Collaborative Research: Specification of Excitatory Fates in the Spinal Cord

这个项目研究了胚胎发育和成长过程中，神经元回路是如何发挥适当功能的。这项工作的重点是斑马鱼胚胎脊髓中的神经回路，这种回路在生命早期发育，比大脑中的神经回路更简单，更容易实验处理。目标是识别关键的调控基因，这些基因指示细胞成长为具有特定功能的神经细胞(中间神经元)，这些神经细胞具有控制精确身体运动的特定功能。该项目的成果将被广泛应用，因为这些类型的神经细胞存在于所有脊椎动物(包括人类)中。在这项研究中发现的基因以及它们影响神经细胞和神经回路发育的方式在许多不同的物种中可能是相似的。该项目将显著增加对基因如何指导细胞以特定特征发育的理解，以及特定基因在构建神经元电路中所起的作用。主要研究人员将把这项研究纳入他们的大学教学和向公众发表的报告中。这项研究将通过妇女和代表性不足的少数群体的积极参与，促进科学工作者的多样化。主要调查人员还将对当地城市高中进行外展，并为9/10年级和高级选修高中的生物学生开发教学模块。这些活动的目的是显著影响传统上服务不足的高中生的生活，并激发代表少数族裔的高中生对科学和研究的热情。理解神经元网络是如何产生的一个重要的第一步是确定不同的中间神经元(INS)是如何被指定为具有特定功能属性的。帮助定义INS的一个关键特性是它们使用哪种神经递质与其他细胞通信。神经递质可以是兴奋性的，也可以是抑制性的，而INS利用不适当的神经递质形成故障电路。因此，确定IN神经递质的特性对于理解IN和电路的发展都是至关重要的。关于兴奋性脊髓神经营养因子的特性是如何被指定的，我们知之甚少。这项建议将以斑马鱼为模型系统，描述一个基因调控网络(GRN)，该网络指定脊髓IN的兴奋特性。研究人员已经确定编码11种不同DNA结合蛋白的基因是GRN的候选成员，GRN指定V0v IN兴奋性(谷氨酸能)特性。V0v INS位于脊髓中部，在运动回路中起着至关重要的作用。研究小组已经证明，其中四个基因evx1/2和lmx1ba/b是正确的V0v INS兴奋命运所必需的。此外，在evx1/2双突变体中，V0v INS失去了lmx1ba/b和其他7个基因的表达，这表明所有这些基因都可能在V0v谷氨酸能特性的Evx1/2下游起作用。这项建议的具体目标是测试这7个其他基因中是否有任何其他基因是正确的V0v神经递质特性所必需的，并确定所有这些受Evx1/2调控的基因之间的上位关系。单一和复合斑马鱼突变体将被用来回答这些问题。这个奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

This project examines how appropriately-functioning neuronal circuits are made as embryos develop and grow. The work focuses on circuits in the spinal cord of zebrafish embryos, which develop early in life and are simpler and more experimentally-tractable than neural circuits in the brain. The goal is to identify key regulatory genes that instruct cells to grow into one particular population of nerve cells (interneurons) with specific functions in controlling precise body movements. The outcomes of this project will be widely applicable, because these types of nerve cell exist in all vertebrates (including humans). The genes identified in this study and the ways that they influence nerve cell and circuit development are likely to be similar in many different species. This project will significantly increase understanding of how genes instruct cells to develop with particular characteristics, and the roles that specific genes play in the construction of neuronal circuitry. The principal investigators will incorporate this research into their university teaching and presentations made to the general public. This research will contribute to the diversification of the scientific workforce through the active participation of both women and under-represented minorities. The principal investigators will also perform outreach to local city high schools and develop teaching modules for both 9th/10th grade and advanced placement high school Biology students. The aim of these activities is to significantly impact the lives of traditionally under-served high school students and enthuse under represented minority high school students about science and research.An essential first step in understanding how neuronal networks are generated is determining how distinct interneurons (INs) are specified with particular functional properties. One key property that helps to define INs is which neurotransmitter they use to communicate with other cells. Neurotransmitters can be excitatory or inhibitory and INs utilizing inappropriate neurotransmitters form malfunctioning circuits. Therefore, determining how IN neurotransmitter properties are specified is crucial for understanding both IN and circuit development. Relatively little is known about how excitatory spinal IN properties are specified. This proposal will characterize a gene regulatory network (GRN) that specifies spinal IN excitatory properties, using zebrafish as a model system. The investigators have identified genes that encode for 11 different DNA-binding proteins as candidate members of a GRN that specifies V0v IN excitatory (glutamatergic) properties. V0v INs reside in the middle of the spinal cord and have crucial roles in locomotor circuitry. The research team has already shown that four of these genes, evx1/2 and lmx1ba/b are required for correct excitatory fates of V0v INs. In addition, in evx1/2 double mutants V0v INs lose expression of both lmx1ba/b and the other 7 genes, suggesting that all 9 of these genes may act downstream of Evx1/2 in specifying V0v glutamatergic properties. The specific objectives of this proposal are to test if any of these 7 other genes are required for correct V0v neurotransmitter properties and determine the epistatic relationships among all of these Evx1/2-regulated genes. Single and compound zebrafish mutants will be used to answer these questions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.