A flexible two-photon fiberscope for fast activity imaging and precise optogenetic photostimulation of neurons in freely moving mice

• Published in: Neuron (Cambridge, Mass.) Mass.), 2023-01, Vol.111 (2), p.176-189.e6
• Main Authors: Accanto, Nicolò, Blot, François G.C., Lorca-Cámara, Antonio, Zampini, Valeria, Bui, Florence, Tourain, Christophe, Badt, Noam, Katz, Ori, Emiliani, Valentina
• Format: Article
• Language: English
• Subjects: 2-photon calcium imaging; 2-photon optogenetic photostimulation; all-optical neuronal circuit manipulation; Animals; Bioengineering; Brain - physiology; computer-generated holography; Engineering Sciences; freely moving mice; GRIN lenses; Holography - methods; Imaging; Life Sciences; Mice; microendoscopy; Neurons - physiology; Neurons and Cognition; Opsins - genetics; optical fiber bundles; Optics; Optogenetics - methods; Photonic
• ISSN: 0896-6273; 1097-4199
• DOI: 10.1016/j.neuron.2022.10.030

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. •2P-FENDO achieves photostimulation at single-cell resolution in freely moving mice•2P-FENDO demonstrates reliable calcium imaging at 20 Hz in freely moving mice•Intrinsic temporal properties of fiber bundles enable single-cell axial resolution•2P-FENDO reaches 2P imaging until a depth of 150 μm in the mouse brain Accanto et al. developed a two-photon fiberscope (2P-FENDO) for the selective and reliable optogenetic photostimulation of neurons and high-speed calcium imaging in freely moving mice. 2P-FENDO enables stable recordings over hours without obstructing animal movement, thus representing a key technology to precisely investigate neuronal circuits and understand how they influence behavior.