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A toolbox of Cre-dependent optogenetic transgenic mice for light-induced activation and silencing

Abstract

Cell type–specific expression of optogenetic molecules allows temporally precise manipulation of targeted neuronal activity. Here we present a toolbox of four knock-in mouse lines engineered for strong, Cre-dependent expression of channelrhodopsins ChR2-tdTomato and ChR2-EYFP, halorhodopsin eNpHR3.0 and archaerhodopsin Arch-ER2. All four transgenes mediated Cre-dependent, robust activation or silencing of cortical pyramidal neurons in vitro and in vivo upon light stimulation, with ChR2-EYFP and Arch-ER2 demonstrating light sensitivity approaching that of in utero or virally transduced neurons. We further show specific photoactivation of parvalbumin-positive interneurons in behaving ChR2-EYFP reporter mice. The robust, consistent and inducible nature of our ChR2 mice represents a significant advance over previous lines, and the Arch-ER2 and eNpHR3.0 mice are to our knowledge the first demonstration of successful conditional transgenic optogenetic silencing. When combined with the hundreds of available Cre driver lines, this optimized toolbox of reporter mice will enable widespread investigations of neural circuit function with unprecedented reliability and accuracy.

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Figure 1: Generation and expression characterization of the Ai27, Ai32, Ai35 and Ai39 Cre-reporter lines.
Figure 2: Photostimulation of pyramidal neurons in cortical slices of E-Ai27, E-Ai32 and Ai32 alone (−Cre) mice.
Figure 3: Effective silencing of cortical pyramidal neurons by Arch-ER2 in E-Ai35 and eNpHR3.0 in E-Ai39 mice.
Figure 4: Alternative light sources for silencing of cortical pyramidal neurons in E-Ai35 and E-Ai39 mice.
Figure 5: Effective silencing of induced population bursting in the hippocampal circuit in E-Ai35 mice.
Figure 6: Optical activation or silencing of pyramidal neuron activities in the neocortex of awake E-Ai27, E-Ai32, E-Ai35 and Camk2a-CreERT2;Ai39 (C-Ai39) mice.
Figure 7: In vivo identification of light-activated neurons in the hippocampus and thalamus of Pvalb-IRES-Cre;Ai32 mice.

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Acknowledgements

We are grateful for the technical support of the Atlas Production Team, led by P. Wohnoutka, and the Technology Team, led by C. Dang, at the Allen Institute. We thank A. Nagy (Mount Sinai Hospital in Toronto) for providing the G4 ES cell line and K. Deisseroth (Stanford University) for providing the eNpHR3.0 construct. The authors wish to thank the Allen Institute founders, P.G. Allen and J. Allen, for their vision, encouragement and support. This work was funded by the Allen Institute for Brain Science and the Howard Hughes Medical Institute, US National Institutes of Health (NIH) grant DA028298 to H.Z., NIH grants MH90478 and MH093667 to E.E.T., NIH grant MH085944 and an Alfred P. Sloan Foundation grant to X.H., NIH grants NS034994 and MH54671 and a US National Science Foundation grant to G.B., and a Marie Curie Fellowship (EU FP7 PEOPLE 2009 IOF 254780) to A.B.

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Contributions

L.M., J.K. and H.G. generated the Cre reporter mouse lines. T.M., B.M.H. and K.S. conducted the slice physiology study on Ai27 and Ai32 mice. H.K., Y.-W.A.H., A.J.G., S.Z., J.M.R. and E.E.T. conducted the slice physiology study on Ai35 and Ai39 mice. J.Z., X.G., Y.M. and X.H. conducted the in vivo cortical recordings. A.B., S.F. and G.B. conducted the in vivo hippocampal and thalamic recordings. A.R.J. provided institutional support. E.S.B. provided the Arch-ER2 construct. L.M., T.M., H.K., J.Z., A.B., S.F., E.S.B., G.B., X.H., E.E.T. and H.Z. analyzed data and wrote the paper.

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Correspondence to Hongkui Zeng.

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Madisen, L., Mao, T., Koch, H. et al. A toolbox of Cre-dependent optogenetic transgenic mice for light-induced activation and silencing. Nat Neurosci 15, 793–802 (2012). https://doi.org/10.1038/nn.3078

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