Genetic screens for analysis of Ca-dependent transmitter specification

描述(由申请人提供)： 在以前的工作中，我们证明了由发育瞬时钙依赖动作电位产生的钙尖峰调节神经递质的表达。我们发现，这种调节是动态平衡的，因此，钙峰活动的增加导致表达抑制性神经递质(甘氨酸和GABA)的细胞比例增加，而兴奋性神经递质(谷氨酸和乙酰胆碱)的比例降低。相反，钙峰活动的减少导致兴奋性神经递质的发生率增加和抑制性神经递质的减少。这项拟议的研究有两个特定的目标，目的是在热带非洲爪哇中建立突变和转基因株系，有助于识别参与钙峰依赖的神经递质规范的分子，并一般用于研究活动依赖的发育。第一个特定目的使用功能增益筛选，第二个特定目的使用基因陷阱插入突变。第三个特定目标描述了第一个和第二个目标中确定的基因的一个子集。这项研究的直接目标是确定与钙峰依赖的神经递质相关的分子，并为这一领域的进一步研究提供试剂。此外，我们的目标是产生对非洲爪哇群落有价值的信息和突变系。长期目标是提供有关控制神经元发育过程的分子信号通路的信息，这将有助于理解神经系统的发育障碍。公共卫生相关性：胚胎发育过程中神经递质表达的正确规范对神经系统的功能至关重要，基因表达和电活动都有助于这一过程。然而，先天(基因)和后天(活动)相互作用以确定神经递质的适当表达的机制尚不清楚。这项研究将确定调节递质表达的活性依赖基因。预计这项工作将有助于理解神经系统的发育障碍，在胚胎和胚胎后发育过程中，神经递质的表达会发生变化。

DESCRIPTION (provided by applicant): In previous work we demonstrated that Ca spikes generated by developmentally transient Ca-dependent action potentials regulate expression of neurotransmitters. We showed that this regulation was homeostatic, such that an increase in the Ca spike activity resulted in an increase in the incidence of cells expressing inhibitory neurotransmitters (glycine and GABA) and a concomitant decrease in the incidence of excitatory neurotransmitters (glutamate and acetylcholine). Conversely, a decrease in Ca spike activity caused an increase in the incidence of excitatory neurotransmitters and a decrease in inhibitory neurotransmitters. The proposed research has two specific aims targeted at generating mutant and transgenic lines in Xenopus tropicalis useful for identifying molecules involved in the Ca spike-dependent specification of neurotransmitters and for studying activity-dependent development in general. The first specific aim uses a gain-of-function screen and the second specific aim uses gene trap insertional mutagenesis. The third specific aim characterizes a subset of the genes identified in the first and second aims. The immediate goal of this research is to identify molecules involved in Ca spike- dependent neurotransmitter specification and to generate reagents for further research in this area. In addition, we aim to generate information and mutant lines that are of value to the Xenopus community. The long-term goal is to provide information about the molecular signaling pathways that govern processes of neuronal development, which will contribute to understanding developmental disorders of the nervous system. PUBLIC HEALTH RELEVANCE: Correct specification of neurotransmitter expression during embryonic development is essential for the function of the nervous system, and both gene expression and electrical activity contribute to this process. However the mechanisms by which nature (genes) and nurture (activity) interact to specify the appropriate expression of neurotransmitters are unclear. This research will identify activity-dependent genes regulating transmitter expression. It is expected that this work will contribute to understanding developmental disorders of the nervous system in which neurotransmitter expression is altered during embryonic and post-embryonic development.