Abstract
Drosophila is widely used in connectome studies due to its small brain size, sophisticated genetic tools, and the most complete single-neuron-based anatomical brain map. Surprisingly, even the brain thickness is only 200-μm, common Ti:sapphire-based two-photon excitation cannot penetrate, possibly due to light aberration/scattering of trachea. Here we quantitatively characterized scattering and light distortion of trachea-filled tissues, and found that trachea-induced light distortion dominates at long wavelength by comparing one-photon (488-nm), two-photon (920-nm), and three-photon (1300-nm) excitations. Whole-Drosophila-brain imaging is achieved by reducing tracheal light aberration/scattering via brain-degassing or long-wavelength excitation at 1300-nm. Our work paves the way toward constructing whole-brain connectome in a living Drosophila.
Highlights
Drosophila is an important model animal to study connectomics since its brain is complex with 105 neurons but still small enough to be completely mapped by optical microscopy with single-cell resolution
The thickness of a Drosophila brain is only about 200 μm, which is much smaller than the typical imaging depth of 2PF microscopy in other model animals like mouse and zebrafish, to the best of our knowledge, no study has demonstrated in vivo whole-brain imaging in Drosophila with single-cell resolution, nor has characterized the image attenuation
We have, for the first time, characterized the optical properties of the Drosophila brain, which is filled with air, with single-photon, two-photon, and three-photon modalities
Summary
Drosophila is an important model animal to study connectomics since its brain is complex with 105 neurons but still small enough to be completely mapped by optical microscopy with single-cell resolution. When observing living mouse or zebrafish brain with 2PF microscopy, the penetration depth approaches 1 mm, which is typically limited to about five scattering lengths [4,5]. The thickness of a Drosophila brain is only about 200 μm, which is much smaller than the typical imaging depth of 2PF microscopy in other model animals like mouse and zebrafish, to the best of our knowledge, no study has demonstrated in vivo whole-brain imaging in Drosophila with single-cell resolution, nor has characterized the image attenuation of a living Drosophila brain. The whole-brain observation capability is a major milestone toward establishing complete connectome in this model animal
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