Ultrafast Genetically Encoded Voltage Indicators Designed from First Principles

根据第一原理设计的超快基因编码电压指示器

基本信息

批准号：
9288761
负责人：
Brian Y Chow
金额：
$ 44.47万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-05-01 至 2022-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9288761
关键词：
Action Potentials Affective Animals Arginine Behavior Biliverdine Binding Biological Biophysical Process Biophysics Brain Brain Diseases Cells Characteristics Chemicals Chronic Cognition Cognitive Communities Contrast Sensitivity Custom Data Detection Directed Molecular Evolution Electric Capacitance Electrophysiology (science)Engineering Ensure Event Exhibits Film Fluorescence Fluorescence Spectroscopy Fluorochrome Frequencies Generations Gold Hippocampus (Brain)Histidine Image Immunohistochemistry Individual Kinetics Lipids Mammalian Cell Measures Membrane Membrane Potentials Membrane Protein Traffic Microscope Neurons Neurosciences Noise Optics Pathology Peptide Signal Sequences Performance Perfusion Phenylalanine Physiology Population Positioning Attribute Preparation Property Propionates Protein Engineering Proteins Protocols documentation Pyrroles Reagent Reporter Reporting Resistance Resolution Safety Scanning Sequence Homology Signal Transduction Slice Spectrum Analysis Speed Structural Protein Structure Synapses Synaptic Potentials Synaptic Transmission System Techniques Technology Thinness Tissues Variant Virus Whole-Cell Recordings Yeasts amphiphilicity awake base brain abnormalities brain tissue cell type chromophore cofactor design dichroism electric field fluorescence imaging improved in vivo innovation insight mathematical model millisecond multi-photon neural circuit next generation novel strategies optical spectra optogenetics patch clamp public health relevance quantum receptor relating to nervous system response scaffold temporal measurement tool trafficking transgene expression voltage

项目摘要

ABSTRACT: The ability to optically record neuronal electrical activity with the temporal and fine-feature waveform resolution on par with whole-cell patch clamp electrophysiology would permit the correlation, of the physiology of individual cells and cell types, to the neural circuit-level activity that underlies behaviors, cognition, and affective states observed in the normal and diseased brain. Genetically encoded voltage indicators (GEVIs) hold great promise for this purpose as cell-type specific probes, if their voltage-sensitivity, brightness, and temporal resolution can be engineered to provide the reliable detection of high frequency action potentials, the detection of sub-threshold “minis” critical to synaptic scaling and homeostatic plasticity, and the ability to resolve waveforms useful for deducing specific channel/receptor contributions to spiking and synaptic transmission. We propose to invent next-generation GEVIs through rational design from first principles of non- biologically derived proteins. We will adapt artificial protein “maquettes,” which are de novo-designed and rigid 4-helix bundle proteins that serve as custom scaffolds for arbitrarily positioning biological co-factors within the scaffold core. Strategic positioning of a biliverdin chromphore within a transmembrane maquette allows for voltage sensing by the optical Stark effect, in which chromophores exhibit electric field-induced changes in absorbance that result in ultrafast changes in observed fluorescence. We call these proteins, “MASTERs” (Maquette Stark Effect Reporters). The ultrafast infrared-fluorescent reporters will recapitulate whole-cell recordings with no observable delay or waveform difference.

抽象的：使用临时和精细波形通过光学记录神经元电活动的能力与全细胞贴片夹电生理学的分辨率将允许生理的相关性基于行为，认知和情感状态在正常和解剖的大脑中观察到。遗传编码的电压指标（GEVIS）如果其电压敏感性，亮度和可以设计临时分辨率，以提供对高频动作电位的可靠检测检测对突触缩放和稳态可塑性至关重要的子阈值“ minis”的检测解析波形可用于推论特定的通道/受体贡献对尖峰和突触的贡献传播。我们建议通过从非 - 生物衍生的蛋白质。我们将适应人造蛋白“ maquettes”，这些蛋白质是从头设计和僵化的 4-螺旋束蛋白作为定制脚手架，以任意定位生物辅助因子脚手架核心。在跨膜maquette中，双列氏蛋白染色体的战略定位允许通过光学鲜明效应传感的电压传感，其中发色团暴露了电场诱导的变化吸光度会导致观察到的荧光的超快变化。我们称这些蛋白质为“大师” （Maquette Stark效果记者）。超快红外荧光记者将概括整个细胞没有可观察到的延迟或波形差的记录。