Continuously tracing brain-wide long-distance axonal projections in mice at a one-micron voxel resolution

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Revealing neural circuit mechanisms is critical for understanding brain functions. Significant progress in dissecting neural connections has been made using optical imaging with fluorescence labels, especially in dissecting local connections. However, acquiring and tracing brain-wide, long-distance neural circuits at the neurite level remains a substantial challenge. Here, we describe a whole-brain approach to systematically obtaining continuous neuronal pathways in a fluorescent protein transgenic mouse at a one-micron voxel resolution. This goal is achieved by combining a novel resin-embedding method for maintaining fluorescence, an automated fluorescence micro-optical sectioning tomography system for long-term stable imaging, and a digital reconstruction-registration-annotation pipeline for tracing the axonal pathways in the mouse brain. With the unprecedented ability to image a whole mouse brain at a one-micron voxel resolution, the long-distance pathways were traced minutely and without interruption for the first time. With advancing labeling techniques, our method is believed to open an avenue to exploring both local and long-distance neural circuits that are related to brain functions and brain diseases down to the neurite level.


▸ New device for 3-D imaging of fluorescence mouse brain at 1 μm voxel resolution ▸ Novel resin-embedding method for transgenic fluorescence labeled mouse brain ▸ Uninterrupted tracing of brain-wide, long-distance axonal projections ▸ Revealed several unreported and putative projection pathways in the mouse brain

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