Chemigenetic voltage indicators for far-red and two-photon imaging in vivo
用于体内远红和双光子成像的化学遗传学电压指示器
基本信息
- 批准号:10731843
- 负责人:
- 金额:$ 216.49万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2023
- 资助国家:美国
- 起止时间:2023-09-01 至 2026-08-31
- 项目状态:未结题
- 来源:
- 关键词:Action PotentialsAmino AcidsAnimal ModelAnimalsBehaviorBiological AvailabilityBrainCellsColorCouplesDependenceDevelopmentDiseaseDyesElectron TransportElectronicsElectronsElectrophysiology (science)EventFamilyFluorescenceFluorescence Resonance Energy TransferFluorescent DyesGeneticGenetic EngineeringHealthImageIndividualInvestigationLanguageLightMapsMeasurementMembraneMembrane PotentialsMolecularMolecular ConformationMotionMusNervous SystemNeuronsNeurosciencesOpticsPenetrationPhotonsPhototoxicityProcessPropertyProtein EngineeringProteinsReporterReportingResearchResolutionSensorySignal TransductionSliceSpecificitySpeedStructureSystemTechniquesTertiary Protein StructureTestingTimeTryptophanValidationVariantWorkcell typedesignexperimental studyfluorophorehybrid proteinimprovedin vivoin vivo evaluationin vivo fluorescence imagingin vivo two-photon imagingmillisecondmodel organismneural circuitnoveloptogeneticspatch clamppostsynapticpreventprototyperesponsescaffoldsensorsmall moleculespatiotemporaltooltransmission processtwo-photonvoltage
项目摘要
PROJECT SUMMARY
DESCRIPTION (provided by applicant): Changes in membrane potential are the fundamental language of the nervous system, but these voltage signals are not directly visible. Existing membrane voltage sensors impose severe constraints on the depth, duration, and field of view of in vivo voltage imaging. The development of brighter, redder, and two-photon (2P) compatible voltage indicators would dramatically increase the number of brain structures accessible to voltage imaging and would also enable qualitatively new types of measurements which could be transformative for neuroscience. This proposal will develop a family of hybrid protein-small molecule (chemogenetic) voltage sensors based on a new sensing mechanism, photoinduced electron transfer (PET). Genetically encoded PET voltage sensors will accept diverse bioavailable HaloTag dyes to report membrane voltage via one-photon (1P) or 2P imaging. This approach combines the exquisite molecular specificity of genetically encoded proteins with the superior photophysical properties of synthetic fluorophores. Proof-of-principle experiments demonstrated chemogenetic voltage sensor proteins (termed HaloVSDs) loaded with a far-red bioavailable dye. These HaloVSDs reported subthreshold voltages and spikes in cultured neurons with excellent sensitivity and speed. In Aim 1, the team will evolve this scaffold to create improved far-red PET-based chemogenetic voltage sensors. The sensors will undergo detailed photophysical characterization and will be validated in mice in vivo. In Aim 2, the team will generate a palette of 2P-compatible voltage sensors (HaloVSD-2P) for accessible 2P imaging using 1000–1300 nm excitation wavelengths. HaloVSD-2P will be a modular platform that can be used with multiple bright, photostable, and bioavailable dyes. In Aim 3, the team will combine the HaloVSDs with channelrhodopsins for a bidirectional optical neuro-electronic interface, i.e., all-optical electrophysiology. These tools will be used to construct functional connectivity maps in vivo. Due to their high brightness, HaloVSDs require ~100-fold less excitation light compared to existing far-red Achaerhodopsin- derived voltage sensors. This will minimize fluorescence background, phototoxicity, and bleaching, and will prevent spurious red-light activation of channelrhodopsins. These tools will enable robust crosstalk-free all- optical electrophysiology experiments in live animals. HaloVSDs will provide neuroscientists with unprecedented means of investigating animal models with all-optical interrogation of circuit dynamics. Because they are genetically encoded, these sensors can be easily introduced to various model organisms and will be of broad use in studies of brain circuit function in health and disease.
项目总结
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Ahmed Abdelfattah其他文献
Ahmed Abdelfattah的其他文献
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{{ truncateString('Ahmed Abdelfattah', 18)}}的其他基金
Lighting up the brain: Optogenetic tools to record, trace, and manipulate brain circuits at cellular resolution
点亮大脑:以细胞分辨率记录、追踪和操纵大脑回路的光遗传学工具
- 批准号:
10244755 - 财政年份:2021
- 资助金额:
$ 216.49万 - 项目类别:
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