Molecular Mechanisms Of Synapse Development And Plasticity

突触发育和可塑性的分子机制

基本信息

批准号：
7734707
负责人：
Bai Lu
金额：
$ 122.11万
依托单位：
EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7734707
关键词：
3&apos Untranslated Regions Acute Address Area Autopsy BDNF gene Behavioral Biological Brain Brain-Derived Neurotrophic Factor Cell Survival Cell model Cell physiology Cells Cellular biology Chemosensitization Chromosome Pairing Class Code Cognition Cognition Disorders Cognitive Communication Complex Dendrites Dendritic Spines Development Diabetes Mellitus Disease Disruption Endopeptidases Environment Exhibits Exons Extinction (Psychology)Fire - disasters Frequencies Fright Gene Expression Genes Genetic Genetic Transcription Goals Green Fluorescent Proteins Hippocampus (Brain)Human Image Impairment Length Light Link Localized Long-Term Depression Long-Term Potentiation Maps Mediating Memory Mental Depression Mental disorders Messenger RNA Methods Modern Medicine Molecular Morphology Mus Mutant Strains Mice NGFR Protein Neurons Parvalbumins Pathogenesis Pattern Peptide Hydrolases Pharmaceutical Preparations Physiological Physiology Play Population Post-Traumatic Stress Disorders Potassium Channel Prefrontal Cortex Primates Protein Biosynthesis Protein Isoforms Protein Overexpression Protein Tyrosine Kinase Proteins Psychotic Disorders Range Regulation Relative (related person)Research Reversal Learning Risk Role Schizophrenia Short-Term Memory Signal Transduction Structure Susceptibility Gene Synapses Synaptic Transmission Synaptic plasticity System Techniques Terminator Codon Time Transcript Vertebral column Work Yin-Yang base brain tissue cognitive function executive function extracellular genetic association genetic linkage genetic manipulation hippocampal pyramidal neuron human disease inhibitory neuron interest long term memory mutant neuronal cell body neuronal survival neurotrophic factor novel postsynaptic promoter receptor relating to nervous system secretory protein synaptic depression synaptic function tool trafficking translational approach

