Objective: We previously developed the first micro-tissue engineered nigrostriatal pathway encased in a tubular hydrogel with 3D structure, multicellular composition, and architecture of long axonal tracts between distinct neuronal populations [1]. In the current study we apply this system as a biofidelic test-bed for modeling Parkinson’s disease (PD) in vitro and evaluating axonal pathophysiology.

Background: Selective loss of long-projecting neural circuitry is a common feature of many neurodegenerative diseases, such as the vulnerable nigrostriatal pathway in PD. Current in vitro approaches generally do not mimic complex anatomical features of the afflicted substrates such as long axonal pathways between stereotypical neural populations. Such exquisite features are crucial for neural systems function and may contribute to the preferential vulnerability of these structures in neurodegenerative disease.

Method: Dopaminergic neurons from the ventral mesencephalon and medium spiny neurons (MSNs) from the striatum were separately isolated from rat embryos, seeded and allowed to integrate in hydrogel micro-columns with a central extracellular matrix several centimeters in length. Constructs were treated with exogenous recombinant human alpha-synuclein active fibrils and immunocytochemistry (ICC) and confocal microscopy were used to assess health, cytoarchitecture, synaptic integration, and mitochondrial dynamics and fibril localization.

Results: TH+ dopaminergic neurons, DARPP-32+ MSNs and extensive axonal-dendritic integration and synapse formation were confirmed by ICC. Active alpha-synuclein fibrils demonstrated cytoplasmic staining and changes to axon length and density. Additionally, mitochondria structure along axonal tracts was observed using MitoTracker staining, revealing dynamic intra-axonal mitochondrial motility in this system. Ongoing studies are evaluating real-time mitochondrial dynamics and axonal physiology under both baseline conditions as well as following the addition of exogenous α-Synuclein fibrils to model synucleinopathy in PD.

Conclusion: This anatomically-inspired platform has neuronal-axonal architecture that structurally and functionally emulates the nigrostriatal pathway in vivo. We are applying this paradigm as a powerful in vitro test-bed for understanding mitochondrial activity and inter-axonal energetics pathways under homeostatic as well as PD pathological conditions.

References: [1] Struzyna et al., J Tiss Eng Regen Med, 2018. Tissue engineered nigrostriatal pathways for treatment of Parkinson’s disease.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/tissue-engineered-nigrostriatal-pathway-as-an-anatomically-inspired-test-bed-for-evaluating-axonal-pathophysiology/