Regulating BDNF Action in Postnatal Development.

调节 BDNF 在产后发育中的作用。

• 批准号: 8206532
• 负责人: BARBARA L HEMPSTEAD
• 金额: $ 36.02万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8206532
• 关键词: Adolescent; Affect; Animals; Anxiety; Axon; Binding; Biological; Brain-Derived Neurotrophic Factor; Cleaved cell; Dendrites; Detection; Development; Electrons; Epitopes; Exhibits; Family member; Genetic; Goals; Growth; Health; In Vitro; Knock-in Mouse; Learning; Light; Location; Mediating; Memory; Mental Depression; Microscopic; Molecular; Molecular Chaperones; Morphology; Mus; Neonatal; Neuraxis; Neuronal Differentiation; Neuronal Plasticity; Neurons; Neurotrophic Tyrosine Kinase Receptor Type 2; Pathway interactions; Play; Process; Protein Isoforms; Protein Tyrosine Kinase; Reagent; Role; Secretory Vesicles; Signal Transduction; Site; Sorting - Cell Movement; Structure; Synapses; Synaptic Cleft; Synaptic Transmission; Synaptic plasticity; Testing; Time; Vertebral column; Vesicle; critical period; density; in vivo; member; nerve stem cell; neuronal survival; neurotransmission; neurotrophic factor; overexpression; postnatal; postsynaptic; presynaptic; promoter; receptor; research study; small hairpin RNA; sortilin; synaptogenesis; tool; trafficking; transcriptional coactivator p75

DESCRIPTION (provided by applicant): Neurotrophins induce structural and functional changes in neurons to modulate synaptic efficacy; our long term goal is to identify molecular mechanisms that regulate BDNF targeting and release at synapses to modulate neuronal structure and neurotransmission. BDNF is initially synthesized as a precursor form (proBDNF) that is sorted to a regulated secretory pathway, and released in an activity-dependent manner. When proBDNF is released at the synapse, it can bind to p75 receptors to induce LTD, and potentially reduce spine density and dendritic complexity. However, if proBDNF is converted to mature BDNF in the secretory vesicle or synaptic cleft, TrkB is selectively activated to enhance synaptic transmission and promote axonal branching and dendritic growth. TrkB receptors are present both pre- and post-synaptically in the Schaffer collateral pathway, and mature BDNF can activate both pre- and post-synaptic TrkB receptors to facilitate neurotransmission. Thus, the molecular mechanisms that regulate conversion of proBDNF to mature BDNF, and that regulate intracellular trafficking to dendrites or axons are critical to modulate structural and functional neuronal plasticity. We have developed new genetic tools to facilitate detection of endogenous BDNF, and identified new sorting receptors that direct BDNF intracellular trafficking. Specifically, we have generated knock-in mice that express HA tagged BDNF to markedly enhance detection of endogenous BDNF. We have also identified intracellular chaperones, including sortilin, and other sortilin family members that bind to proBDNF. With these tools, three aims are proposed to dissect BDNF trafficking, cleavage, and depolarization dependent release: (1) Using neurons from the BDNF-HA tagged mouse, identify if conversion of proBDNF to mature BDNF occurs during synthesis and sorting to secretory vesicles, or whether conversion occurs following vesicle release. We predict that the location of BDNF conversion may differ among neuronal subtypes and across early postnatal time points when synaptic connections are being refined and synaptogenesis is robust. (2) We will investigate how sortilin family members alter intracellular cleavage of proBDNF and modulate pro- vs. mature BDNF release in neuronal cultures. (3) We will identify the sortilin family members that chaperone proBDNF to the constitutive or regulated secretory pathway, and to dendrites or axons. We posit that different sortilin family members direct intracellular trafficking to different subcellular compartments and regulate cleavage to mature BDNF and its release. These studies will rely on the BDNF- HA tagged mouse, and overexpression or shRNA knockdown of different chaperones. These studies will identify molecular mechanisms that regulate BDNF processing and trafficking, to induce structural and functional changes in the developing postnatal central nervous system. PUBLIC HEALTH RELEVANCE: The neurotrophin BDNF plays critical roles in regulating neuronal survival, morphology, and activity-dependent forms of synaptic plasticity. Interestingly, BDNF is initially synthesized as a precursor, proBDNF, which exhibits biological actions that are distinct, and even opposing those of its mature form. Here we will identify molecular mechanisms that (1) regulate the trafficking of BDNF to synapses, (2) regulate the conversion of proBDNF to mature BDNF and (3) determine how these processes are regulated in early postnatal development, a critical period of robust synaptogenesis. It is well established that even modest changes in the level of BDNF secreted by neurons has significant effects on learning, memory, anxiety states and depression, and these studies will test the hypothesis that BDNF trafficking and release is regulated by differential use of intracellular chaperones.

