Journal
LIGHT-SCIENCE & APPLICATIONS
Volume 6, Issue -, Pages -Publisher
CHINESE ACAD SCIENCES, CHANGCHUN INST OPTICS FINE MECHANICS AND PHYSICS
DOI: 10.1038/lsa.2016.255
Keywords
biophotonics; multiphoton microscopy; optogenetics; temporal focusing; three-photon
Categories
Funding
- NIH [5-P41-EB015871-27, DP3-DK101024 01, 1-U01-NS090438-01, 1-R01-EY017656 -0, 6A1, 1-R01-HL121386-01A1, 1R24MH106075, 2R01DA029639, 1R01MH103910, 1R01GM104948, 5U54 CA151884-04, 9-P41-EB015871-26A1]
- Biosym IRG of Singapore-MIT Alliance Research and Technology Center
- Koch Institute for Integrative Cancer Research Bridge Initiative
- Hamamatsu Inc.
- Samsung GRO program
- Wellcome Trust [093831/Z/10/Z]
- MIT Media Lab
- New York Stem Cell Foundation-Robertson Award
- NSF [CBET 1053233]
- EMBO Longterm Fellowship
- NCI [R35 CA197743, P01 CA080124]
- Wellcome Trust [093831/Z/10/Z] Funding Source: Wellcome Trust
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Three-photon wide-field depth-resolved excitation is used to overcome some of the limitations in conventional point-scanning two-and three-photon microscopy. Excitation of chromophores as diverse as channelrhodopsins and quantum dots is shown, and a penetration depth of more than 700 mu m into fixed scattering brain tissue is achieved, approximately twice as deep as that achieved using two-photon wide-field excitation. Compatibility with live animal experiments is confirmed by imaging the cerebral vasculature of an anesthetized mouse; a complete focal stack was obtained without any evidence of photodamage. As an additional validation of the utility of wide-field three-photon excitation, functional excitation is demonstrated by performing three-photon optogenetic stimulation of cultured mouse hippocampal neurons expressing a channelrhodopsin; action potentials could reliably be excited without causing photodamage.
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