MOLECULAR GENETIC ANALYSIS OF DEVELOPING SYNAPSES

描述（由申请人提供）：神经细胞相互交流的主要方式是通过化学突触释放神经递质。大脑处理信息的能力依赖于在许多不同类型的神经元之间精确而可靠地形成突触连接。这一建议旨在发展对调节突触发生的突触伙伴之间信号传导的分子理解。众所周知，即使在简单的后生动物中，如秀丽隐杆线虫，突触活动的变化也会引起突触强度和结构的代偿性变化。我们建议结合遗传学，细胞生物学，分子生物学和实时成像来识别和表征协调神经突触突触发育的信号通路的分子成分的作用。首先，我们的目标是描述新生突触形成的细胞事件的顺序，通过可视化荧光标记的成分招募到新形成的突触。我们将定义线粒体、突触囊泡、活性区成分和粘附分子出现在突触位点的顺序。我们还将定义介导突触前特化的后续生长的细胞机制。其次，我们将定义被鉴定为不能形成突触的突变体的新分子成分的作用。利用各种分子、遗传和蛋白质相互作用的研究，我们将在突触组装的当前分子模型中定位基因。第三，我们将使用一种新的突触标签，在荧光解剖范围下可以很容易地在活体动物中检测到，利用遗传学方法分离和表征秀丽隐杆线虫机械感觉神经元与其突触伙伴之间信号传导的基因。总之，这些方法将有助于确定细胞在突触形成和突触维持过程中用于识别和相互交流的机制。虽然秀丽隐杆线虫的突触发生无疑没有脊椎动物那么复杂，但已经很清楚的是，在这两个系统中，类似的途径起作用。因此，对秀丽隐杆线虫中参与这一过程的分子的分析应该有助于定义一套一般的和可能保守的原则，这些原则在一般的突触发生机制中是共同的。公共卫生相关性：突触连接是大脑中主要的神经元通信结构。在阿尔茨海默病中，现在已经确定突触密度的变化（即突触连接的丧失）与认知障碍的关系比与该疾病相关的标志性斑块和缠结病变更好。我们的工作重点是了解突触连接是如何形成的。这种对大脑发育和功能的基本科学理解将有助于开发在疾病早期进行干预的疗法，从而减缓或阻止突触丧失。

DESCRIPTION (provided by applicant): The primary means by which nerve cells communicate with each other is through the release of neurotransmitter at chemical synapses. The ability of the brain to process information depends on synaptic connections forming precisely and reliably between many different types of neurons. This proposal is directed towards developing a molecular understanding of the signaling between synaptic partners that regulate synaptogenesis. It is well established that even in simple metazoans like the worm C. elegans changes in synaptic activity induce compensatory changes in synaptic strength and structure. We propose to use a combination of genetics, cell biology, molecular biology and live imaging to identify and characterize the role of molecular components of the signaling pathways that coordinate synaptic development at nerve-nerve synapses. First, we aim to describe the order of cellular events in nascent synapse formation by visualizing the recruitment of fluorescent-tagged components to newly forming synapses. We will define the order in which mitochondria, synaptic vesicles, active zone components and adhesion molecules appear at synaptic sites. We will also define the cellular mechanisms that mediate subsequent growth of the presynaptic specializations. Second, we will define the role of novel molecular components that were identified as mutants that fail to form synapses. Using a variety of molecular, genetic and protein interaction studies we will position the genes within the current molecular models of synapse assembly. Third, we will use genetic approaches to isolate and characterize genes which disrupt signaling between mechanosensory neurons and their synaptic partners in C. elegans using a novel synaptic tag which can be easily detected in live animals under a fluorescent dissecting scope. Together these approaches will help define mechanisms that cells use to identify and communicate with one another during the process of synapse formation and synaptic maintenance. While synaptogenesis is undoubtedly less complex in C. elegans than in vertebrates, it is already clear that similar pathways operate in both systems. Thus, analysis of the molecules participating in the process in C. elegans should help define a set of general and likely conserved principles that are common to synaptogenesis mechanisms in general. PUBLIC HEALTH RELEVANCE: Synaptic connections are the primary neuronal communication structures in the brain. In Alzheimer's disease, it is now well established that changes in synaptic density (i.e. loss of synaptic connections) correlate better with cognitive impairment that the hallmark plaque and tangle lesions that are also associated with the disease. Our work is focused on understanding how synaptic connections are formed. Such basic scientific understanding of brain development and function will aid in developing therapies that intervene early in disease hence slowing or arresting synaptic loss.