Dynamic regulation of SER during development and synaptic plasticity

发育和突触可塑性过程中 SER 的动态调节

• 批准号: 8701399
• 负责人: DEBORAH JEAN WATSON
• 金额: $ 5.7万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8701399
• 关键词: AMPA Receptors; Adult; Age; Basic Science; Calcium; Carrying Capacities; Cell model; Collaborations; Complex; Dendrites; Dendritic Spines; Dependence; Development; Electron Microscopy; Elements; Equilibrium; Goals; Hippocampus (Brain); Hour; Indium; Knowledge; Laboratories; Learning; Light; Long-Term Potentiation; Membrane; Membrane Protein Traffic; Memory; Mission; Modeling; Modification; NMDA receptor antagonist; National Institute of Child Health and Human Development; National Institute of Mental Health; Neurons; Organelles; Outcome; Pattern; Perfusion; Phase; Phosphorylation; Process; Protein Biosynthesis; Proteins; Pyramidal Cells; Regulation; Relative (related person); Research; Research Proposals; Resolution; Resources; Role; Route; Science; Slice; Smooth Endoplasmic Reticulum; Specificity; Structure; Sum; Synapses; Synaptic plasticity; Testing; Time; Tubular formation; United States National Institutes of Health; Vertebral column; brain behavior; cognitive function; density; developmental disease; experience; in vivo; indexing; light microscopy; nanometer; postnatal; public health relevance; reconstruction; research study; synaptogenesis; therapeutic target

DESCRIPTION (provided by applicant): New research from a collaboration between the Harris and Ehlers laboratories shows that the mobility of cargo along the smooth endoplasmic reticulum (SER) of hippocampal dendrites is reduced where the SER is highly elaborated, particularly along spiny segments and at dendritic branch points. The primary objective of this research proposal is to investigate whether the SER network within developing dendrites is locally regulated in the vicinity of spines and synapses. We will examine changes in dendritic SER organization during a period of robust synaptogenesis from postnatal day (P) 15 to P21. During this one postnatal week, oblique dendrites mature substantially in spine number and synapse size. Aim 1 will examine how the dendritic SER accommodates new spine formation during normal development. We will test the hypothesis that the local organization of dendritic SER becomes more elaborate as spine density and synapse size increase from P15 to P21. To test this hypothesis, we will use the nanometer resolution of serial section electron microscopy to make three-dimensional (3D) reconstructions of dendrites, synapses, and the dendritic SER network of perfusion-fixed hippocampus in vivo at P15 and P21. To estimate an index of local elaboration, we will compare regions of SER that form complex SER cisterns to regions that mainly consist of simple tubules along spiny and spine-free segments of dendrites. We will also induce long-term potentiation (LTP) in hippocampal slices (as a cellular model of learning) to investigate changes in the organization of dendritic SER during synaptic plasticity. We will use theta-burst stimulation (TBS) to study multiple phases of LTP. The AMPA receptor-dependent, early-phase LTP (E-LTP) lasts up to an hour, while the protein-synthesis dependent, late- phase LTP (L-LTP) endures for more than 3 hrs. Aim 2 will test whether dendritic SER undergoes plasticity- dependent elaboration during E-LTP. Aim 3 will test the hypothesis that plasticity-dependent elaboration of dendritic SER is reduced during L-LTP relative to E-LTP. In these experiments, slices that received TBS (1 each for E-LTP and L-LTP) will be compared to a slice that only received control-stimulation. After expression of LTP, we will compare spine density and synapse size between control and TBS dendrites at P15 and P21, and evaluate whether the organization of the dendritic SER near spines is altered relative to the spine-free portions of the dendritic shaft in these slices. LTP-specific effects will be controlled for by administering APV, a competitive NMDA receptor antagonist known to block LTP. The implications of these findings will establish a comprehensive description of the role of SER in the development and plasticity of spines and synapses at a postnatal age where developmental disorders often manifest dendritic abnormalities. The goals put forth in this application serve the mission of a variety of NIH agencies that use basic research to understand the evolving science of brain, behavior, and experience (e.g., NIMH and NICHD) and will help to identify potential therapeutic targets for developmental disorders associated with malformed spines and dysfunctional SER.

描述（由申请人提供）：Harris 和 Ehlers 实验室合作的新研究表明，在海马树突的平滑内质网 (SER) 高度精细的地方，货物沿着海马树突的平滑内质网 (SER) 的流动性会降低，特别是沿着多刺节段和树突分支点。本研究计划的主要目的是研究发育中树突内的 SER 网络是否在树突棘和突触附近受到局部调节。我们将检查从出生后第 15 天到第 21 天突触发生期间树突 SER 组织的变化。在出生后的这一周内，斜树突的棘数量和突触大小显着成熟。目标 1 将检查树突 SER 在正常发育过程中如何适应新脊柱的形成。我们将检验这样的假设：随着树突棘密度和突触大小从 P15 增加到 P21，树突 SER 的局部组织变得更加复杂。为了验证这一假设，我们将使用连续切片电子显微镜的纳米分辨率对 P15 和 P21 体内灌注固定海马的树突、突触和树突 SER 网络进行三维 (3D) 重建。为了估计局部精细化指数，我们将比较形成复杂 SER 池的 SER 区域与主要由沿着树突的带刺和无刺部分的简单小管组成的区域。我们还将在海马切片（作为学习的细胞模型）中诱导长时程增强（LTP），以研究突触可塑性期间树突 SER 组织的变化。我们将使用 theta 突发刺激 (TBS) 来研究 LTP 的多个阶段。 AMPA 受体依赖性早期 LTP (E-LTP) 可持续长达 1 小时，而蛋白质合成依赖性晚期 LTP (L-LTP) 可持续 3 小时以上。目标 2 将测试树突 SER 在 E-LTP 期间是否经历可塑性依赖的加工。目标 3 将检验以下假设：相对于 E-LTP，L-LTP 期间树突 SER 的可塑性依赖性加工会减少。在这些实验中，接收 TBS 的切片（E-LTP 和 L-LTP 各 1 个）将与仅接收控制刺激的切片进行比较。 LTP 表达后，我们将比较 P15 和 P21 时对照树突和 TBS 树突之间的树突密度和突触大小，并评估树突附近树突 SER 的组织是否相对于无树突刺部分发生改变。 这些切片中的树突轴。 LTP 特异性效应可通过施用 APV 来控制，APV 是一种已知可阻断 LTP 的竞争性 NMDA 受体拮抗剂。这些发现的意义将全面描述 SER 在出生后脊柱和突触发育和可塑性中的作用，在出生后发育障碍通常表现为树突异常。本申请中提出的目标服务于各种 NIH 机构的使命，这些机构利用基础研究来了解不断发展的大脑、行为和经验科学（例如 NIMH 和 NICHD），并将有助于确定发育障碍的潜在治疗靶点与脊柱畸形和 SER 功能失调有关。