Molecular Analysis of BDNF-TrkB Regulation of Synapse Formation and Maintenance

BDNF-TrkB 突触形成和维持调节的分子分析

• 批准号: 9056348
• 负责人: Eric J Huang
• 金额: $ 34.67万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9056348
• 关键词: Acute; Addictive Behavior; Adult; Affect; Alleles; Appearance; Behavior; Binding; Biochemical; Brain; Brain region; Brain-Derived Neurotrophic Factor; Cell Culture Techniques; Cell Surface Proteins; Cell surface; Cells; Cerebellum; Detergents; Development; Disease; Excision; Excitatory Synapse; Genes; Genetic study; Health; Heart Diseases; Human; Human Genetics; Image; In Vitro; Inhibitory Synapse; Kinetics; Knowledge; Ligands; LoxP-flanked allele; Maintenance; Mediating; Memory; Mental Retardation; Mental disorders; Microscopy; Modeling; Molecular; Molecular Analysis; Moods; Mutate; Mutation; Names; Obesity; Optics; Performance; Phosphorylation; Post-Translational Protein Processing; Protein Biosynthesis; Proteins; Proteomics; Receptor Protein-Tyrosine Kinases; Regulation; Resistance; Resolution; Role; Running; Scaffolding Protein; Schizophrenia; Side; Signal Transduction; Site; Staging; Synapses; Walking; Work; expectation; gephyrin; granule cell; in vivo; insight; microscopic imaging; molecular assembly/self assembly; motor control; neuronal circuitry; neuronal survival; neurotrophic factor; new therapeutic target; postsynaptic; presynaptic; protein degradation; protein function; protein protein interaction; receptor; reconstruction; scaffold; synaptogenesis

DESCRIPTION (provided by applicant): We will characterize mechanisms through which TrkB regulates inhibitory synapse formation and maintenance in the cerebellum with the expectation that the insights obtained will prove important for understanding the role of TrkB in many regions of the brain. Extending our prior demonstrations that TrkB controls inhibitory synapse formation throughout the cerebellum and has important pre- and postsynaptic cell- autonomous roles, we will use high resolution stochastic optical reconstruction microscopy (STORM) imaging to characterize the localizations of inhibitory synapse-associated cell surface and synaptic scaffold proteins and determine the effects of TrkB activation and inhibition on their presence at the synapse. Extending our recent observation that adult TrkB activity is required to maintain inhibitory synapses and that reactivation of TrkB signaling after earlier inhibition results in reappearance at the synapse of many synaptic proteins, we will examine the effects of adult TrkB inhibition and reactivation on the molecular composition of these synapses. Using cell culture we will examine in more detail the appearance and disappearance of proteins associated with the synapse following TrkB activation and inactivation. We shall determine whether TrkB functions in part through control of protein synthesis or turnover. We shall also examine the effects of TrkB activity on the kinetics of gephyrin stability, insertion and removal a synaptic sites in cell culture. Finally, we more critically examine our model that TrkB acts in par through control molecular assembly of the proteins that form the synaptic scaffold. We will determine the effects of TrkB activation and inactivation in vivo and in vitro on the distribution f gephyrin and other postsynaptic scaffold proteins in detergent soluble and resistant fractions, the interactions of these proteins with binding partners and on phosphorylation and other post-translational modifications.

描述（由申请人提供）：我们将描述TrkB调节小脑中抑制性突触形成和维持的机制，期望获得的见解将证明对理解TrkB在大脑许多区域中的作用是重要的。我们之前已经证明TrkB控制整个小脑的抑制性突触形成，并具有重要的突触前和突触后细胞自主作用，我们将使用高分辨率随机光学重建显微镜（STORM）成像来表征抑制性突触相关细胞表面和突触支架蛋白的定位，并确定TrkB激活和抑制对它们在突触中的存在的影响。扩展我们最近的观察，即维持抑制性突触需要成体TrkB活性，并且在早期抑制后TrkB信号的再激活导致许多突触蛋白的突触重新出现，我们将研究成体TrkB抑制和再激活对这些突触分子组成的影响。通过细胞培养，我们将更详细地研究TrkB激活和失活后与突触相关的蛋白质的出现和消失。我们将确定TrkB是否部分通过控制蛋白质合成或周转发挥作用。我们还将研究TrkB活性对细胞培养中卟啉稳定性、插入和移除突触位点动力学的影响。最后，我们更严格地检验了我们的模型，即TrkB通过控制形成突触支架的蛋白质的分子组装而发挥作用。我们将在体内和体外确定TrkB激活和失活对洗涤剂可溶性和抗性部分中卟啉和其他突触后支架蛋白分布的影响，这些蛋白与结合伙伴的相互作用以及磷酸化和其他翻译后修饰。