项目摘要

This year we have made a significant progress in revealing several new aspects of neurotrophin functions. 1) Distinct Role of Long 3UTR BDNF mRNA in Spine Morphology and Synaptic Plasticity in Hippocampal Pyramidal Neurons A quite common and yet puzzling phenomenon in cell biology is the existence of transcripts that code for exactly the same protein but with different 3 untranslated region (3UTR). One such example is the messenger RNA for BDNF. There are two pools of BDNF mRNAs in neurons with similar abundance: one with a short 3UTR) and the other with a long 3UTR. It is not known why a neuron would need two mRNAs of different length, coding for the same BDNF protein. We have now provided evidence suggesting that mRNAs of different 3UTR confer differential functions due to their distinct subcellular localization. We show that the short 3UTR BDNF mRNAs are restricted in neuronal cell bodies whereas the long 3UTR BDNF mRNAs are also localized in dendrites. In a mouse mutant where the long 3UTR is truncated, dendritic targeting of BDNF mRNAs is impaired. There is little BDNF in hippocampal dendrites despite normal levels of total BDNF protein. This mutant exhibits deficits in pruning and enlargement of dendritic spines, as well as selective impairment in long-term potentiation in dendrites, but not somata, of hippocampal neurons. These results are significant for several reasons. First, it elucidates a cellular mechanism why BDNF, a key regulator for brain development and plasticity, could elicit diverse cellular functions ranging from cell survival to synaptic transmission. Second, it provides clear experimental evidence that dendritically localized mRNAs play crucial roles in regulating spine morphologies, a long-sought result in the field of local protein synthesis and synaptic plasticity. Finally, as far as we know, this is the first demonstration that different transcripts with the same coding sequence have distinct roles in a cell. 2) Activity-dependent BDNF transcription and secretion In the first study, we address the role of activity-dependent BDNF gene expression. Transcription of the BDNF gene is controlled by multiple promoters, which drive the expression of multiple transcripts that encode for the same protein. Promoter-IV contributes significantly to activity-dependent BDNF transcription. We have generated promoter-IV mutant mice (BDNF-KIV) by inserting the GFP gene followed by a stop codon into exon-IV. This genetic manipulation resulted in disruption of promoter-IV-mediated BDNF expression, particularly in the prefrontal cortex (PFC), an area involved in working memory and other executive functions. Interestingly, the BDNF-KIV exhibited a selective impairment in parvalbumin-positive GABAergic neurons and inhibitory but not excitatory postsynaptic currents in the PFC, leading to an aberrant spike-timing dependent synaptic potentiation (STDP). Behaviorally, BDNF-KIV mice are significantly impaired in PFC-mediated reversal learning and fear memory extinction, but not working memory. These results demonstrate the importance of promoter-IV-dependent BDNF transcription in GABAergic function, and reveal an unexpected role of BDNF in behavioral perseverance. Our study may shed light on the pathogenesis of several cognitive disorders in which perseverance is prominent, including schizophrenia and Post-traumatic stress disorder (PTSD). In the second study, we address the role of activity-dependent BDNF gene expression. We have previously shown that pro- and mature BDNF often elicit opposing biological effects. For instance, mature BDNF (mBDNF) is critical for long-term potentiation (LTP) induced by high-frequency stimulation (HFS), whereas proBDNF facilitate long-term depression (LTD) induced by low-frequency stimulation (LFS). Since mBDNF is derived from proBDNF by endoproteolytic cleavage, mechanisms regulating the cleavage of proBDNF may control the direction of BDNF regulation. Using methods that selectively detect proBDNF or mBDNF, we show that LFS induced predominant proBDNF secretion in cultured hippocampal neurons. In contrast, HFS preferentially increased extracellular mBDNF. Inhibition of extracellular, but not intracellular cleavage of proBDNF greatly reduced HFS-induced extracellular mBDNF. Moreover, HFS, but not LFS, selectively induced the secretion of tPA, a key protease involved in extracellular proBDNF to mBDNF conversion. Thus, high-frequency neuronal activity controls the ratio of extracellular proBDNF/mBDNF by regulating the secretion of extracellular proteases. Our study demonstrates activity-dependent control of extracellular proteolytic cleavage of a secretory protein, and reveals an important mechanism that controls diametrically opposed functions of BDNF isoforms. In addition to revealing the first example of how neuronal activity can control the cleavage of a secreted protein, this work provides useful tools to study distinct function of proBDNF and mBDNF. 3) Studies of genes involved in schizophrenia An important goal of modern medicine is to understand the basic mechanisms underlying common and complex human diseases, such as diabetes and schizophrenia. Conventional genetic linkage and association studies have confirmed that these diseases involve multiple genes, and each gene elicits very small effects across populations. Moreover, because of the complexity of genetic factors and their interaction with the environment, it is often difficult to link genetic findings to important biological or mechanistic aspects of the illness. To overcome these obstacles, we have taken a powerful translational approach that dissects the complex phenomenology of psychosis into several neural system and molecular components and maps genetic association onto these multiple levels of analysis. Using this approach, we identify a potentially new class of schizophrenia susceptibility genes, the KCNH2 potassium channel, but more importantly, a novel isoform of this type of potassium channel that is primate and brain specific and increased 2.5 fold in schizophrenia brain tissue. Expression of this novel isoform is predicted by risk associated SNPs in the gene, even in normal brain tissue, and risk SNPs also predict cognition and related brain physiology in normal subjects. Postmortem expression analysis shows a 2.5-fold increase in Isoform 3.1 relative to KCNH2-1A in schizophrenic hippocampus. Structurally, Isoform 3.1 lacks most of the PAS domain critical for slow channel deactivation. Electrophysiological characterization in primary cortical neurons reveals that overexpression of Isoform 3.1 results in a rapidly deactivating K+ current and a high-frequency, non-adapting firing pattern. These results identify a novel KCNH2 channel involved in cortical physiology, cognition, and psychosis, providing a potential new target for psychotherapeutic drugs.

