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Simultaneous, Cell-Resolved, Bioluminescent Recording From Microcircuits

微电路同步、细胞解析、生物发光记录

基本信息

批准号：
10463819
负责人：
Nozomi Nishimura
金额：
$ 24.6万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10463819
关键词：
3-Dimensional Action Potentials Address Adopted Aequorin Affect Axon Behavioral Paradigm Bioluminescence Calcium Cells Code Collection Color Complex Coupled Data Dendrites Dependovirus Electrodes Electrophysiology (science)Equilibrium Fluorescence Functional Imaging Genetic Geometry Goals Grant Head Image Individual Infection Interneurons Ion Channel Knowledge Label Learning Light LoxP-flanked allele Maps Measurement Measures Methodology Methods Microinjections Microscope Microscopy Modernization Neurites Neurons Neurosciences Optics Pattern Plasmids Positioning Attribute Proteins Rabies Rabies virus Scanning Scheme Signal Transduction Structure Synapses Techniques Technology Tetanus Helper Peptide Tracer Viral Viral Vector Virus Work adeno-associated viral vector anterograde transport base brain volume calcium indicator cell type experimental study genetic approach high resolution imaging improved instrumentation light emission optical fiber optical imaging optogenetics postsynaptic presynaptic presynaptic neurons recombinase relating to nervous system retrograde transport technology development tool vector

项目摘要

Summary Measuring the activity of many individual neurons at once while knowing their wiring diagrams would provide exciting information on how the components of a network interact. Knowledge of wiring diagrams has rapidly improved due to advances in the field of connectomics, and capabilities for simultaneous measurement of many individual neurons has increased exponentially with large-scale recording techniques. However, it is still difficult to combine such measurements. Registering high-resolution imaging for tracing neural projections with electrophysiological measurements, such as electrode arrays, is extremely difficult. With optical imaging, such tracing is possible, but neural activity measurements are often limited to particular geometries, most commonly a single plane in z. Although new imaging advances for volumetric imaging have eased this limitation somewhat, complicated instrumentation puts such technologies out of reach for most labs. This proposal addresses this challenge by using multicolor aequorin-fluorescent proteins (Aeq-FPs) as both fluorescent structural tracers and functional indicators for recording calcium activity. Aeq-FPs are bioluminescent indicators of calcium concentration that emit light from the entire cell including the dendritic and axonal arbors. In the proposed scheme, each neuron will express a unique combination of Aeq-FP colors so that it is color-coded to have its own spectral signature. The activity of individual neurons can be distinguished from the spectrum of the emitted bioluminescence without resolving the spatial position of the origin of the light. This enables simultaneous recording of the activity of many cells in arbitrary spatial arrangements including from different layers in the cortex. Connected networks are identified by limiting expression of the Aeq-FPs to neurons that are one synapse away from “starter” cells using transsynaptic viral vectors (modified rabies for retrograde transport and adeno- associated viruses (AAVs) for anterograde transport). The unique color combinations expressed in each cell also facilitate structural tracing. With these combined technologies, the network of microcircuits defined by connectivity to a single “starter” cell will be traced in three dimensions and correlated to measurements of activity in a single trial. In Aim 1, the starter cell is postsynaptic from the network, so this data will show how the presynaptic network involving multiple different types of cells from across cortical layers affects starter cell activity. In Aim 2, the starter cell is presynaptic to the labeled network and will express channelrhodopsin. Optically stimulating the starter cell will show how the network activity is affected by the modulation of the single cell. Such measurement capabilities will enable new types of experiments relating structure and activity and could be readily adopted by many labs.

总结