New Genetic Tracers for Concurrent Labeling of Multiple Gustatory Neural Pathways

用于同时标记多个味觉神经通路的新基因示踪剂

基本信息

批准号：
8109237
负责人：
ICHIRO MATSUMOTO
金额：
$ 14.33万
依托单位：
MONELL CHEMICAL SENSES CENTER
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-15 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8109237
关键词：
Affect Animals Antibodies Area Binding Cells Chemicals Code Computer Simulation Cultured Cells Engineering Enhancers Evaluation FLAG peptide Future Genes Genetic Grant In Vitro Label Lectin Mediating Methods Modality Molecular Molecular Conformation Mus Nervous system structure Neural Pathways Neuraxis Neurosciences Research Oligosaccharides Pathway interactions Peptides Phospholipase C Plant Lectins Plasmids Protein Engineering Proteins Receptor Cell Research Research Personnel Screening procedure Signal Transduction Structure System Taste Perception Testing Tracer Transgenic Animals Transgenic Mice Virus Wheat Germ Agglutinins Work barley lectin base comparative efficacy enhanced green fluorescent protein immunocytochemistry in vitro activity in vivo mRNA Expression molecular dynamics neural circuit promoter protein distribution public health relevance relating to nervous system research study somatosensory tool

项目摘要

DESCRIPTION (provided by applicant): Although in recent decades our understanding of the molecular mechanisms of taste reception has advanced, how the taste information is transmitted and coded in the nervous system remains unclear. Our long-term objective is to understand the coding system of gustatory information, from taste receptor cells to the central nervous system. In this proposal, we aim to develop genetic tools that will greatly contribute to our understanding by concurrently elucidating precise gustatory neural pathways within the same animal, and then use these tools to test our hypothesis that gustatory information for different taste modalities is transmitted via specific ascending neural pathways. One well-known genetic tracer is wheat germ agglutinin (WGA), which was developed ten years ago. In the research we propose here, we will engineer various WGAs that integrate specific tags such as Myc peptide, FLAG peptide, and enhanced green fluorescent protein, and will evaluate their tertiary structure by molecular dynamic simulation. We will evaluate the resultant engineered WGAs in terms of their intracellular stability and lectin activity in vitro. We will then induce their expression in mice in a subset of taste receptor cells under the control of promoters/enhancers of taste reception-related genes and evaluate their utility in vivo. From these studies, we expect to develop a minimum of two tagged WGA- based genetic tracers that are available to label multiple gustatory pathways concurrently and distinguishably. These tracers should be of great use not only in our future gustatory research but in studies of olfactory and somatosensory systems and many other areas of neuroscience research that study the genetics of interconnected pathways. PUBLIC HEALTH RELEVANCE: This proposed research aims to develop new genetic tools to label multiple neural circuitries concurrently and distinguishably. These tools will contribute to elucidating relationships between multiple neural pathways for different taste modalities. They will also contribute to other neuroscience research, including studies of olfactory and somatosensory systems.

描述（由申请人提供）：尽管近几十年来我们对味觉接收的分子机制的理解有所进步，但味觉信息如何在神经系统中传递和编码仍不清楚。我们的长期目标是了解从味觉感受器细胞到中枢神经系统的味觉信息编码系统。在本提案中，我们的目标是开发遗传工具，通过同时阐明同一动物内精确的味觉神经通路，极大地促进我们的理解，然后使用这些工具来检验我们的假设，即不同味觉模式的味觉信息是通过特定的上升传递的神经通路。一种著名的遗传示踪剂是十年前开发的麦芽凝集素（WGA）。在我们提出的研究中，我们将设计集成Myc肽、FLAG肽和增强型绿色荧光蛋白等特定标签的各种WGA，并通过分子动力学模拟评估它们的三级结构。我们将评估由此产生的工程 WGA 的细胞内稳定性和体外凝集素活性。然后，我们将在味觉接收相关基因的启动子/增强子的控制下，在小鼠的味觉受体细胞子集中诱导它们的表达，并评估它们在体内的效用。从这些研究中，我们期望开发至少两种基于 WGA 的标记遗传示踪剂，可同时且可区分地标记多个味觉通路。这些示踪剂不仅在我们未来的味觉研究中，而且在嗅觉和体感系统的研究以及研究互连通路的遗传学的神经科学研究的许多其他领域都有很大的用处。公共健康相关性：这项拟议的研究旨在开发新的遗传工具来同时和区分地标记多个神经回路。这些工具将有助于阐明不同味觉模式的多个神经通路之间的关系。他们还将为其他神经科学研究做出贡献，包括嗅觉和体感系统的研究。