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bioRxiv · 10.1101/339531

Whole-brain imaging and characterization of Drosophila brains based on one-, two-, and three-photon excitations

Chu, S.-W.·Lin, Y.-Y.·Chiang, A.-S.·Hsu, K.-J.
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Abstract

To study functional connectome, optical microscopy provides the advantages of in vivo observation, molecular specificity, high-speed acquisition, and sub-micrometer spatial resolution. Now, the most complete single-neuron-based anatomical connectome is built upon Drosophila; thus it will be a milestone to achieve whole-brain observation with sub-cellular resolution in living Drosophila. Surprisingly, two-photon microscopy cannot penetrate through the 200-m-thick brain, due to the extraordinarily strong aberration/scattering from tracheae. Here we achieve whole-Drosophila-brain observation by degassing the brain or by using three-photon microscopy at 1300-nm, while only the latter provides in vivo feasibility, reduced aberration/scattering and exceptional optical sectioning capability. Furthermore, by comparing one-photon (488-nm), two-photon (920-nm), and three-photon (1300-nm) excitations in the brain, we not only demonstrate first quantitative reduction of both scattering and aberration in trachea-filled tissues, but unravel that the contribution of aberration exceeds scattering at long wavelengths. Our work paves the way toward constructing functional connectome in a living Drosophila.

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Chu, S.-W., Lin, Y.-Y., Chiang, A.-S., Hsu, K.-J.. 2018-06-05. Whole-brain imaging and characterization of Drosophila brains based on one-, two-, and three-photon excitations. https://doi.org/10.1101/339531

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