Synaptic Dissection of Cell Adhesion Molecule Function within Subicular Circuits

毛细血管内细胞粘附分子功能的突触解剖

• 批准号: 8827859
• 负责人: Jason Aoto
• 金额: $ 9.27万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8827859
• 关键词: Acute; Adverse effects; Affect; Alternative Splicing; Animals; Behavior; Binding; Biological; Biological Assay; Brain; Cell Adhesion Molecules; Cells; Complex; Corpus striatum structure; DRD2 gene; Data; Development; Disease; Dissection; Dopamine; Dorsal; Economic Burden; Effectiveness; Electrophysiology (science); Family; Fiber; Functional disorder; Future; Gene Expression Profiling; Genome; Genomics; Goals; Grant; Health; Hippocampus (Brain); Human; Impaired cognition; Impulsivity; Individual; Infection; Injection of therapeutic agent; Interneurons; Investigation; Label; Ligands; Light; Link; Measures; Mediating; Mental Depression; Mental disorders; Mentors; Messenger RNA; Methods; Modeling; Molecular; Mus; Mutant Strains Mice; Mutation; Neurons; Nucleus Accumbens; Output; Patients; Phase; Play; Population; Probability; Property; Proteins; RNA Sequences; RNA Splicing; Recruitment Activity; Regulation; Relative (related person); Research; Rewards; Rhodopsin; Role; Schizophrenia; Shapes; Site; Slice; Specific qualifier value; Stress; Structure; Surface; Synapses; Synaptic Transmission; Synaptic plasticity; Techniques; Technology; Training; Virus; base; cell type; differential expression; dopamine system; hippocampal pyramidal neuron; in vivo; insight; mutant; neuropsychiatry; next generation; novel; novel therapeutics; optogenetics; postsynaptic; presynaptic; recombinase; research study; response; screening; small hairpin RNA; social; synaptic function; tool; transcriptome sequencing; transcriptomics; transmission process; treatment strategy; virus Cre recombinase

DESCRIPTION (provided by applicant): Increasing evidence suggests that abnormalities in synaptic transmission may be a key mechanism underlying some neuropsychiatric disorders. How might synaptic properties be specified an maintained? Synaptic cell adhesion molecules (SCAMs) are primary candidates that play a critical role regulating synapse function. Not surprisingly, genomic studies have found mutations in neurexin-3 (Nrx3) a prototypical presynaptic SCAM that is part of the neurexin/neuroligin complex. that are linked to Schizophrenia (SZ) and reward-seeking behavior. These disorders are associated with an enormous social and economic burden and share a common pathophysiological basis - dopamine system dysregulation due to abnormalities in synaptic transmission in the ventral subiculum (vSub) within the vSub-nucleus accumbens (NAc) shell circuit. Two distinct populations of subicular projection neurons serve as the major output of the hippocampus, receive input from CA1 and project to cortical and subcortical regions, including to two distinct medium spiny interneuron (MSN) cell- types in the NAc shell. A fundamental understanding how Nrx3 shapes cell-type specific synaptic properties at two subicular synapses involved in DA dysregulation - CA1 input to the vSub and subicular output to the NAc shell - is unexplored; thus, the dissection of cell-type specific pre- and post-synaptic functions of subicular neurons within this disease circuit may open new avenues for treatment strategies. To this end, we generated a Nrx3 mouse where splice site 4 (SS4), the splice site that dictates binding to nearly all neurexin ligands, is constitutively included (SS4+) but can be conditionally excluded (SS4-). We observed a selective reduction in AMPAR synaptic transmission, due to reduced surface AMPAR stability, in the dorsal subiculum of Nrx3SS4+ mice. Altered transsynaptic interactions in the Nrx3SS4 mouse may imitate disease-related Nrx3 mutations because most mutations affect surface exposed residues and alternative splicing - rarely resulting in complete protein loss. The overarching goal of this grant is to dissect presynaptic Nrx3SS4-dependent functions in distinct cell-types at the CA1-vSub synapse and the vSub-NAc shell synapse. The mentored aims will 1.) assess cell-type specific synaptic responses in vSub neurons that project to the NAc shell at the CA1-vSub synapse using stereotactic co-injection of retrograde virus and cre-recombinase in WT and Nrx3SS4 animals and 2.) assay vSub projections to two distinct populations of NAc shell MSNs in the striatal circuit by utilizing stereotactic injection of channelrhodopsin to selectively recruit ventral subicular fibers. For the R00 phase of this grant, will 1.) expand the study of synapse-specific striatal circuitry by dissecting the dorsal Sub-NAc core circuit using techniques acquired during the K99 phase and 2.) identify novel SCAMs by next-generation single-cell RNA sequencing of subicular neurons with the goal long-term of assessing their synaptic function in disease-relevant circuits. I anticipate that this proposal wil uncover new insights into cell-type specific synaptic properties in the subicular-striatal circuit and create a platform for future independent investigations into how other SCAMs function in subicular and striatal circuitry.

