Mechanism and functional role of AKAP79/150 in M current control and excitability

AKAP79/150 在 M 电流控制和兴奋性中的机制和功能作用

• 批准号: 7728381
• 负责人: MARK S SHAPIRO
• 金额: $ 37.92万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7728381
• 关键词: A kinase anchoring protein; Adrenergic Receptor; Agonist; Angiotensins; Animals; Binding; Binding Proteins; Biological; Biological Assay; Bradykinin; Calcineurin; Calmodulin; Cardiac Myocytes; Catecholamines; Cell physiology; Cells; Chinese Hamster; Chinese Hamster Ovary Cell; Co-Immunoprecipitations; Coculture Techniques; Confocal Microscopy; Cyclic AMP-Dependent Protein Kinases; Disease; Dominant-Negative Mutation; Electrophysiology (science); Emotional; Ensure; Fluorescence; Fluorescence Resonance Energy Transfer; G-Protein-Coupled Receptors; Goals; Health; Image; Knockout Mice; Light; Link; Measures; Mediating; Mediator of activation protein; Memory; Methods; Modeling; Molecular; Moods; Mus; Muscarinic Agonists; Muscarinics; Nervous System Physiology; Nervous system structure; Neurons; Nodose Ganglion; Ovary; Personality; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositols; Phosphoric Monoester Hydrolases; Physiological; Play; Potassium Channel; Property; Protein Kinase C; Proteins; Rattus; Receptor Signaling; Regulation; Research; Role; Sensory Ganglia; Shapes; Signal Pathway; Signal Transduction; Signaling Molecule; Small Interfering RNA; Specificity; Stimulus; Structure of superior cervical ganglion; System; Techniques; Testing; Ventricular; carbon fiber; cell preparation; chronotropic; detector; in vitro Model; knock-down; mutant; neuronal excitability; neurophysiology; neurotransmitter release; patch clamp; public health relevance; receptor; receptor coupling; receptor sensitivity; research study; response; scaffold; voltage

DESCRIPTION (provided by applicant): A-kinase anchoring proteins (AKAPs) organize numerous intracellular signaling pathways by bringing together their molecular components into discrete sub-cellular microdomains. One such AKAP, AKAP79/150 interacts with protein kinase A, protein kinase C (PKC), calmodulin (CaM), calcineurin, and phosphatidylinositol 4,5- bisphosphate (PIP2), along with effectors such as M-type (KCNQ, Kv7) K+ channels and certain G protein- coupled receptors. In this project, we will study which KCNQ1-5 subunits are targets of AKAP79/150, and which Gq/11-coupled receptors of sympathetic and nodose ganglia neurons (muscarinic M1, bradykinin B2, angiotensin AT1 and purinergic P2Y) use AKAP79/150 to modulate M-type channels. We will also investigate the interactions between the CaM and PIP2 molecules with AKAP79/150 that are critical to the function of both M channels and AKAP79/150. The role of AKAP79/150 in Gq/11-coupled receptor control of neuronal discharge properties of the neurons, and on their release of neurotransmitter will be explored, both at the single-cell level, and via an in vitro model of the chronotropic response of cardiomyocytes to sympathetic neuron activity. We will use a heterologous expression system in which M-channels, receptors and signaling molecules are expressed in Chinese hamster ovary (CHO) cells, preparations of rat and mouse superior cervical ganglia (SCG) and nodose ganglia (NG) neurons, and a co-culture of SCG neurons and ventricular cardiomyocytes. Techniques to be used include fluorescence resonance energy transfer (FRET), total internal reflection fluorescence (TIRF), confocal microscopy, patch-clamp electrophysiology, carbon-fiber amperometry and video imaging. We aspire to discover the mechanisms endowing AKAP79/150 in specificity towards receptors and M-type K+ channels, and its functional role in shaping the neurophysiological and neurotransmitter release properties of neurons. PUBLIC HEALTH RELEVANCE: The signaling pathways that comprise a common motif in biological signaling underlie how the nervous system is regulated, including alterations in emotional state and mood, personality and the acquisition and use of memories. We will study the regulation of the M-type potassium ion channel, which plays dominant role in regulation of nerve-cell excitability, mediated by the A-kinase Anchoring Protein, AKAP79/150. The elucidation of the mechanisms and functional role of AKAP79/150 in M-channel activity will shed light on how the nervous system functions in health and disease.

描述（由申请人提供）：A-激酶锚定蛋白（AKAP）通过将其分子成分聚集到离散的亚细胞微区域，来组织许多细胞内信号通路。一种这样的AKAP AKAP79/150与蛋白激酶A，蛋白激酶C（PKC），钙调蛋白（CAM），钙调蛋白和磷脂酰肌醇4,5-双磷酸（PIP2）相互作用，以及M-Type（KCNQ，KCNQ，KV7）K+通道和某些g pentersels和某些g pentersels和某些效果。在这个项目中，我们将研究哪些KCNQ1-5亚基是AKAP79/150的靶标，以及哪些syaltic和Nodose神经元神经元的GQ/11偶联受体（毒蕈碱M1，Bradylyinic M1，Bradykinin B2，AT1和Purinergic P2Y）使用AKAP79/150的Modunce M-M-M--我们还将研究CAM和PIP2分子与AKAP79/150之间的相互作用，这些相互作用对于M通道和AKAP79/150的功能至关重要。 AKAP79/150在神经元神经元放电特性的GQ/11耦合受体控制中的作用，以及在单细胞水平上的神经递质的释放，以及通过单细胞水平的释放，以及通过体外模型的心肌细胞对同情神经元的表达的计量。我们将使用一个异源表达系统，其中M通道，受体和信号分子在中国仓鼠卵巢（CHO）细胞中表达，大鼠和小鼠上颈神经节（SCG）的制剂和Nodose Ganglia（NG）神经元，以及SCG神经元和scG神经元和心脏室性心脏造型的培养。要使用的技术包括荧光共振能量转移（FRET），总内反射荧光（TIRF），共聚焦显微镜，斑块钳电生理学，碳纤维安培时间表和视频成像。我们渴望发现针对受体和M型K+通道的特异性赋予AKAP79/150的机制，以及其在塑造神经生理学和神经递质释放性能的功能作用。公共卫生相关性：在生物信号传导中构成一个共同主题的信号传导途径是如何调节神经系统的基础，包括情绪状态和情绪的改变，个性以及记忆的获取和使用。我们将研究M型钾离子通道的调节，该通道在调节神经细胞兴奋性方面起着显着作用，这是由A-激酶锚定蛋白AKAP79/150介导的。 AKAP79/150在M通道活性中的机制和功能作用的阐明将阐明神经系统在健康和疾病中的功能。