Objective: 1) Biofabricate constructs that mimic the cytoarchitecture and function of the nigrostriatal pathway, and 2) Microinject constructs to assess restoration of dopaminergic input to the striatum in a rat model of Parkinson’s disease.

Background: Currently available therapeutic strategies to ameliorate the loss of nigrostriatal input to the striatum either replace dopamine systemically or place dopaminergic neurons directly into the striatum, both of which have significant limitations.

Methods: Dopaminergic neurons were dervived from either rat embryonic ventral mesencephalon or human stem cells. Green florescent protein expression on the tyrosine hydroxylase promoter enabled fluorescent activated cell sorting (FACS). Neuronal micro-spheres were generated via “forced cell aggregation” and then seeded within hydrogel micro-columns to facilitate axonal tract extension in vitro as described previously (Struzyna, et al. 2017). Constructs were stereotaxically microinjected into rats with the aggregate end placed in the substantia nigra and the fully-grown axonal terminals ending in the striatum. Constructs were assessed via phase contrast microscopy, immunochemistry, and confocal microscopy, and dopaminergic release was assessed using fast-scanning cyclic voltammetry (FSCV).

Results: Dopaminergic micro-sphere seeding resulted in axonal extension at rates exceeding 350μm/day. Micro-tubular constructs were generated with unidirectional dopaminergic axonal tracts spanning >1cm by 1 month in vitro. FACS-based enrichment resulted in >50% dopaminergic neurons. Electrical stimulation resulted in evoked dopamine release as measured by FSCV. Similar constructs were also fabricated using human stem cell derived neurons, with >50% dopaminergic neurons. Following stereotaxic microinjection in rats, implanted neurons survived in the substantia nigra and integrated via local neurite extension, while the axonal projections to the striatum were maintained to >1 month post-transplant.

Conclusions: Implantable tissue engineered nigrostriatal pathways are capable of physically replacing lost dopaminergic neurons in the substantia nigra as well as their long-projecting axonal inputs to the striatum. Ongoing studies are assessing the ability of this strategy to restore striatal dopaminergic tone from the substantia nigra, thereby anatomically and functionally recapitulating the native pathway.

References: Struzyna, L. A., Adewole, D. O., Gordián-Vélez, W. J., Grovola, M. R., Burrell, J. C., Katiyar, K. S., et al. Anatomically Inspired Three-dimensional Micro-tissue Engineered Neural Networks for Nervous System Reconstruction, Modulation, and Modeling. J. Vis. Exp. (123), e55609, doi:10.3791/55609 (2017).

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MDS Abstracts - https://www.mdsabstracts.org/abstract/biofabricated-tissue-constructs-to-recapitulate-the-anatomy-and-functionality-of-the-nigrostriatal-pathway-in-parkinsons-disease/