All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins

By D. Hochbaum, Yongxin Zhao, S. Farhi, Nathan C. Klapoetke, C. Werley, Vikrant Kapoor, Peng Zou, Joel M. Kralj, Dougal Maclaurin, Niklas Smedemark-Margulies and 13 more. Nature Methods, 2014

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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.

Subfields: Cellular and Molecular Neuroscience, Cognitive Neuroscience

View Paper: https://www.semanticscholar.org/paper/a1370fe973b2e4762dadf90b74efef68a8d04f60

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Citations in the corpus

Machine learning to design integral membrane channelrhodopsins for efficient eukaryotic expression and plasma membrane localization (2017, background): “Two of the three had localization properties similar to the CheRiff parent (Fig 6B).”
A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters (2018, positive): “The Ace2N-4aa-mNeon, Archer1-KGC-EGFP-ER2 and Quasar2-mOrange-KGC-ER2 genes were synthesized de novo by GenScript, based on the sequences reported in the original publications 12,13,16 .”
Closed-loop and activity-guided optogenetic control. (2015, positive): “…set point rather than aiming to simply silence neural firing, allowing a more nuanced approach to optogenetic inhibition. from Mattis et al., 2012; Hochbaum et al., 2014; Inoue et al., 2015; Wu et al., 2013; Yizhar et al., 2011b; Klapoetke et al., 2014; Lin et al., 2013; Akerboom et al., 2012,…”
The growing and glowing toolbox of fluorescent and photoactive proteins (2016, background): “With the rapid adoption of optogenetic approaches and recent advances towards achieving all optical observation and manipulation of biological pathways [159], the potential utilization of FPs for simultaneous observation and manipulation may further revolutionize the way intracellular events are stu”