Defining the Connectivity and Dynamics of Peripheral Sour Taste Circuits

定义外围酸味电路的连接性和动态

• 批准号: 10658843
• 负责人: Shannon Marie Landon
• 金额: $ 3.8万
• 依托单位: UNIVERSITY OF TEXAS SAN ANTONIO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-28 至 2025-01-27
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10658843
• 关键词: Biological Assay; Brain; Calcium; Cell Differentiation process; Cells; Circumvallate Papilla; Complex; Consummatory Behavior; Data; Development; Engineering; Fiber; Food; Functional disorder; Fungiform Papilla; Ganglia; Goals; Grant; Half-Life; Hand; Health; Homeostasis; Hour; Image; Individual; Label; Larynx; Light; Location; Longevity; Maintenance; Malnutrition; Maps; Methods; Microscopy; Modeling; Molecular; Mus; Neurons; Nutrient; Peripheral; Population; Process; Proteins; Quality of life; Receptor Cell; SHH gene; Sampling; Signal Transduction; Site; Stimulus; Structure of geniculate ganglion; Synapses; Synaptic Cleft; Synaptic Membranes; System; Taste Buds; Taste Disorders; Taste Perception; Techniques; Technology; Testing; Time; Tissues; Toxin; Transgenic Mice; Variant; Viral; Visualization; Work; cohort; density; design; experimental study; in vivo; inhibitor; innovation; nerve supply; neurogenetics; novel; postsynaptic; precursor cell; presynaptic; prevent; promoter; reconstitution; sensory system; taste system; tool; two photon microscopy

Abstract The sense of taste is a complex and integrative chemosensory system that guides consummatory behavior. Taste receptor cells (TRCs) in the taste bud are continuously renewed and must reconnect with peripheral gustatory neurons (GNs) to relay taste signals to the brain. The turnover and re-establishment of peripheral taste synapses is vital to maintain this complex sensory system. Studies have demonstrated the molecular mechanisms governing the differentiation, maturation and turnover of TRCs as well as the identification of molecularly and functionally separate populations of geniculate ganglion neurons. However, the synaptic connectivity and dynamics between TRCs and GNs remains unknown due to the lack of technologies to examine them. To overcome this, I have developed GFP Reconstitution Across Synaptic Partners (GRASP) to visualize synaptic contacts in the taste bud. GRASP consists of pre- and post-GRASP split GFP molecules expressed, directed, and anchored into the synaptic membranes of pre- and post-synaptic cells, respectively. Because expression of the GRASP components can be controlled genetically, we can use it to assess the connectivity of specific populations. Moreover, GRASP produces bright fluorescent signals in native tissues which grants the unique ability to study synapse dynamics in time-lapse studies. In preliminary work, I have created transgenic mice that express the cyan variant of the pre-GRASP protein under the PKD2L1 promoter, a marker for sour TRCs. Additionally, I have engineered Flex-switch post-GRASP AAVs which enables us to express the post-synaptic GRASP in GNs. With this novel technique at hand, my goal is to genetically identify potential GN-TRC synaptic partners and to characterize synaptic dynamics during taste cell turnover, using sour TRCs as a model. To achieve these goals, this proposal will investigate two aims, 1) Identify synaptic partners for sour TRCs in the fungiform, circumvallate, and laryngeal taste buds, and 2) Determine the longevity and dynamics of sour TRC synapses over time. This proposal is designed to use GRASP, a novel and innovative technique, to elucidate synaptic connectivity and dynamics in the peripheral taste system, which will have far-reaching impact in the fields of taste development, connectivity, and taste disorders.

抽象的 味觉是一个复杂且综合的化学感觉系统，指导完善的行为。品尝 味蕾中的受体细胞（TRC）不断更新，必须与周围味觉重新连接 神经元（GN）将味觉信号传递给大脑。外周味觉突触的更新和重建 对于维持这个复杂的感觉系统至关重要。研究证实了分子机制 控制 TRC 的分化、成熟和周转以及分子和 功能上独立的膝状神经节神经元群。然而，突触连接和 由于缺乏检查技术，TRC 和 GN 之间的动态仍然未知。到 为了克服这个问题，我开发了 GFP Reconstitution Across Synaptic Partners (GRASP) 来可视化突触 味蕾的接触。 GRASP 由 GRASP 前和后分裂的 GFP 分子组成，表达、定向和 分别固定在突触前和突触后细胞的突触膜上。因为表达的 GRASP成分可以通过基因控制，我们可以用它来评估特定人群的连通性。 此外，GRASP 在天然组织中产生明亮的荧光信号，这赋予了独特的研究能力 延时研究中的突触动力学。在前期工作中，我创造了表达 PKD2L1 启动子下 pre-GRASP 蛋白的青色变体，是酸 TRC 的标记。另外，我有 设计的 Flex-switch post-GRASP AAV 使我们能够在 GN 中表达突触后 GRASP。和 这项新技术，我的目标是从基因上识别潜在的 GN-TRC 突触伙伴，并 使用酸味 TRC 作为模型来表征味觉细胞更新过程中的突触动力学。为了实现这些目标， 该提案将研究两个目标，1）识别菌状中酸 TRC 的突触伙伴， 周围味蕾和喉部味蕾，以及 2) 确定酸味 TRC 突触的寿命和动态 时间。该提案旨在使用 GRASP 这种新颖的创新技术来阐明突触 外围味觉系统的连通性和动态性，这将对以下领域产生深远影响： 味觉发育、连通性和味觉障碍。