VISUAL SYSTEM DEVELOPMENT AND SYNAPTOGENESIS

本项目的总体目标是研究控制 形成感光突触的数量和质量组成 在果蝇视叶的第一个神经桩或椎板中 果蝇和苍蝇 这个神经堆的模块，叫做“盒子”， 包括少量的、固定数量的已识别神经元， 两种方法：1）我们将使用定量单节EM来估计 突触频率，测量突触接触部位和细胞表面 地区2)我们还将使用串行EM来追踪突触连接 并进行细胞的计算机三维重建 形态学 基于我们之前对正常突触发生的分析， 我们特别想了解对发育的控制 多接触突触（二联体、三联体等）的组装，为此 果蝇感受器四分体突触是一个模型。 跨物种移植 旨在允许嵌合突触发生的实验将测试 同源神经元之间突触发生信号的保守性 不同的物种 三重突变体rol/sol/mnb将导致 四分体的突触后贡献者的不同组合， 通过突触发生前的自发细胞变性，并允许 存活的细胞有机会取代那些失去的细胞， 表现出他们的突触发生偏好。 我们将照射苍蝇， 包皮环切术杀死包裹板层的上皮神经胶质细胞 墨盒，并探讨突触的后果，和速度， 预期从完整的中间神经元发芽， 邻近的药筒已经被视网膜神经电刺激急性去传入 光烧蚀 我们将扩大我们最初的做法， 通过光感受器的共培养体外突触发生， 视叶细胞 我们将继续进行大型电脑三维重建 一个完整的椎板钉仓EM系列，以量化空间 分布在神经元上的突触群的特征 表面，特别是那些构成相互连接的表面 神经元之间。 我们将研究椎板的动态变化 盒：1）在昼夜节律期间在突触群体之间，以及 冷胁迫后，两者都引起短期变化，和2）之间 interneurons，显示每天的大小变化;以及调查这些 从长期ERG记录功能。 最后，我们将采用 增强子捕获方法：1）研究鉴定的视神经细胞中的基因表达 叶中间神经元，尤其是在已知的条件下发生的， 促进突触发生，以及2）产生特定的分子 这些细胞的标记。 这里提出的研究，因为他们的目的是 产生适用于多接触的突触发生的基本模型 突触，如在视觉系统中广泛发现的二分体和三分体， 应该有助于对扰动的一般理解， 疾病状态，视觉突触在他们的 增长和发展。

The general aims of this project are to study the factors controlling the numerical and qualitative composition of photoreceptor synapses that form in the first neuropile, or lamina, of the optic lobe of the flies Drosophila and Musca. The modules of this neuropile, called cartridges, comprise small, fixed numbers of identified neurons, which we will sample in two ways: 1) We will use quantitative single-section EM to estimate synaptic frequencies, measure synaptic contact sites and cell surface areas. 2) We will also use serial EM to trace out synaptic connections between elements and undertake computer 3-D reconstructions of cell morphologies. Based on our previous analysis of normal synaptogenesis, we especially want to understand the control of the developmental assembly of multiple-contact synapses (dyads, triads, etc.), for which the fly receptor tetrad synapses are a model. Cross-species transplant experiments designed to allow chimeric synaptogenesis will test the conservation of synaptogenetic signals between homologous neurons in different species. The triple mutant rol/sol/mnb will procure the loss of differing combinations of postsynaptic contributors to the tetrads, through spontaneous cell degeneration prior to synaptogenesis, and allow surviving cells the opportunity to replace those lost, and thereby exhibit their synaptogenetic preferences. We will irradiate flies as prepupae to kill the epithelial glial cells that envelop the lamina cartridges, and explore the synaptic consequences, and rapidity, of anticipated sprouting from the interneurons of intact cartridges to adjacent cartridges which have been acutely deafferented by retinal photo-ablation. We will extend our initial approaches to procure synaptogenesis in vitro through the co-culture of photoreceptors and optic lobe cells. We will continue a major computer 3-D reconstructions of a complete lamina cartridge EM series to quantify the spatial characteristics of synaptic populations distributed over neuronal surfaces, especially those which constitute reciprocal connections between neurons. We will examine dynamic changes in the lamina cartridge: 1) amongst synaptic populations during a circadian rhythm, and after cold stress, both of which cause short-term changes, and 2) amongst interneurons, which show daily size changes; as well as investigate these functionally from long-term ERG recordings. Lastly, we will employ enhancer trap methods: 1) to study gene expression in identified optic lobe interneurons, especially that occurring under conditions known to promote synaptogenesis, as well as 2) to generate specific molecular markers to these cells. The studies proposed here, because they aim to produce a basic model of synaptogenesis applicable to multiple-contact synapses, such as the dyads and triads found widely in visual systems, should contribute to a general understanding of the perturbations in disease states to which visual synapses are susceptible during their growth and development.