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Nat Methods. 2014 Aug;11(8):825-33. doi: 10.1038/nmeth.3000. Epub 2014 Jun 22.

All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins.

Author information

1
1] Applied Physics Program, School of Engineering and Applied Science, Harvard University, Cambridge, Massachusetts, USA. [2].
2
1] Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada. [2].
3
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, USA.
4
1] The MIT Media Laboratory, Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, USA. [2] Department of Biological Engineering, MIT, Cambridge, Massachusetts, USA. [3] Department of Brain and Cognitive Sciences, MIT, Cambridge, Massachusetts, USA. [4] McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts, USA.
5
Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts, USA.
6
Department of Physics, Harvard University, Cambridge, Massachusetts, USA.
7
Department of Neurobiology, Harvard Medical School, Boston, Massachusetts, USA.
8
Institute of Botany, Cologne Biocenter, University of Cologne, Cologne, Germany.
9
1] Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada. [2] Department of Medicine, University of Alberta, Edmonton, Alberta, Canada. [3] Beijing Genomics Institute-Shenzhen, Shenzhen, China.
10
Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada.
11
1] Department of Neurobiology, Harvard Medical School, Boston, Massachusetts, USA. [2] Howard Hughes Medical Institute, Harvard University, Cambridge, Massachusetts, USA.
12
1] The MIT Media Laboratory, Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, USA. [2] Department of Biological Engineering, MIT, Cambridge, Massachusetts, USA. [3] Department of Brain and Cognitive Sciences, MIT, Cambridge, Massachusetts, USA. [4] McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts, USA. [5].
13
1] Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, USA. [2] Department of Physics, Harvard University, Cambridge, Massachusetts, USA. [3] Howard Hughes Medical Institute, Harvard University, Cambridge, Massachusetts, USA.

Abstract

All-optical electrophysiology-spatially resolved simultaneous optical perturbation and measurement of membrane voltage-would open new vistas in neuroscience research. We evolved two archaerhodopsin-based voltage indicators, QuasAr1 and QuasAr2, which show improved brightness and voltage sensitivity, have microsecond response times and produce no photocurrent. We engineered a channelrhodopsin actuator, CheRiff, which shows high light sensitivity and rapid kinetics and is spectrally orthogonal to the QuasArs. A coexpression vector, Optopatch, enabled cross-talk-free genetically targeted all-optical electrophysiology. In cultured rat neurons, we combined Optopatch with patterned optical excitation to probe back-propagating action potentials (APs) in dendritic spines, synaptic transmission, subcellular microsecond-timescale details of AP propagation, and simultaneous firing of many neurons in a network. Optopatch measurements revealed homeostatic tuning of intrinsic excitability in human stem cell-derived neurons. In rat brain slices, Optopatch induced and reported APs and subthreshold events with high signal-to-noise ratios. The Optopatch platform enables high-throughput, spatially resolved electrophysiology without the use of conventional electrodes.

PMID:
24952910
PMCID:
PMC4117813
DOI:
10.1038/nmeth.3000
[Indexed for MEDLINE]
Free PMC Article

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