Synapse growth and elimination in mature CNS

 描述(由申请人提供)：长期目标是了解成人中枢神经系统中的突触结构如何支持学习和记忆。我们研究了海马长时程增强(LTP)，这是一种突触可塑性的形式，被视为学习和记忆的细胞底物。研究的重点是树突棘，这是大脑中90%以上的兴奋性突触所在的微小突起，在LTP、学习和记忆过程中被修改，并在各种神经疾病中严重扭曲。最近，我们已经证明，如果LTP诱导间隔超过1.5小时，最初饱和的LTP可以随后增加，在成年小鼠的海马区，间隔4小时就可以达到100%的成功率。这些发现支持这样一种假设，即LTP的间隔发作参与了可能是间隔学习相对于集中学习的优势的机制。利用连续切片电子显微镜(3DEM)的三维重建，我们发现了突触结构的几个变化，这些变化在LTP最初饱和后随着时间的推移而显现。在体内和体外，许多突触都有新生区域，即动态的边缘区域，具有突触后密度，但缺乏通常在活动区发现的突触前小泡。初生区迅速获得突触前小泡，从而在30分钟后转变为活动区。2小时后，初生区和活动区均扩大，最大的突触扩大发生在含有光滑内质网(SER)和多核糖体的棘上。我们将测试这样的假设，即修改突触结构的能力最初被LTP饱和，突触需要时间来恢复或生长，为以后的增强做准备。在进行更耗时的3DEM和免疫标记之前，我们将在成熟的小鼠海马区进行各种药物和遗传操作后，用生理学方法验证成功的LTP诱导。我们的目标是验证以下关于增强机制的假设：1)受体和突触前对接位置必须在LTP初始饱和后扩大的突触处积累。2)SER依赖的突触生长和脊髓聚集有助于LTP的增强。3)LTP的增强需要依赖蛋白质合成的突触生长。4)缺乏参与构建或稳定突触的候选分子会扰乱LTP的饱和或增强。在完成这些目标后，我们将知道结构突触可塑性的哪些元素是LTP增强所必需的。这些结果将为新生的区域转换、突触生长和脊柱聚集的机制提供新的理解，以及这些过程是否与SER扩张、局部蛋白质合成和突触黏附相耦合，为随后在成熟小鼠大脑中最初饱和的LTP的增强做准备。这一结果最终应该会为修复功能失调的突触电路的新策略的开发提供信息。

 DESCRIPTION (provided by applicant): The long-term goal is to understand how synapse structure in the adult central nervous system supports learning and memory. We study hippocampal long-term potentiation (LTP), a form of synaptic plasticity that is viewed as a cellular substrate for learning and memory. The focus is on dendritic spines, the tiny protrusions that host more than 90% of excitatory synapses in the brain, are modified during LTP, learning, and memory, and are severely distorted in a variety of neurological disorders. Recently, we have shown that initially saturated LTP can be subsequently augmented if more than 1.5 hours elapse between episodes of LTP induction, with 100% success achieved in adult mouse hippocampus after a 4 hour interval. These findings support the hypothesis that spacing episodes of LTP engages mechanisms that might underlie the advantage of spaced over massed learning. Using 3D reconstruction from serial section electron microscopy (3DEM), we have discovered several changes in synapse structure that manifest over time after the initial saturation of LTP. Both in vivo and in vitro, many synapses had nascent zones, dynamic edge regions that have a postsynaptic density but lack the presynaptic vesicles normally found at active zones. Nascent zones rapidly acquired presynaptic vesicles, thereby converting to active zones however by 30 min. By 2 hr., both nascent and active zones were enlarged, and the greatest synapse enlargement occurred on spines that contained smooth endoplasmic reticulum (SER) and polyribosomes. We will test the hypothesis that the capacity to modify synaptic structure is initially saturated by LTP, and time is required for synapses to recover or grow in preparation for later augmentation. We will verify successful LTP induction with physiology following various pharmacological and genetic manipulations in mature mouse hippocampus prior to performing the more time consuming 3DEM and immunolabeling. We aim to test the following hypotheses regarding mechanisms of augmentation: 1) That receptors and presynaptic docking sites must accumulate at synapses enlarged after the initial saturation of LTP. 2) That SER-dependent synapse growth and spine clustering serve the augmentation of LTP. 3) That protein synthesis-dependent growth of synapses is required for augmentation of LTP. 4) That absence of candidate molecules involved in building or stabilizing synapses disrupts saturation or augmentation of LTP. Upon completion of these aims we will know which elements of structural synaptic plasticity are integral to the augmentation of LTP. Outcomes will provide new understanding of the mechanisms of nascent zone conversion, synapse growth, and spine clustering, and whether these processes are coupled to SER expansion, local protein synthesis, and synapse adhesion in preparation for the subsequent augmentation of initially saturated LTP in the mature mouse brain. The results should ultimately inform the development of new strategies to repair dysfunctional synaptic circuits.