title: In vivothree-photon microscopy of subcortical structures within an intact mouse brain.
abstract: Two-photon fluorescence microscopy (2PM)1enables scientists in various fields including neuroscience2,3, embryology4, and oncology5to visualizein vivoandex vivotissue morphology and physiology at a cellular level deep within scattering tissue. However, tissue scattering limits the maximum imaging depth of 2PM within the mouse brain to the cortical layer, and imaging subcortical structures currently requires the removal of overlying brain tissue3or the insertion of optical probes6,7. Here we demonstrate non-invasive, high resolution,in vivoimaging of subcortical structures within an intact mouse brain using three-photon fluorescence microscopy (3PM) at a spectral excitation window of 1,700 nm. Vascular structures as well as red fluorescent protein (RFP)-labeled neurons within the mouse hippocampus are imaged. The combination of the long excitation wavelength and the higher order nonlinear excitation overcomes the limitations of 2PM, enabling biological investigations to take place at greater depth within tissue.
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Nicholas G
authors.initials: NG
authors.lastname: Horton
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Ke
authors.initials: K
authors.lastname: Wang
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Demirhan
authors.initials: D
authors.lastname: Kobat
authors.affiliation: Department of Biomedical Engineering, Cornell University, Ithaca, New York, 14853, USA.
authors.forename: Catharine G
authors.initials: CG
authors.lastname: Clark
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Frank W
authors.initials: FW
authors.lastname: Wise
authors.affiliation: Department of Biomedical Engineering, Cornell University, Ithaca, New York, 14853, USA.
authors.forename: Chris B
authors.initials: CB
authors.lastname: Schaffer
authors.affiliation: School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Chris
authors.initials: C
authors.lastname: Xu