In vivo three-photon microscopy of subcortical structures within an intact mouse brain
Nicholas G. Horton,Ke Wang,Demirhan Kobat,Catharine G. Clark,Frank W. Wise,Chris B. Schaffer,Chris Xu +6 more
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TLDR
Non-invasive, high-resolution, in vivo imaging of subcortical structures (the external capsule and hippocampus) within an intact mouse brain is demonstrated using three-photon fluorescence microscopy at the new spectral window of 1700 nm.Abstract:
Two-photon fluorescence microscopy (2PM)1 enables scientists in various fields including neuroscience2,3, embryology4, and oncology5 to visualize in vivo and ex vivo tissue 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 tissue3 or the insertion of optical probes6,7. Here we demonstrate non-invasive, high resolution, in vivo imaging 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.read more
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References
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Book
The Mouse Brain in Stereotaxic Coordinates
TL;DR: The 3rd edition of this atlas is now in more practical 14"x11" format for convenient lab use and includes a CD of all plates and diagrams, as well as Adobe Illustrator files of the diagrams, and a variety of additional useful material.
Journal ArticleDOI
Two-Photon Laser Scanning Fluorescence Microscopy
TL;DR: The fluorescence emission increased quadratically with the excitation intensity so that fluorescence and photo-bleaching were confined to the vicinity of the focal plane as expected for cooperative two-photon excitation.
Journal ArticleDOI
Deep tissue two-photon microscopy
Fritjof Helmchen,Winfried Denk +1 more
TL;DR: Fundamental concepts of nonlinear microscopy are reviewed and conditions relevant for achieving large imaging depths in intact tissue are discussed.
Journal ArticleDOI
Measurement of two-photon excitation cross sections of molecular fluorophores with data from 690 to 1050 nm
Chris Xu,Watt W. Webb +1 more
TL;DR: In this paper, the two-photon fluorescence excitation (TPE) spectra were measured for 11 common molecular fluorophores in the excitation wavelength range 690 nm < λ < 1050 nm.
Journal ArticleDOI
Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system
Jean Livet,Tamily A. Weissman,Hyuno Kang,Ju Lu,Robyn A Bennis,Joshua R. Sanes,Jeff W. Lichtman +6 more
TL;DR: Strategies to visualize synaptic circuits by genetically labelling neurons with multiple, distinct colours are presented and may facilitate the analysis of neuronal circuitry on a large scale.