Novel Luminescence Reporters of Neural Activity Partnered with Optogenetics

与光遗传学合作的新型神经活动发光记者

基本信息

批准号：
9130311
负责人：
CARL Hirschie JOHNSON
金额：
$ 19.61万
依托单位：
VANDERBILT UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9130311
关键词：
Accelerometer Animal Model Animals Bacteriorhodopsins Behavior Biological Assay Biological Models Bioluminescence Brain Brain region Cations Cells Circadian Rhythms Complex Corpus striatum structure Development Electrophysiology (science)Energy Transfer Exposure to Family Fluorescence Fluorescent Probes Health Hippocampus (Brain)Hypothalamic structure In Vitro Ions Kinetics Laboratories Light Light Cell Measurement Measures Mental Health Methodology Methods Monitor Mus National Institute of Mental Health Neurobiology Neurons Neurosciences Optical Methods Optics Penetration Pharmaceutical Preparations Photobleaching Phototoxicity Preparation Rattus Recording of previous events Reporter Research Research Project Grants Rodent Slice Synapses Techniques Technology Tissues Viral Vector base cell type cost effective in vivo in vivo imaging innovation luminescence luminescence resonance energy transfer minimally invasive neural circuit neural model neural stimulation new technology novel optogenetics physical conditioning presynaptic protein protein interaction ratiometric reconstitution relating to nervous system response screening sensor spatiotemporal tool vector voltage

项目摘要

DESCRIPTION (provided by applicant): A novel luminescence-based methodology for monitoring neural activity as a new tool for functional neuroscience will be developed in this project. Optogenetic methods for stimulating neural activity are revolutionizing neurobiological research in vitro and in vivo. Brief exposure to light of cells expressing channelrhodopsin-2 (ChR2) can elicit excitatory cation fluxes (or inhibitory ion fluxes with the bacteriorhodopsin bR) To date, the impact of optogenetic stimulation has usually been monitored by electrophysiological methods that are accurate and well characterized, but are difficult and expensive to implement in freely behaving animals in vivo and/or in multiple neurons simultaneously. Optogenetic stimulation would optimally be partnered with less invasive methods to monitor activity among many cells, such as by optical methods. Unfortunately, the currently preferred methods for optically measuring neural activity are based on fluorescence methods that are poorly matched with ChR2/bR because the fluorescence excitation needed to monitor synaptic activity can trigger ChR2 and/or bR. Moreover, fluorescence can photobleach probes and excite tissue autofluorescence that generates undesirable background. Luminescence is an alternate optical technology that avoids problems associated with fluorescence. This project will develop novel luminescence probes for neuronal activity that are genetically encodable and can be targeted to specific cell types and to specific cellular loci that are involved in neural activity. These probes will respond to neuronal activity by changing their luminescence intensity and/or luminescence spectrum. In the latter case, probes based on Bioluminescence Resonance Energy Transfer (BRET) will be modulated by neural activity so that the spectrum of luminescent emission changes when neurons are activated. Our new luminescence methodology will avoid the drawbacks of electrophysiology and fluorescence excitation (esp. off- target optogenetic stimulation, photobleaching & tissue autofluorescence), and will therefore optimally partner with optogenetic methods for in vitro and in vivo stimulation. These luminescence reporters of neural activity will be characterized in conjunction with optogenetic stimulation of hippocampal primary neurons and of brain slices that reconstitute neural circuits in vitro. In addition, viral vectors encoding these reporters will be used to introduce the probes to the brain in a minimally invasive manner so as to monitor neural activity in freely behaving rodents (i) over the circadian cycle from the hypothalamus, and (ii) before and after optogenetic brain stimulation of the cortex in vivo. This project is appropriate for the R21 Exploratory/Developmental Research Grant mechanism of the NIMH because it will develop new technologies and tools to advance spatiotemporal analyses of complex circuits and cellular interactions in the brains of multiple model animal species.

描述（由申请人证明）：新型工具功能将在该项目中开发出来。 BR）通常通过在体内和/或在多个神经元中实施的e遗传学刺激用于测量神经活动的光学是与CHR2/BR相匹配的荧光，因为荧光性静脉疾病可以触发CHR2和/或BR，荧光可以光线探针和激发探针产生LE弹性。与荧光相关。这些探针参与了神经元活动，通过改变其发光强度和/或发光，基于生物发光谐振能量转移（BRET）将受到神经活动的调节被激活。神经活性的发光记者将与海马神经元的光遗传学刺激和大脑在体外的脑部切片，并以微创的方式进行体外。该项目适用于R21探索性/发展研究授予技术机理，用于多种模型动物物种的大脑中遵守的阶梯层次的临时性。