VISUAL SYSTEM DEVELOPMENT AND SYNAPTOGENESIS

本项目的总体目标是研究控制经济增长的因素 脑内形成的光感受器突触的数量和质量组成 果蝇视叶的第一神经丛或板层 还有马斯卡。我们特别想了解对 多接触突触的发育组装(二联体、三联体等)， 苍蝇受体四分体突触就是一个模型。的模块 这种薄片被称为墨盒，由少量固定数量的已识别的 神经元。我们将对这些进行抽样：1)使用定量的单截面EM来 估计突触频率，测量突触接触部位和细胞 表面积。2)用串口电磁法进行计算机三维 未成熟细胞形态重建与树突状细胞生长的关系 形成突触。在发生快速变化的状态下 光或温度的函数反转，我们将绘制位置 新的突触位置相对于旧的突触位置，以检查一个 在现有地点附近形成一个新的突触地点，并 突触前带在新的位置形成的模式。我们还将 观察突触细胞器之间膜的循环 果蝇突变体Shibire。解剖性突触的功能结局 将从ERG记录中寻求改变，以及活动依赖于 突触形成将被评估具有受损的Pre-or突变 突触后板层功能。生物体内昼夜节律的调控 由两组广泛分布的神经元组成的板层，其中一个被认为释放5-羟色胺， 另一种多肽PDF，将作为可塑性的进一步例子进行分析 在椎板上。我们将照射苍蝇作为蛹来杀死上皮细胞 包裹椎板盒的神经胶质细胞，探索突触 预期的中间神经元萌发的后果和迅速 将完整的墨盒转移到相邻的墨盒，这些墨盒已经严重损坏 通过视网膜光消融术去感觉神经。三重突变体rol/sol/mnb 会导致突触后不同组合的丢失 四分体的贡献者，通过先前的自发细胞退化 突触发生，并让存活的细胞有机会展示 它们替代突触伙伴的能力。我们将利用 我们已经开发了组织培养系统来检查选择性丛生组织 光感受器轴突及其与板层细胞的相互作用 靶点，并通过共培养获得体外突触发生。 光感受器和视叶细胞。在与其他实验室的合作中，我们 将检查参与形成或形成的基因的表达 突触的维持。我们将研究GAL4果蝇系，在这些系中 基因表达是用外源报告基因来标记的，以识别 在已知促进突触发生的条件下发生的表达，如 以及将特定的分子标记固定到所识别的视叶 细胞。我们还将研究两个可能的突触表达 有希望的候选基因，unC和demo，使用免疫-EM。这些研究 之所以在这里提出，是因为他们的目标是产生一个基本的突触发生模型 适用于多触点突触，如二联体和三联体 广泛存在于视觉系统中，将有助于对 视觉突触在疾病状态下的扰动 在它们的生长和发育过程中容易受到影响。

The general aims of this project are to study the factors controlling the numerical and qualitative composition of photoreceptor synapses formed in the first neuropile, or lamina, of the optic lobe of the flies Drosophila and Musca. 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. The modules of the lamina, called cartridges, comprise small, fixed numbers of identified neurons. We will sample these: 1) using quantitative single-section EM to estimate synaptic frequencies, measure synaptic contact sites and cell surface areas. 2) using serial EM to undertake computer 3-D reconstructions of immature cell morphologies, to relate dendritic growth to synapse formation. Under states of rapid change occurring after functional reversals of light or of temperature, we will plot the location of new synaptic sites relative to old ones, to examine the influence of an existing site on the formation of a new synaptic site nearby, and the presynaptic ribbon's mode of formation at the new site. We will also examine the recycling of membrane between synaptic organelles, using the Drosophila mutant shibire. The functional outcome of anatomical synaptic changes will be sought from ERG recordings, and the activity-dependence of synapse formation will be assessed in mutants with impaired pre- or postsynaptic lamina function. The control of circadian modulations in the lamina by two sets of widespread neurons, one thought to release 5-HT, the other the peptide PDF, will be analyzed as a further example of plasticity in the lamina. We will irradiate flies as pupae to kill the epithelial glial cells that envelop the lamina cartridges, and explore the synaptic consequences, and rapidly, of anticipated sprouting from the interneurons of intact cartridges to adjacent cartridges which have been acutely deafferented by retinal photo-ablation. 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 exhibit their ability to substitute synaptic partners. We will exploit the tissue-culture system we have developed to examine selective fasciculation amongst photoreceptor axons and their interactions with their lamina cell targets, and to procure synaptogenesis in vitro from co-cultures of photoreceptors and optic lobe cells. In collaboration with other labs, we will examine the expression of genes involved in either the formation or maintenance of synapses. We will examine GAL4 Drosophila lines, in which gene expression is marked by an exogenous reporter gene, to identify expression occurring under conditions known to promote synaptogenesis, as well as to secure specific molecular markers to identified optic lobe cells. We will also examine the possible synaptic expression of two promising candidate genes, irreC and demo, using immuno-EM. 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, will contribute to a general knowledge of the perturbations in disease states to which visual synapses are susceptible during their growth and development.