RESUMEN
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.
Asunto(s)
Holografía , Optogenética , Ratones , Animales , Optogenética/métodos , Holografía/métodos , Encéfalo/fisiología , Neuronas/fisiología , Opsinas/genéticaRESUMEN
Fiber-based micro-endoscopes are a critically important tool for minimally-invasive deep-tissue imaging. However, current micro-endoscopes cannot perform three-dimensional imaging through dynamically-bent fibers without the use of bulky optical elements such as lenses and scanners at the distal end, increasing the footprint and tissue-damage. Great efforts have been invested in developing approaches that avoid distal bulky optical elements. However, the fundamental barrier of dynamic optical wavefront-distortions in propagation through flexible fibers limits current approaches to nearly-static or non-flexible fibers. Here, we present an approach that allows holographic, bend-insensitive, coherence-gated, micro-endoscopic imaging using commercially available multi-core fibers (MCFs). We achieve this by adding a partially-reflecting mirror to the distal fiber-tip, allowing to perform low-coherence full-field phase-shifting holography. We demonstrate widefield diffraction-limited reflection imaging of amplitude and phase targets through dynamically bent fibers at video-rate. Our approach holds potential for label-free investigations of dynamic samples.