In a nutshell: Finding the way is easier with a map and the same is true when doing brain research. With new techniques like CLARITY, it’s now possible to follow nerve cell extensions into the spinal cord in unprecedented detail.

The big picture:

Just a few short years ago, working out how the brain connected to the spinal cord depended on dissection, and the microscopic analysis of tissue slices. But cutting distorts tissue, introducing inaccuracies. What’s more, the maps these techniques produce are usually two-dimensional – less than ideal for navigating the three-dimensional nervous system.

Fast forward to 2015, and this team has used a combination of conventional analysis and new 3D imaging techniques such as CLARITY to trace individual nerve cell projections or axons — some extending 4 cm — between the brain and the spinal cord of the mouse.

CLARITY is a technique that removes fat molecules from the tissues of the brain and spinal column, while preserving 3D structure. Proteins and nucleic acid molecules remain intact, and they can be coloured using antibodies and other markers tagged with fluorescent labels. The result: a semi-transparent brain with the business parts — for example, the wiring between nerve cells, the chemicals they release, and the genes that are active — captured in a 3-D colour snapshot.

Figure: CLARITY video of serotonergic fibres in mouse spinal cord.

Led by CIBF chief investigator George Paxinos at Neuroscience Research Australia and The University of New South Wales in Sydney, the team used CLARITY and other techniques to focus on axons extending between regions of the spinal cord and the hind brain that process information affecting movement and balance.

“Seeing the projections in three dimensions instead of two [means] we can follow the nerve fibres down and see exactly where they are within the spinal cord and where each one ends,” says Andy (Huazheng) Liang, post-doc and lead author on three of the studies. That continuity isn’t possible in two dimensions because the fibres move in and out of the view plane.

Next steps:
Mapping of neurons in ever-finer detail will continue, eventually creating a complete 3D atlas of the mouse brain and spinal cord. The maps will be used in studies of spinal cord injury, stroke and other diseases and conditions.

Sengul, G., Fu, Y., Yu, Y., & Paxinos, G. (2015). Spinal cord projections to the cerebellum in the mouse. Brain Structure and Function, 220(5), 2997-3009.

Liang, H., Bácskai, T., & Paxinos, G. (2015). Termination of vestibulospinal fibers arising from the spinal vestibular nucleus in the mouse spinal cord. Neuroscience, 294, 206-214.

Liang, H., Watson, C., & Paxinos, G. (2016). Terminations of reticulospinal fibers originating from the gigantocellular reticular formation in the mouse spinal cord. Brain Structure and Function, 221(3), 1623-1633.

Liang, H., Wang, S., Francis, R., Whan, R., Watson, C., & Paxinos, G. (2015). Distribution of raphespinal fibers in the mouse spinal cord. Molecular pain, 11(1), 1.

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