%0 Journal Article %A Miao-Ping Chien %A Daan Brinks %A Yoav Adam %A William Bloxham %A Simon Kheifets %A Adam E. Cohen %T Two-photon photoactivated voltage imaging in tissue with an Archaerhodopsin-derived reporter %D 2017 %R 10.1101/211946 %J bioRxiv %P 211946 %X Robust voltage imaging in tissue remains a technical challenge. Existing combinations of genetically encoded voltage indicators (GEVIs) and microscopy techniques cannot simultaneously achieve sufficiently high voltage sensitivity, background rejection, and time resolution for highresolution mapping of sub-cellular voltage dynamics in intact brain tissue. We developed a pooled high-throughput screening approach to identify Archaerhodopsin mutants with unusual photophysical properties. After screening ~105 cells, we identified a novel GEVI, NovArch, whose 1-photon near infrared fluorescence is reversibly enhanced by weak 2-photon excitation. Because the 2-photon excitation acts catalytically rather than stoichiometrically, high fluorescence signals, optical sectioning, and high time resolution are achieved simultaneously, at modest 2-photon laser power. We developed a microscopy system optimized for NovArch imaging in tissue. The combination of protein and optical engineering enhanced signal contrast sufficiently to enable optical mapping of back-propagating action potentials in dendrites in acute mouse brain slice. %U https://www.biorxiv.org/content/biorxiv/early/2017/10/31/211946.full.pdf