Real-time 3D movement correction for two-photon imaging in behaving animals

Two-photon microscopy is widely used to investigate brain function across multiple spatial scales. However, measurements of neural activity are compromised by brain movement in behaving animals. Brain motion-induced artifacts are typically corrected using post hoc processing of two-dimensional image...

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Bibliographic Details
Published in:	Nature methods Vol. 17; no. 7; pp. 741 - 748
Main Authors:	Griffiths, Victoria A., Valera, Antoine M., Lau, Joanna YN, Roš, Hana, Younts, Thomas J., Marin, Bóris, Baragli, Chiara, Coyle, Diccon, Evans, Geoffrey J., Konstantinou, George, Koimtzis, Theo, Nadella, K. M. Naga Srinivas, Punde, Sameer A., Kirkby, Paul A., Bianco, Isaac H., Silver, R. Angus
Format:	Journal Article
Language:	English
Published:	New York Nature Publishing Group US 01-07-2020 Nature Publishing Group
Subjects:	631/1647/328 631/1647/328/2057 631/1647/334/1874/345 631/378 Acousto-optics Animal models Animals Balances (scales) Bioinformatics Biological Microscopy Biological Techniques Biomedical and Life Sciences Biomedical Engineering/Biotechnology Brain Dendrites Digital integrated circuits Female Field of view Field programmable gate arrays Fluorescence Imaging Imaging, Three-Dimensional - methods Life Sciences Male Mice Mice, Inbred C57BL Microscopy Microscopy, Fluorescence, Multiphoton - methods Movement Neuroimaging Neurons Photons Proteomics Real time Synapses Three dimensional motion Zebrafish United Kingdom
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Summary:	Two-photon microscopy is widely used to investigate brain function across multiple spatial scales. However, measurements of neural activity are compromised by brain movement in behaving animals. Brain motion-induced artifacts are typically corrected using post hoc processing of two-dimensional images, but this approach is slow and does not correct for axial movements. Moreover, the deleterious effects of brain movement on high-speed imaging of small regions of interest and photostimulation cannot be corrected post hoc. To address this problem, we combined random-access three-dimensional (3D) laser scanning using an acousto-optic lens and rapid closed-loop field programmable gate array processing to track 3D brain movement and correct motion artifacts in real time at up to 1 kHz. Our recordings from synapses, dendrites and large neuronal populations in behaving mice and zebrafish demonstrate real-time movement-corrected 3D two-photon imaging with submicrometer precision. Real-time 3D movement correction by tracking a fluorescent bead in the field of view enables functional imaging with 3D two-photon random-access microscopy in behaving mice and zebrafish.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 VG, and RAS conceived the project and RAS supervised it. VG, GK, K.M.N.S.N., P.A.K. and R.A.S designed the RT-3DMC system. VG and GK developed the closed loop FPGA code. A.V., B.M. and G.E. developed the MATLAB imaging software; P.A.K., K.M.N.S.N., G.K. and R.A.S. designed and built the AOL microscope. K.M.N.S.N., T.K. and S.A.P. developed and tested the LabVIEW imaging software. V.G., A.V., and R.A.S. designed the experiments on mice. D.C., T.Y., H.R. and C.B. performed viral transduction and surgeries. A.V, H.R., TY and C.B. performed the mouse experiments. J.Y.N.L., A.V., I.H.B. and R.A.S. designed the experiments on fish. J.Y.N.L. and A.V. performed the fish experiments. A.V. and V.G. analysed the data and AV made most of the figures. V.G., A.V. and R.A.S. wrote the manuscript with contributions from all authors. Equal contribution Victoria A. Griffiths and Antoine M. Valera Author contributions
ISSN:	1548-7091 1548-7105
DOI:	10.1038/s41592-020-0851-7