through the actions of NHE/NBCe and NCX, respectively. We further hypothesize that Slc26a6 plays important roles in the dynamic pHi regulation in the heart regulating cardiac pacemaking activities and contractility. We will test our hypothesis using multidisciplinary approaches including functional electrophysiological recordings, imaging, biochemical, molecular and genetic approaches as well as ex vivo and in vivo functional studies. Wild type and cardiac-specific Slc26a6 knockout mouse model as well as human cardiomyocytes will be tested. Three specific aims are: 1. To determine the regulatory mechanisms of Slc26a6 on cardiac pHi and function. We will test how Slc26a6 regulates dynamic cardiac pHi, Na+ and Ca2+ homeostasis, hence, excitability and contractility. The relationship between pHi and cardiac function will be directly tested to gain mechanistic insights into the functional roles of Slc26a6 in the heart. We will use novel techniques including multimodal second harmonic generation (SHG) microscopy and our recently established dynamic pH recording techniques. 2. To determine the mechanistic roles of Slc26a6 in cardiac ischemia/reperfusion (I/R). We will test the contributions of Slc26a6 to cardiac function in the I/R mouse model. Mechanistic roles of Slc26a6 in cardiac I/R injury will be tested using ex vivo confocal imaging of pHi, intracellular Na+ and Ca2+ concentrations. I/R injury will be employed in control and Slc26a6-/- mice. 3. To determine the functional roles and regulatory mechanisms of Slc26a6 in cardiac pacemaking activities. We will test the mechanistic roles of Slc26a6 in the regulation of AP firing frequency, pacemaker currents, Ca2+ signaling, and pHi in SAN cells. Additionally, ECG telemetry will be used to test the roles of Slc26a6 in conscious control and SAN-specific Slc26a6-/- mice. Our studies will unravel a missing molecular link between pHi regulation and Na+, and Ca2+ homeostasis in the heart. The anticipated results will provide novel insights into the roles of Slc26a6 in cardiac pHi regulation, cardiac excitability, and function under physiological and pathological conditions. At the translational level, Slc26a6 may represent a novel therapeutic target for cardioprotection in cardiac ischemia and arrhythmia.

心脏病是美国死亡率最高的疾病，死亡人数超过所有癌症 加起来冠心病（或缺血性心脏病，IHD）是最常见的心脏病类型， 伴随心肌局部pH值的显著下降。然而，pH调节的机制和 心肌细胞中H+中和缓冲液（如HCO 3-和Cl-）的稳态仍不完全 这使得难以设计针对pH调节的治疗策略。最近我们 从心肌细胞中鉴定并克隆了溶质载体Slc 26 a6的不同亚型。slc 26 a6是 心脏中的主要Cl-/HCO 3-交换器。我们证明Slc 26 a6介导产电Cl-/HCO 3- 在心房和心室肌细胞中的交换活动。我们的发现提高了Slc 26 a6可能 代表心脏中的主要Cl-/HCO 3-调节机制。我们获得了令人兴奋的数据， 支持Slc 26 a6在心脏兴奋性和收缩性中的关键作用。我们记录了空删除 Slc 26 a6在小鼠中导致缩短的动作电位（AP）、窦性心动过缓、碎裂的QRS复合波 和受损的心脏功能。我们已经确定并描述了两个 人心脏中的人SLC 26 A6的同种型，其也是产电的，类似于小鼠心脏Slc 26 a6。在 此外，我们最近发现并报道了一种动态的逐拍细胞内pH（pHi）调节系统， 称为“pHi瞬变”，其与流行的三种已知动态系统，即电， Ca 2+和机械系统。然而，关键问题仍然没有答案。Slc 26 a6活动如何影响 不仅是pHi，还有心脏AP和收缩力？研究的目的是确定机械环节 Slc 26 a6活性与心脏AP和收缩力之间的关系。Slc 26 a6介导的Cl-/HCO 3-的贡献 也将测试pHi瞬变。综上所述，我们假设Slc 26 a6在 pHi将通过Na+/HCO 3-协同转运蛋白（NBCe）直接促进细胞内Na+稳态， Na+/H+交换器（NHE），并随后通过肌膜Na+-Na+通道调节细胞内Ca 2+浓度。 钙离子交换剂（NCX）。因此，切除Slc 26 a6将导致细胞内Na+和Ca 2+的减少 分别通过NHE/NBCe和NCX的作用。我们进一步假设Slc 26 a6在这一过程中起重要作用。 在调节心脏起搏活动和收缩性的心脏中的动态pHi调节中的作用。我们将 使用多学科方法测试我们的假设，包括功能性电生理记录， 成像、生物化学、分子和遗传方法以及离体和体内功能研究。野生 型和心脏特异性Slc 26 a6敲除小鼠模型以及人心肌细胞进行测试。三 具体目标是：1.探讨Slc 26 a6对心脏pHi和功能的调节机制。我们将 测试Slc 26 a6如何调节动态心脏pHi、Na+和Ca 2+稳态，从而调节兴奋性和收缩性。 将直接测试pHi和心脏功能之间的关系，以获得对心脏功能的机制性见解。 Slc 26 a6在心脏中的作用我们将使用新技术，包括多模态二次谐波 第二代（SHG）显微镜和我们最近建立的动态pH记录技术。2.以确定 Slc 26 a6在心肌缺血/再灌注（I/R）中的机制作用。我们将测试Slc 26 a6的贡献 对心脏功能的影响。Slc 26 a6在心脏I/R损伤中的机制作用将使用 pHi、细胞内Na+和Ca 2+浓度的离体共聚焦成像。I/R损伤将用于控制 和Slc 26 a6-/-小鼠。3.目的：探讨Slc 26 a6在心肌细胞凋亡中的作用及其调控机制。 pacemaking活动我们将测试Slc 26 a6在AP放电频率调节中的机制作用， 起搏器电流、Ca 2+信号和SAN细胞中的pHi。此外，ECG遥测将用于测试 Slc 26 a6在清醒对照和SAN特异性Slc 26 a6-/-小鼠中的作用。我们的研究将解开一个失踪的 pHi调节与心脏中Na+和Ca 2+稳态之间的分子联系。预期结果将 提供了新的见解Slc 26 a6在心脏pHi调节，心脏兴奋性和功能的作用， 生理和病理条件。在翻译水平上，Slc 26 a6可能代表一种新的治疗药物 在心脏缺血和心律失常中心脏保护靶点。