title: Production and validation of recombinant adeno-associated virus for channelrhodopsin expression in neurons.
abstract: Recent discovery of the light-activated ion channel, channelrhodopsin (ChR), has provided researchers a powerful and convenient tool to manipulate the membrane potential of specific cells with light. With genetic targeting of these channels and illumination of light to a specific location, the experimenter can selectively activate the voltage-gated ion channels (VGICs) of ChR-expressing cells, initiating electrical signaling in temporally and spatially precise manners. In neuroscience research, this can be used to study electrical signal processing within one neuron at the cellular level, or the synaptic connectivity between neurons at the circuitry level. To conduct experiments with ChRs, these exogenous channels need to be introduced into the cells of interest, commonly through a viral approach. This chapter provides an overview of the design, production, and validation of recombinant adeno-associated virus (rAAV) for ChR expression that can be used in vitro or in vivo to infect neurons. The virus produced can be used to conduct "optogenetic" experiments in behaving animals, in vitro preparations and cultured cells, and can be used to study signal transduction and processing at a cellular or circuitry level.
authors.affiliation: Department of Pharmacology, University of California San Diego, San Diego, CA, USA.
authors.forename: John Y
authors.initials: JY
authors.lastname: Lin
mesh_terms: D003001:Cloning, Molecular; D004274:DNA, Recombinant / Q000235:genetics*; D000229:Dependovirus / Q000235:genetics* / Q000502:physiology*; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D015203:Reproducibility of Results; D012243:Rhodopsin / Q000235:genetics*; D020079:Terminal Repeat Sequences / Q000235:genetics; D014779:Virus Replication
chemical_list: D004274:DNA, Recombinant; D012243:Rhodopsin