Fluorescent Indicators for Imaging Synaptic Zinc in Cortical Sound Processing

用于皮质声音处理中突触锌成像的荧光指示剂

基本信息

批准号：
10599603
负责人：
Huiwang Ai
金额：
$ 68.42万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-30 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10599603
关键词：
Address Affinity Alzheimer&apos s Disease Animals Auditory area Brain Cells Chelating Agents Color Cytosol Development Directed Molecular Evolution Engineering Epilepsy Frequencies Generations Genes Hearing problem Image Imaging Device Imaging technology In Vitro Ischemia Knockout Mice Link Location Membrane Mental Depression Mus Neurobiology Neuromodulator Neurons Neurotransmitters Oranges Organelles Outcome Property Proteins Research Personnel Role SLC30A3 gene Sensory Shapes Signal Transduction Specificity Stimulus Stroke Synapses Testing Time Transgenic Mice Variant Vesicle Viral Vector Zinc auditory processing awake base cell type extracellular fluorescence imaging imaging capabilities improved in vivo information processing innovation knockout gene nervous system disorder neurotransmission new technology novel presynaptic protein transport public health relevance response sensory cortex sensory stimulus sound sound frequency spatiotemporal synergism trafficking two-photon

项目摘要

Abstract Although the importance of synaptic Zn2+, as an emerging neuromodulator throughout the brain, has been widely appreciated, the dynamics of synaptic Zn2+ release in response to naturally occurring stimuli remains largely elusive. Genetically encoded Zn2+ indicators (GEZIs) derived from fluorescent proteins are popular tools for imaging Zn2+ in the cytosol and intracellular organelles. However, fluorescence imaging of Zn2+ secretion in the brain in live animals has not yet been achieved due to the limitations of current GEZIs (e.g., insufficient extracellular membrane localization, mismatching affinity, and/or inadequate dynamic range and photostability). This interdisciplinary multi-PI 4-year R01 project, led by Dr. Huiwang Ai with expertise in genetically encoded indicators and fluorescence imaging and Dr. Thanos Tzounopoulos with expertise in studying the role of Zn2+ in auditory processing, aims to (1) develop a new generation of GEZIs to address the hurdles for imaging secreted Zn2+ in the brain in vivo, and (2) integrate the new GEZIs with our innovative ZnT3 cKO mice, which, for the first time, allow for Cre-dependent expression of exogenous genes in ZnT3-expressing neurons and Dre-dependent region- and cell type-specific conditional ZnT3 gene knockout, to identify the cell- and circuit-specificity of Zn2+ dynamics that shape cortical sound processing. The project will lead to a novel capability of imaging synaptically released Zn2+ in the brain in awake behaving animals. Our innovative strategy to optimize the exoplasmic location of GEZIs may be generalized to enhance other genetically encoded indicators. Furthermore, because synaptic Zn2+ is a potent modulator throughout the cortex, our findings on Zn2+ dynamics in the primary auditory cortex (A1) during sound processing will improve the understanding of the roles of synaptic Zn2+ in cortical information processing beyond sensory cortices. We expect our studies to catalyze an extensive array of studies on Zn2+-related neurobiology and neurological diseases.

抽象的尽管突触 Zn2+ 作为整个大脑中新兴的神经调节剂的重要性已被广泛认识到。众所周知，响应自然发生的刺激的突触 Zn2+ 释放动态在很大程度上仍然存在难以捉摸。源自荧光蛋白的基因编码 Zn2+ 指示剂 (GEZI) 是流行的工具对细胞质和细胞内细胞器中的 Zn2+ 进行成像。然而，Zn2+分泌的荧光成像由于当前 GEZIs 的局限性（例如，不足），活体动物大脑尚未实现细胞外膜定位、亲和力不匹配和/或动态范围和光稳定性不足）。这个跨学科、多 PI 为期 4 年的 R01 项目，由具有基因编码专业知识的艾辉旺博士领导指标和荧光成像以及 Thanos Tzounopoulos 博士在研究 Zn2+ 在听觉处理，旨在 (1) 开发新一代 GEZI 以解决分泌成像的障碍体内大脑中的 Zn2+，以及 (2) 将新的 GEZIs 与我们创新的 ZnT3 cKO 小鼠相结合，这是第一个时间，允许表达 ZnT3 的神经元中外源基因的 Cre 依赖性表达和 Dre 依赖性区域和细胞类型特异性条件 ZnT3 基因敲除，以确定 Zn2+ 的细胞和电路特异性塑造皮质声音处理的动力学。该项目将带来一种新的能力，可以对清醒时大脑中突触释放的 Zn2+ 进行成像动物。我们优化 GEZIs 外质位置的创新策略可以推广到增强其他基因编码指标。此外，由于突触 Zn2+ 是整个过程中的有效调节剂皮层，我们对声音处理过程中初级听觉皮层 (A1) 中 Zn2+ 动态的研究结果将会改善了解突触 Zn2+ 在感觉皮层以外的皮层信息处理中的作用。我们期望我们的研究能够促进 Zn2+ 相关神经生物学和神经病学的广泛研究疾病。