All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins

Hochbaum, D.R.*, Zhao, Y.*, Farhi, S.L., Klapoetke, N.C., Werley, C.A., Kapoor, V., Zou, P., Kralj, J.M., Maclaurin, D., Smedemark-Margulies, N., Saulnier, J., Boulting, G.L., Straub, C., Cho, Y., Melkonian, M., Wong, G.K.-S., Harrison, D. J., Murthy, V.N., Sabatini, B., Boyden, E.S.**, Campbell, R.E.**, Cohen, A.E. (2014) All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins, Nature Methods, 11(8):825-33. (*, equal contribution, **, jointly directed work)

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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.

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