描述（由申请人提供）：越来越多的证据表明，突触传递异常可能是某些神经精神疾病的关键机制。如何指定和维护突触属性？突触细胞粘附分子（SCAM）是在调节突触功能中发挥关键作用的主要候选分子。毫不奇怪，基因组研究发现 neurexin-3 (Nrx3) 发生突变，这是一种典型的突触前 SCAM，是 neurexin/neuroligin 复合物的一部分。与精神分裂症（SZ）和寻求奖励行为有关。这些疾病与巨大的社会和经济负担相关，并且具有共同的病理生理学基础——由于伏隔核 (NAc) 壳回路内的腹侧下托 (vSub) 突触传递异常导致多巴胺系统失调。两个不同的皮下投射神经元群作为海马的主要输出，接收来自 CA1 的输入并投射到皮质和皮质下区域，包括 NAc 壳中两种不同的中型多棘中间神经元 (MSN) 细胞类型。 Nrx3 如何在参与 DA 失调的两个下突触（CA1 输入到 vSub 和下突输出到 NAc 壳）上塑造细胞类型特异性突触特性的基本了解尚未被探索；因此，对该疾病回路内的下神经元的细胞类型特异性突触前和突触后功能的剖析可能会为治疗策略开辟新的途径。为此，我们生成了 Nrx3 小鼠，其中剪接位点 4 (SS4)（该剪接位点决定与几乎所有神经毒素配体的结合）被组成型包含 (SS4+)，但可以有条件地排除 (SS4-)。我们观察到，在 Nrx3SS4+ 小鼠的背下托中，由于表面 AMPAR 稳定性降低，AMPAR 突触传递选择性减少。 Nrx3SS4 小鼠中突触相互作用的改变可能会模仿疾病相关的 Nrx3 突变，因为大多数突变影响表面暴露的残基和选择性剪接 - 很少导致蛋白质完全丢失。该资助的总体目标是剖析 CA1-vSub 突触和 vSub-NAc 壳突触不同细胞类型中突触前 Nrx3SS4 依赖性功能。指导目标将 1.) 在 WT 和 Nrx3SS4 动物中使用逆行病毒和 cre 重组酶的立体定向共注射来评估投射到 CA1-vSub 突触处的 NAc 外壳的 vSub 神经元中的细胞类型特异性突触反应，以及 2.)通过利用视紫红质立体定向注射选择性招募腹侧下皮层，对纹状体回路中两个不同的 NAc 壳 MSN 群进行 vSub 投影纤维。对于这笔资助的 R00 阶段，将 1.) 通过使用 K99 阶段获得的技术解剖背侧 Sub-NAc 核心电路来扩展突触特异性纹状体电路的研究，2.) 通过下一代单-NAc 识别新颖的 SCAM对皮下神经元进行细胞 RNA 测序，长期目标是评估其在疾病相关回路中的突触功能。我预计该提案将揭示对皮下-纹状体回路中细胞类型特异性突触特性的新见解，并为未来独立研究其他 SCAM 在皮下和纹状体回路中的功能提供一个平台。