Dysregulation of Cav1.2 by beta amyloid peptide

β 淀粉样肽导致 Cav1.2 失调

• 批准号: 10521735
• 负责人: JOHANNES W HELL
• 金额: $ 161.05万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-15 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10521735
• 关键词: Adenylate Cyclase; Adrenergic Receptor; Affect; Alzheimer' s Disease; Alzheimer' s disease model; Amyloid beta-Protein; Antibodies; Binding; Biotinylation; Brain; C-terminal; Chronic; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dendritic Spines; Development; Endocytosis; Endosomes; Etiology; Event; Excision; Excitatory Postsynaptic Potentials; Functional disorder; Gene Expression; Genetic Transcription; Glutamates; Hippocampus (Brain); Image; Impairment; Intervention; Isradipine; Knock-in; Knock-in Mouse; Label; Left; Link; Long-Term Depression; Long-Term Potentiation; Measures; Mediating; Mus; Neuronal Dysfunction; Neurons; Norepinephrine; Pathway interactions; Peptides; Permeability; Pharmacology; Phase; Phosphorylation; Phosphorylation Site; Proteins; Rattus; Recycling; Regulation; Regulator Genes; Reporting; Rodent; Rodent Model; Role; Science; Secretory Vesicles; Serine; Signal Pathway; Signal Transduction; Site; Surface; Symptoms; Synapses; Synaptic Transmission; Synaptic plasticity; Testing; Toxic effect; Up-Regulation; Western Blotting; abeta accumulation; aged; amyloid peptide; density; improved; inhibitor; innovation; mouse model; neuron loss; neuronal excitability; neurotoxic; neurotoxicity; peptide L; postsynaptic; protein kinase A kinase; recruit; reuptake; synaptic function; trafficking; uptake; Adenylate Cyclase; Adrenergic Receptor; Affect; Alzheimer' s Disease; Alzheimer' s disease model; Amyloid beta-Protein; Antibodies; Binding; Biotinylation; Brain; C-terminal; Chronic; Clinical; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dendritic Spines; Development; Endocytosis; Endosomes; Etiology; Event; Excision; Excitatory Postsynaptic Potentials; Functional disorder; Gene Expression; Genetic Transcription; Glutamates; Hippocampus (Brain); Image; Impairment; Intervention; Isradipine; Knock-in; Knock-in Mouse; Label; Left; Link; Long-Term Depression; Long-Term Potentiation; Measures; Mediating; Mus; Neuronal Dysfunction; Neurons; Norepinephrine; Pathway interactions; Peptides; Permeability; Pharmacology; Phase; Phosphorylation; Phosphorylation Site; Proteins; Rattus; Recycling; Regulation; Regulator Genes; Reporting; Rodent; Rodent Model; Role; Science; Secretory Vesicles; Serine; Signal Pathway; Signal Transduction; Site; Surface; Symptoms; Synapses; Synaptic Transmission; Synaptic plasticity; Testing; Toxic effect; Up-Regulation; Western Blotting; abeta accumulation; aged; amyloid peptide; density; improved; inhibitor; innovation; mouse model; neuron loss; neuronal excitability; neurotoxic; neurotoxicity; peptide L; postsynaptic; protein kinase A kinase; recruit; reuptake; synaptic function; trafficking; uptake

