Design and characterization of a DNA-encoded, voltage-sensitive fluorescent protein.
Optical imaging of electrical activity has been suggested as a promising approach to investigate the multineuronal representation of information processing in brain tissue. While considerable progress has been made in the development of instrumentation suitable for high-speed imaging, intrinsic or extrinsic dye-mediated optical signals are often of limited use due to their slow response dynamics, low effective sensitivity, toxicity or undefined cellular origin. Protein-based and DNA-encoded voltage sensors could overcome these limitations. Here we report the design and generation of a voltage-sensitive fluorescent protein (VSFP) consisting of a voltage sensing domain of a potassium channel and a pair of cyan and yellow emitting mutants of green fluorescent protein (GFP). In response to a change in transmembrane voltage, the voltage sensor alters the amount of fluorescence resonance energy transfer (FRET) between the pair of GFP mutants. The optical signals respond in the millisecond time-scale of fast electrical signalling and are large enough to allow monitoring of voltage changes at the single cell level.
Sakai
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Laboratory for Neuronal Circuit Dynamics, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan.
Repunte-Canonigo
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V
Raj
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Knöpfel
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D000200:Action Potentials / Q000187:drug effects* / Q000502:physiology; D000818:Animals; D001921:Brain / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000138:chemical synthesis / Q000235:genetics*; D001330:Electronic Data Processing; D004594:Electrophysiology; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000138:chemical synthesis / Q000235:genetics*; D008856:Microscopy, Fluorescence; D009154:Mutation / Q000502:physiology; D009474:Neurons / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D009482:Neurophysiology; D010777:Photochemistry; D015221:Potassium Channels / Q000138:chemical synthesis / Q000235:genetics*; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary / Q000235:genetics
D008164:Luminescent Proteins; D015221:Potassium Channels; D049452:Green Fluorescent Proteins; D004247:DNA