描述（由申请人提供）：神经营养因子诱导神经元的结构和功能变化以调节突触功效;我们的长期目标是鉴定调节BDNF靶向和在突触处释放以调节神经元结构和神经传递的分子机制。BDNF最初合成为前体形式（proBDNF），其被分选到受调节的分泌途径，并以活性依赖性方式释放。当proBDNF在突触处释放时，它可以与p75受体结合以诱导LTD，并可能降低棘密度和树突复杂性。然而，如果proBDNF在分泌囊泡或突触间隙中转化为成熟BDNF，则TrkB被选择性地激活以增强突触传递并促进轴突分支和树突生长。TrkB受体在Schaffer侧支通路中存在于突触前和突触后，成熟的BDNF可以激活突触前和突触后TrkB受体以促进神经传递。因此，调节proBDNF向成熟BDNF转化以及调节细胞内向树突或轴突运输的分子机制对于调节结构和功能神经元可塑性至关重要。我们已经开发了新的遗传工具，以促进内源性BDNF的检测，并确定了新的分选受体，直接BDNF细胞内运输。具体地，我们已经产生了表达HA标记的BDNF的敲入小鼠，以显著增强内源性BDNF的检测。我们还确定了细胞内伴侣，包括分拣蛋白，和其他分拣蛋白家族成员结合的proBDNF。利用这些工具，提出了三个目的来剖析BDNF运输、切割和去极化依赖性释放：（1）使用来自BDNF-HA标记的小鼠的神经元，鉴定是否在合成和分选成分泌囊泡期间发生proBDNF向成熟BDNF的转化，或者是否在囊泡释放之后发生转化。我们预测，BDNF转换的位置可能不同的神经元亚型和出生后早期的时间点时，突触连接正在完善和突触是强大的。(2)我们将研究分拣蛋白家族成员如何改变脑源性神经营养因子原的细胞内切割和调节神经元培养物中前与成熟脑源性神经营养因子的释放。(3)我们将确定sortilin家族成员，伴侣proBDNF的组成或调节分泌途径，树突或轴突。我们证实不同的分拣蛋白家族成员直接将细胞内运输到不同的亚细胞区室，并调节成熟BDNF的切割及其释放。这些研究将依赖于BDNF-HA标记的小鼠，以及不同分子伴侣的过表达或shRNA敲低。这些研究将确定调节BDNF加工和运输的分子机制，以诱导发育中的出生后中枢神经系统的结构和功能变化。 公共卫生关系：神经营养因子BDNF在调节神经元存活、形态和突触可塑性的活性依赖形式中起关键作用。有趣的是，BDNF最初是作为前体proBDNF合成的，proBDNF表现出不同的生物学作用，甚至与其成熟形式相反。在这里，我们将确定分子机制，（1）调节BDNF的运输到突触，（2）调节proBDNF转化为成熟的BDNF和（3）确定这些过程是如何在出生后早期发育，一个关键时期的强大的突触发生调节。已经确定的是，即使是神经元分泌的BDNF水平的适度变化也对学习、记忆、焦虑状态和抑郁具有显著影响，并且这些研究将检验BDNF运输和释放受细胞内伴侣的差异使用调节的假设。