Harnessing plasticity to repair spinal cord injury

Background
Unlike mammals, zebrafish possess the ability to regenerate their spinal cords following injury. Thus, this presents a fascinating model organism to explore spinal cord injury in and an alternative method of researching it, as rather than investigating what is 'going wrong' in the mammal, we can investigate what is 'going right' in the fish. The Becker group are interested in elucidating the mechanisms occurring during this regeneration process so that they may be applied to spinal cord injury in humans. The group have previously found that a new neuronal cell type is generated from spinal stem cells and bridges across the injury site, which is hypothesised to contribute to the functional recovery of the fish (Ohnmacht et al., 2016 and Tsarouchas et al., 2018). They expresses marker genes for motor neurons but are not present in the uninjured fish and are thought to act as relay neurons connecting the rostral and caudal spinal cord, potentially showing similarity to transplanted neurons that have been used in mammalian spinal cords and therefore this cell type is of high interest.
Project description
This project is focused on this new neuronal cell type. It will aim to shed further light on the characteristics of these new neurons to better understand their contribution to functional recovery. Firstly, since the neurotransmitter identity of these newly formed neurons is not yet known, there will be an investigation of the specific neurotransmitter(s) by genetic and/or immunohistochemical methods. This can be further validated by the utilisation of various agonists and/or antagonists of their receptors, and by genetic manipulation using the CRISPR/Cas9 system. Another aspect of the project will involve using a previously established dataset involving genes that are upregulated in the zebrafish following injury, to guide knockout studies via CRISPR. Study of swimming behaviour and immunohistochemistry will help to identify the impact of the gene knockout on the ability to regain swimming function and the number of new neurons bridging the injury site. This will elucidate any genes that are useful/essential for the spinal cord regeneration process.
Finally, cell lineage tracing and electrophysiology will be utilised to determine some functional electrophysiological characteristics of these new neurons at different timepoints and provide information on the network changes that have occurred as a result of the injury and regeneration. It can be hypothesised that there may be increased firing as the neurons mature and that these may initially have more depolarised resting membrane potentials which will gradually reach normal potentials, these parameters could be expected to be aligned with the ventral root output and swimming recovery of the fish larvae. All initial work will be carried out on the zebrafish embryo/larvae and potentially taken further into adults if findings are of interest. Information obtained from this project will allow a greater understanding of the role that these neurons play in regeneration, as well as the plastic network changes that have occurred, and may provide potential future therapeutic targets for spinal cord repair in mammalian systems.
References
Ohnmacht, J., Yang, Y., Maurer, G., Barreiro-Iglesias, A., Tsarouchas, T., Wehner, D., Sieger, D., Becker, C. and Becker, T. (2016). Spinal motor neurons are regenerated after mechanical lesion and genetic ablation in larval zebrafish. Development, 143(9), pp.1464-1474
Tsarouchas, T., Wehner, D., Cavone, L., Munir, T., Keatinge, M., Lambertus, M., Underhill, A., Barrett, T., Kassapis, E., Ogryzko, N., Feng, Y., van Ham, T., Becker, T. and Becker, C. (2018). Dynamic control of proinflammatory cytokines Il-1 and Tnf-a by macrophages in zebrafish spinal cord regeneration. Nature Communications, 9(1).