A hybrid approach to measuring electrical activity in genetically specified neurons.
The development of genetically encoded fluorescent voltage probes is essential to image electrical activity from neuronal populations. Previous green fluorescent protein (GFP)-based probes have had limited success in recording electrical activity of neurons because of their low sensitivity and poor temporal resolution. Here we describe a hybrid approach that combines a genetically encoded fluorescent probe (membrane-anchored enhanced GFP) with dipicrylamine, a synthetic voltage-sensing molecule that partitions into the plasma membrane. The movement of the synthetic voltage sensor is translated via fluorescence resonance energy transfer (FRET) into a large fluorescence signal (up to 34% change per 100 mV) with a fast response and recovery time (0.5 ms). Using this two-component approach, we were able to optically record action potentials from neuronal cell lines and trains of action potentials from primary cultured neurons. This hybrid approach may form the basis for a new generation of protein-based voltage probes.
Chanda
Baron
B
Department of Physiology and Anesthesiology, David Geffen School of Medicine at the University of California Los Angeles, Los Angeles, California 90095, USA.
Blunck
Rikard
R
Faria
Leonardo C
LC
Schweizer
Felix E
FE
Mody
Istvan
I
Bezanilla
Francisco
F
D000200:Action Potentials / Q000502:physiology*; D000818:Animals; D001897:Boronic Acids / Q000633:toxicity; D002460:Cell Line; D004558:Electric Stimulation / Q000379:methods; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics*; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D007093:Imidazoles / Q000633:toxicity; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques / Q000379:methods; D010851:Picrates / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D013997:Time Factors; D014162:Transfection / Q000379:methods
C502590:4-(4,5-diphenyl-1H-imidazol-2-yl)phenylboronic acid; D001897:Boronic Acids; D007093:Imidazoles; D010851:Picrates; D049452:Green Fluorescent Proteins; C011454:dipicrylamine