今年，我们在揭示神经营养蛋白功能的几个新方面取得了重大进展。 1）长3UTR BDNF mRNA在海马锥体神经元中的脊柱形态和突触可塑性中的独特作用细胞生物学中的一种非常普遍但令人困惑的现象是成绩单的存在，这些转录本与完全相同的蛋白质编码，但具有不同的3个未翻译区域（3UTR）。一个这样的例子是BDNF的Messenger RNA。神经元中有两个BDNF mRNA池，具有相似的丰度：一个短3UTR），另一个具有长3UTR。尚不清楚为什么神经元需要两个不同长度的mRNA，以编码相同的BDNF蛋白。现在，我们提供了证据，表明不同3UTR的mRNA由于其独特的亚细胞定位而赋予差异功能。我们表明，短3UTR BDNF mRNA受到神经元细胞体的限制，而长3UTR BDNF mRNA也位于树突中。在将长3UTR截断的小鼠突变体中，bdnf mRNA的树突状靶向受损。尽管总BDNF蛋白水平正常，但海马树突中的BDNF很少。该突变体在树突状刺的修剪和扩大方面表现出缺陷，以及在海马神经元的树突增强（而不是somata）长期增强中的选择性损害。这些结果很重要，原因有几个。首先，它阐明了一种细胞机制，为什么BDNF是脑发育和可塑性的关键调节剂，可能会引起从细胞存活到突触传播的各种细胞功能。其次，它提供了明确的实验证据，表明树突定位的mRNA在调节脊柱形态中起着至关重要的作用，这是局部蛋白质合成和突触可塑性领域的长期追求的。最后，据我们所知，这是第一次证明具有相同编码序列的不同转录本在单元中具有不同的作用。 2）依赖于活动的BDNF转录和分泌在第一项研究中，我们解决了依赖活性BDNF基因表达的作用。 BDNF基因的转录由多个启动子控制，这些启动子驱动了对同一蛋白质编码的多个转录本的表达。启动子-IV对活动依赖性BDNF转录做出了重大贡献。我们通过将GFP基因随后将终止密码子插入外显子IV来产生启动子IV突变小鼠（BDNF-KIV）。这种遗传操作导致启动子IV介导的BDNF表达中断，特别是在前额叶皮层（PFC）中，这是参与工作记忆和其他执行功能的领域。有趣的是，BDNF-KIV在白蛋白阳性阳性GABA能神经元和PFC中抑制性抑制性神经元和抑制性抑制性抑制作用中表现出选择性损害，导致异常的尖峰触发依赖性突触增强（STDP）。从行为上讲，BDNF-KIV小鼠在PFC介导的逆转学习和恐惧记忆灭绝中受到显着损害，但没有工作记忆。这些结果证明了启动子IV依赖性BDNF转录在GABA能函数中的重要性，并揭示了BDNF在行为毅力中的意外作用。我们的研究可能会阐明几种毅力突出的认知障碍的发病机理，包括精神分裂症和创伤后应激障碍（PTSD）。在第二项研究中，我们解决了依赖活性的BDNF基因表达的作用。我们以前已经表明，亲和成熟的BDNF经常引起相反的生物学作用。例如，成熟的BDNF（MBDNF）对于高频刺激（HFS）引起的长期增强（LTP）至关重要，而ProbDNF促进了由低频刺激（LFS）诱导的长期抑郁症（LTD）。由于MBDNF是通过内蛋白溶解裂解从ProbDNF得出的，因此调节ProbDNF裂解的机制可能控制BDNF调节的方向。使用选择性检测ProbDNF或MBDNF的方法，我们表明LFS在培养的海马神经元中诱导了主要的ProbDNF分泌。相反，HFS优先增加了细胞外MBDNF。抑制细胞外的，但不能大大降低HFS诱导的细胞外MBDNF。此外，HFS而非LFS选择性地诱导了TPA的分泌，TPA是针对MBDNF转换涉及的关键蛋白酶。因此，高频神经元活性通过调节细胞外蛋白酶的分泌来控制细胞外探针/MBDNF的比率。我们的研究表明，分泌蛋白的细胞外蛋白水解裂解的活性依赖性控制，并揭示了一种重要的机制，该机制控制了BDNF同工型的直截了当功能。除了揭示神经元活性如何控制分泌蛋白的裂解的第一个示例外，这项工作还提供了研究ProbDNF和MBDNF不同功能的有用工具。 3）研究中涉及精神分裂症的基因现代医学的一个重要目标是了解糖尿病和精神分裂症等常见和复杂人类疾病的基本机制。常规的遗传联系和关联研究已经证实，这些疾病涉及多种基因，每个基因在人群中产生非常小的影响。此外，由于遗传因素的复杂性及其与环境的相互作用，通常很难将遗传发现与疾病的重要生物学或机械方面联系起来。为了克服这些障碍，我们采取了一种强大的翻译方法，将精神病的复杂现象学剖析到几个神经系统和分子成分，并将遗传关联映射到这些多个级别的分析中。使用这种方法，我们确定了一种潜在的新型精神分裂症易感基因，KCNH2钾通道，但更重要的是，这种类型的钾通道的新型同工型是灵长类动物和大脑特异性的，并且在精神分裂症脑组织中增加了2.5倍。即使在正常脑组织中，这种新型同工型的表达也可以通过基因中的风险相关SNP来预测，而风险SNP也可以预测正常受试者中认知和相关的脑生理。验尸表达分析表明，在精神分裂症海马中，同工型3.1的同工型3.1增加了2.5倍。从结构上讲，同工型3.1缺少大多数PAS域对于慢速通道停用至关重要。原发性皮质神经元中的电生理表征表明，同工型的过表达3.1导致迅速失活的K+电流和高频，非适应性放电模式。这些结果确定了涉及皮质生理，认知和精神病的新型KCNH2通道，为心理治疗药物提供了潜在的新靶标。

项目成果

期刊论文数量（43）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

BDNF-induced recruitment of TrkB receptor into neuronal lipid rafts: roles in synaptic modulation.

DOI：
10.1083/jcb.200404106
发表时间：
2004-12-20
期刊：
The Journal of cell biology
影响因子：
0
作者：
Suzuki S;Numakawa T;Shimazu K;Koshimizu H;Hara T;Hatanaka H;Mei L;Lu B;Kojima M
通讯作者：
Kojima M

Signaling mechanisms mediating BDNF modulation of synaptic plasticity in the hippocampus.

介导海马突触可塑性 BDNF 调节的信号机制。