A novel approach to examine slow synaptic transmission in vivo

一种检查体内缓慢突触传递的新方法

基本信息

批准号：
9604295
负责人：
Tianyi Mao
金额：
$ 10万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-05-01 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9604295
关键词：
Accelerometer Adrenergic Antagonists Animal Behavior Animals Axon Behavior Benchmarking Brain Calcium Cells Chemicals Communication Complement Cyclic AMP Cyclic AMP-Dependent Protein Kinases Defect Development Dopamine Electrodes Event Exposure to Fluorescence Resonance Energy Transfer Foxes Functional disorder Future Glutamates Habitats Head Image In Situ In Vitro Individual Investigation Light Measurement Measures Mediating Methods Microdialysis Modernization Monitor Mus Neuromodulator Neurons Noise Norepinephrine Pathway interactions Pattern Performance Periodicity Play Reagent Regulation Reporting Reproducibility Resolution Role Scanning Signal Transduction Signal Transduction Pathway Smell Perception Somatosensory Cortex Source Stress Synaptic Transmission Synaptic plasticity Technology Urine Walking adaptive optics adenylate kinase base brain tissue contrast imaging experimental study fluorescence lifetime imaging gamma-Aminobutyric Acid hippocampal pyramidal neuron imaging approach imaging modality improved in vivo in vivo imaging light scattering locus ceruleus structure nervous system disorder neuroregulation noradrenergic novel novel strategies optogenetics public health relevance response sensor sound spatiotemporal treadmill two-photon voltage

项目摘要

DESCRIPTION (provided by applicant): Two primary modes of chemical communication occur between neurons in the brain: fast synaptic transmission, such as that mediated by glutamate and GABA, which directly control the electrical activities of neurons, and slow synaptic transmission, such as that mediated by norepinephrine and dopamine, which regulate subcellular signaling events that cannot be measured directly from neuronal electrical activities. Slow synaptic transmission, which is also called neuromodulation, plays important modulatory roles in regulating excitability, synaptic plasticity and other aspects of neuronal function, and eventually imposes powerful control over the function of fast synaptic transmission. However, unlike fast synaptic transmission, which can be monitored directly via an increasing number of modern approaches such as multi-electrode recording, voltage imaging and calcium imaging methods, much less is known about the precise neuromodulatory events that occur in living animals because there has not been an established method to reliably record the relevant activities triggered by neuromodulation in individual neurons in vivo. To overcome this problem, we propose a novel approach for examining neuromodulatory activities with single-neuron resolution in vivo by imaging the activity of cyclic AMP (cAMP) and protein kinase A (PKA). The cAMP/PKA pathway is a common downstream signal transduction pathway for both dopamine and norepinephrine. Although genetically encoded cAMP/PKA sensors based on Förster resonance energy transfer (FRET) have been used for experiments in vitro, their application in vivo has been difficult due to lower signal-to-noise ratios under the more challenging in vivo imaging conditions. We propose a multipronged approach to eliminate several bottlenecks encountered with current FRET imaging approaches to maximize the signal-to-noise ratio. Our approach includes: 1) developing and improving cAMP/PKA sensors, 2) implementing a FRET imaging modality that is more effective than conventional FRET measures in light-scattering brain tissue, 3) correcting light aberrations associated with in vivo imaging conditions, and 4) developing novel mouse reagents for high-contrast, reproducible FRET imaging. We will validate the utility of this method for monitoring neuromodulatory activities by determining the spatiotemporal patterns of norepinephrine action in anesthetized mice using optogenetic approaches and in behaving mice using different stress stimulations. If successful, our efforts will provide a previously unattainable ability to conduct large- scale monitoring of neuromodulatory activities in the brain at the cellular and circuitry levels. This ability to quantitate neuromodulation will complement the measurements of fast synaptic transmission to enhance our understanding of brain function underlying animal behavior.

描述（由申请人提供）：大脑神经元之间发生化学通讯的两种主要模式：快速突触传递，例如由谷氨酸和 GABA 介导的传递，它们直接控制神经元的电活动；以及慢速突触传递，例如由谷氨酸和 GABA 介导的传递。由去甲肾上腺素和多巴胺介导，调节无法直接从神经元电活动测量的亚细胞信号传导事件，也称为神经调节，在调节中发挥重要的调节作用。兴奋性、突触可塑性和神经功能的其他方面，并最终对快速突触传递的功能施加强大的控制。然而，与可以通过越来越多的现代方法（例如多电极记录）直接监测的快速突触传递不同，电压成像和钙成像方法，对活体动物中发生的精确神经调节事件知之甚少，因为还没有一种既定的方法来可靠地记录体内单个神经元的神经调节触发的相关活动来克服这个问题，我们提出了一种通过对环 AMP (cAMP) 和蛋白激酶 A (PKA) 的活性进行成像来检查体内单神经元分辨率的神经调节活性的新方法。cAMP/PKA 途径是多巴胺和蛋白激酶的常见下游信号转导途径。尽管基于福斯特共振能量转移（FRET）的基因编码cAMP/PKA传感器已用于体外实验，但由于信噪比较低，其在体内的应用一直很困难。在更具挑战性的体内成像条件下，我们提出了一种多管齐下的方法来消除当前 FRET 成像方法遇到的几个瓶颈，以最大限度地提高信噪比，我们的方法包括：1）开发和改进 cAMP/PKA 传感器，2）在光散射脑组织中实施比传统 FRET 测量更有效的 FRET 成像方式，3) 校正与体内成像条件相关的光像差，以及 4) 开发新型小鼠试剂我们将通过使用光遗传学方法确定麻醉小鼠和使用不同应激刺激的行为小鼠的去甲肾上腺素作用的时空模式来验证这种方法用于监测神经调节活动的实用性。在细胞和电路水平上对大脑中的神经调节活动进行大规模监测的能力是以前无法实现的，这种定量神经调节的能力将补充测量结果。快速突触传递以增强我们对动物行为背后的大脑功能的理解。