Principles of two-photon excitation microscopy and its applications to neuroscience
Open accessAbstract
The brain is complex and dynamic. The spatial scales of interest to the neurobiologist range from individual synapses (approximately 1 microm) to neural circuits (centimeters); the timescales range from the flickering of channels (less than a millisecond) to long-term memory (years). Remarkably, fluorescence microscopy has the potential to revolutionize research on all of these spatial and temporal scales. Two-photon excitation (2PE) laser scanning microscopy allows high-resolution and high-sensitivity fluorescence microscopy in intact neural tissue, which is hostile to traditional forms of microscopy. Over the last 10 years, applications of 2PE, including microscopy and photostimulation, have contributed to our understanding of a broad array of neurobiological phenomena, including the dynamics of single channels in individual synapses and the functional organization of cortical maps. Here we review the principles of 2PE microscopy, highlight recent applications, discuss its limitations, and point to areas for future research and development.
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Two-photon microscopy rapidly became the method of choice for deep tissue imaging and has even enabled intravital brain imaging in mice.
“Two-photon microscopy rapidly became the method of choice for deep tissue imaging and has even enabled intravital brain imaging in mice (Svoboda and Yasuda, 2006).”
From Barriers in the brain: resolving dendritic spine morphology and compartmentalization (2014) · cites this paper as Svoboda and Yasuda, 2006
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- Authors
- Svoboda, K., & Yasuda, R.
- Affiliations
- Howard Hughes Medical Institute (US)
- Duke Medical Center (US)
- Corresponding author
- Svoboda, Karel
- Yasuda, Ryohei
- Venue
- Neuron
- Publisher
- Elsevier BV
- Published
- 2006-06
- Volume
- 50
- Issue
- 6
- Pages
- 823-839
- ISSN
- 0896-6273 (ISSN-L)
- OpenAlex
- W2041060053
- Cited by
- 1,027
- References
- 18
- Citations by year
- 2026: 30 · 2025: 54 · 2024: 44 · 2023: 43 · 2022: 65 · 2021: 51 · 2020: 56 · 2019: 75 · 2018: 75 · 2017: 67 · 2016: 60 · 2015: 51 · 2014: 48 · 2013: 35 · 2012: 65
- FWCI
- 3.87
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- www.elsevier.com/tdm/userlicense/1.0/ (tdm)
- www.elsevier.com/legal/tdmrep-license (tdm)
- www.elsevier.com/open-access/userlicense/1.0/ (vor, 2639 d)
- doi.org/10.15223/policy-017 (stm-asf)
- doi.org/10.15223/policy-037 (stm-asf)
- doi.org/10.15223/policy-012 (stm-asf)
- doi.org/10.15223/policy-029 (stm-asf)
- doi.org/10.15223/policy-004 (stm-asf)
- Locations
- Neuron (publishedVersion)
- PubMed (publishedVersion)
- Cold Spring Harbor Laboratory Institutional Repository (Cold Spring Harbor Laboratory) (acceptedVersion)
- Funders
- Burroughs Wellcome Fund
- Dana Foundation
- National Institutes of Health
- Howard Hughes Medical Institute
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
- Physical Sciences › Engineering › Biomedical Engineering › Nonlinear Optical Materials Studies
- Keywords
- two-photon microscopy, laser scanning microscopy, fluorescence microscopy, photostimulation, synapses, cortical maps, neural circuits
- MeSH terms
- Animals, Humans, Neurosciences, Microscopy, Confocal, Microscopy, Fluorescence, Multiphoton, Photobleaching
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