Abstract
We developed a flexible two-photon microendoscope (2P-FENDO) capable of all-optical brain investigation at near cellular resolution in freely moving mice. The system performs fast two-photon (2P) functional imaging and 2P holographic photostimulation of single and multiple cells using axially confined extended spots. Proof-of-principle experiments were performed in freely moving mice co-expressing jGCaMP7s and the opsin ChRmine in the visual or barrel cortex. On a field of view of 250 μm in diameter, we demonstrated functional imaging at a frame rate of up to 50 Hz and precise photostimulation of selected groups of cells. With the capability to simultaneously image and control defined neuronal networks in freely moving animals, 2P-FENDO will enable a precise investigation of neuronal functions in the brain during naturalistic behaviors.
Original language | American English |
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Pages (from-to) | 176-189.e6 |
Journal | Neuron |
Volume | 111 |
Issue number | 2 |
DOIs | |
State | Published - 18 Jan 2023 |
Bibliographical note
Funding Information:We acknowledge support from the IHU FOReSIGHT grant (grant P-ALLOP3-IHU-000 ), the Fondation Bettencourt Schueller (Prix Coups d’élan pour la recherche française), the Axa research funding, the Agence National de la Recherche (grant ANR 2MEnHoloMD - 19-CE19-0001-01 ), the Human Frontiers Science Program (grant RGP0015/2016 ), Imagine Nano, the ERC Advanced Grant HOLOVIS ( ERC-2019-AdG ; award no. 885090 ), the ERC Horizon 2020 H2020-ICT (DEEPER, 101016787 ), the Israel Science Foundation ( 1361/18 ), and the Israeli Ministry of Science and Technology (grant 712845 ). We would like to thank Serge Charpak, Marine Tournissac, and Manon Omnès (Vision Institute, Paris) for their helpful discussions and pilot imaging experiments; Andrea Giovannucci for the help with CaImAn; Aaron Benson Wong (ErasmusMC Rotterdam) for the help with the calcium imaging data analysis; Vincent de Sars for the WaveFront IV software development; and Manuel Simonutti, Julie Degardin, and Quenol Cesar from the animal facility of the Institut de la vision for their help and support with the animal experimentation.
Funding Information:
We acknowledge support from the IHU FOReSIGHT grant (grant P-ALLOP3-IHU-000), the Fondation Bettencourt Schueller (Prix Coups d’élan pour la recherche française), the Axa research funding, the Agence National de la Recherche (grant ANR 2MEnHoloMD - 19-CE19-0001-01), the Human Frontiers Science Program (grant RGP0015/2016), Imagine Nano, the ERC Advanced Grant HOLOVIS (ERC-2019-AdG; award no. 885090), the ERC Horizon 2020 H2020-ICT (DEEPER, 101016787), the Israel Science Foundation (1361/18), and the Israeli Ministry of Science and Technology (grant 712845). We would like to thank Serge Charpak, Marine Tournissac, and Manon Omnès (Vision Institute, Paris) for their helpful discussions and pilot imaging experiments; Andrea Giovannucci for the help with CaImAn; Aaron Benson Wong (ErasmusMC Rotterdam) for the help with the calcium imaging data analysis; Vincent de Sars for the WaveFront IV software development; and Manuel Simonutti, Julie Degardin, and Quenol Cesar from the animal facility of the Institut de la vision for their help and support with the animal experimentation. N.A. A.L.-C. and C.T. built the optical setup. N.A. and A.L.-C. developed the acquisition software and performed the optical characterization. O.K. conceived the experiment for measuring the ICDD. N.B. measured and analyzed the ICDD. F.G.C.B. and C.T. designed and developed the implant for freely moving experiments. F.G.C.B. V.Z. and F.B. developed surgical procedures and performed viral injections. V.Z. and F.B. acquired in vivo and in vitro z stacks and preliminary calcium imaging data in vivo. F.G.C.B. and A.L.-C. performed imaging and photostimulation experiments in freely moving mice. N.A. F.G.C.B. and A.L.-C. analyzed the data. N.A. F.G.C.B. and V.E. wrote the paper with the contribution of all the authors. N.A. and V.E. conceived and supervised the project. The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Publisher Copyright:
© 2022 Elsevier Inc.
Keywords
- 2-photon calcium imaging
- 2-photon optogenetic photostimulation
- GRIN lenses
- all-optical neuronal circuit manipulation
- computer-generated holography
- freely moving mice
- microendoscopy
- optical fiber bundles