In vivo three-photon microscopy of subcortical structures within an intact mouse brain
Open accessAbstract
Two-photon fluorescence microscopy (2PM)1 enables scientists in various fields including neuroscience 2,3, embryology 4, and oncology 5 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 tissue 3 or the insertion of optical probes 6,7. 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.
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- Authors
- Horton, N. G., Wang, K., Kobat, D., Clark, C. G., Wise, F. W., Schaffer, C. B., & Xu, C.
- Affiliations
- Cornell University (US)
- Corresponding author
- Xu, Chris
- Venue
- Nature Photonics
- Published
- 2013-01-20
- Volume
- 7
- Issue
- 3
- Pages
- 205-209
- ISSN
- 1749-4885 (ISSN-L)
- 1749-4893
- PMC
- PMC3864872
- OpenAlex
- W2150708165
- Cited by
- 1,505
- References
- 32
- Citations by year
- 2026: 84 · 2025: 102 · 2024: 136 · 2023: 98 · 2022: 146 · 2021: 126 · 2020: 123 · 2019: 122 · 2018: 119 · 2017: 127 · 2016: 105 · 2015: 111 · 2014: 70 · 2013: 36
- FWCI
- 65.07
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Photonics (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Science Foundation
- National Institutes of Health
- National Cancer Institute (R01CA133148)
- National Institute of General Medical Sciences (R21GM103461)
- National Institute of Biomedical Imaging and Bioengineering (R01EB014873)
- National Center for Research Resources (R21RR032392)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Optical Imaging and Spectroscopy Techniques
- Physical Sciences › Engineering › Biomedical Engineering › Nonlinear Optical Materials Studies
- Keywords
- three-photon microscopy, mouse brain, hippocampus, deep tissue imaging, tissue scattering, vascular imaging, infrared excitation, noninvasive in vivo imaging
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