Abstract Dysregulation of Cav1.2 by beta amyloid peptide L-type Ca2+ channels (LTCC) are key regulators of gene transcription, neuronal excitability, and synaptic functions. Cav1.2 is the prevalent LTCC in brain (Hell et al., 1993, JCB 123, 949-962). Chronically increased Ca2+ influx via LTCCs has been implicated early on in senile symptoms and Alzheimer’s disease (AD) (e.g., Science 272, 1017). We found that aged rats have significantly increased PKA-mediated phosphorylation of Cav1.2 on Serine 1928 (Davare and Hell, 2003, PNAS 100, 16018-23), which increases Cav1.2 channel activity (Qian, …, Hell, 2017, Sci Sig 10, eaaf9659). We also found that Cav1.2 forms a unique signaling complex with the b2 adrenergic receptor (b2 AR) that also contains all other proteins necessary for the cAMP-mediated regulation of Cav1.2 (e.g., Davare et al., 2001, Science 293, 98), making Cav1.2 a prime target for b2 AR signaling. b amyloid peptide 1-42 (Ab) is a hallmark of AD. Oligomeric Abo stimulates the b2 AR. Supportive evidence from single channel recording indicates that Abo increase Cav1.2 activity via the b2 AR. We hypothesize that this increase is mediated by phosphorylation of S1928, the main PKA site of Cav1.2, and triggers downstream events that ultimately cause neuronal damage. Aim 1 is to test whether Cav1.2 dysregulation by Abo via Cav1.2- associated b2 AR occurs in dendrites and spines (Ca2+ imaging), whether it is in part mediated by increased surface insertion of Cav1.2, and whether blocking this signaling pharmacologically or in innovative S1928A knock-in (KI) mice alleviates Abo neurotoxicity. Aim 2 is to test whether Abo - b2 AR - S1928 signaling stimulates surface insertion of Ca2+ permeable (CP) AMPARs (GluA1 homomers) and blocking CP-AMPARs alleviates Abo neurotoxicity. We will use cutting edge live imaging of SEP-tagged GluA1 and GluA2 and analyze field EPSPs and mEPSC before and after inhibition of glutamate re-uptake to measure AMPAR activity in the perisynaptic space. Aim 3 will test the role of Abo - b2 AR - S1928 signaling in augmentation of long-term depression by Abo, and in long-term potentiation, which is impaired by Abo. Crossing our S1928A KI mice with highly amyloidogenic 5xFAD mice will show whether Abo - b2 AR - S1928 signaling is important for Ab-induced dysfunction of Cav1.2 activity and synaptic transmission and its plasticity. This project is focused on what we hypothesize is an early effect of Abo (minute range) that triggers subsequent neurotoxic events because we need to understand all aspects of Abo toxicity. Also, as an early event the Abo - b2 AR - S1928 signaling constitutes a potential target for early pharmacological intervention in AD. We will to some degree explore later events downstream of Cav1.2 dysregulation by testing whether neurotoxicity in the 6-48h range can be improved by blockers of Abo - b2AR - CaV1.2 signaling and of CP-AMPAR in neuronal cultures. We will also explore whether blocking this signaling in the 5xFAD mouse model of AD will improve synapse function in these mice. The development of our peptide that disrupts binding of the b2 AR to Cav1.2 is highly innovative because it allows to specifically block signaling by Abo via the b2 AR without affecting other signaling pathways that involve the b2 AR.

抽象的 Beta淀粉样胡椒的CAV1.2失调 L型CA2+通道（LTCC）是基因转录，神经元和突触的关键调节剂 功能。 CAV1.2是大脑中普遍存在的LTCC（Hell等，1993，JCB 123，949-962）。长期增加 通过LTCCS的Ca2+影响已经与老年症状和阿尔茨海默氏病（AD）有关（例如， 科学272，1017）。我们发现，老年大鼠已显着增加了PKA介导的磷酸化 Cav1.2在丝氨酸1928（Davare and Hell，2003，PNAS 100，16018-23）上，增加CAV1.2频道活动 （Qian，…，地狱，2017年，Sci Sig 10，EAAF9659）。我们还发现Cav1.2与 B2肾上腺素受体（B2 AR），还包含CAMP介导的所有其他蛋白质 CAV1.2的调节（例如，Davare等，2001，Science 293，98），使CAV1.2成为B2 AR信号传导的主要目标。 B淀粉样蛋白肽1-42（AB）是AD的标志。寡聚ABO刺激B2 AR。支持的证据 单个通道记录表明ABO通过B2 AR增加CAV1.2活性。我们假设这是 增加是由S1928的磷酸化（CAV1.2的主要PKA部位）和下游事件触发的磷酸化介导 最终导致神经元损害。 AIM 1是测试CAV1.2 ABO是否通过CAV1.2-进行失调 相关的B2 AR发生在树突和棘中（Ca2+成像），是否部分是由增加的 CAV1.2的表面插入，以及是否在物理上还是在创新的S1928A中阻止此信号 敲入（Ki）小鼠减轻了ABO神经毒性。 AIM 2是测试ABO -B2 AR -S1928信号刺激是否刺激 Ca2+渗透性（CP）AMPAR（GLUA1同源物）的表面插入并阻止CP-Ampars减轻ABO 神经毒性。我们将使用SEP标记的GLUA1和GLUA2的尖端实时成像并分析现场EPSP MEPSC在抑制谷氨酸之前和之后重新摄取以测量过度突触中的AMPAR活性 空间。 AIM 3将测试ABO -B2 AR -S1928的作用 在长期增强中，ABO损害了。用高度淀粉样蛋白跨越我们的S1928A Ki小鼠 5xFAD小鼠将显示ABO -B2 AR -S1928信号是否对于AB诱导的CAV1.2的功能障碍很重要 活动和突触传播及其可塑性。该项目的重点是我们假设是早期的 ABO（分钟范围）的影响会触发随后的神经毒性事件，因为我们需要了解所有事件 ABO毒性的各个方面。另外，作为早期事件，ABO -B2 AR -S1928信号构成潜在目标 用于AD的早期药理干预。我们将在某种程度上探索Cav1.2下游的以后事件 通过测试6-48H范围内的神经毒性是否可以通过ABO -B2AR的阻滞剂来改善失调。 CAV1.2神经元培养中的信号传导和CP-Ampar。我们还将探讨是否将此信号阻止 AD的5xFAD小鼠模型将改善这些小鼠的突触功能。我们肽的发展 破坏B2 AR与CAV1.2的结合是高度创新的，因为它允许专门阻止信号 通过ABO通过B2 AR不影响涉及B2 AR的其他信号通路。