Abstract
Parkinson’s disease (PD) is the second most common neurodegenerative disorder, with a rapidly growing prevalence worldwide. PD occurs from progressive degeneration of dopaminergic neurons in the midbrain, leading to symptoms such as motor deficits and tremour. The neuronal degeneration is linked to hallmarks such as mitochondrial and lysosomal dysfunction, and aggregation of the protein α-synuclein. While dopaminergic neurons are dying, other neurons in the same brain environment can evade death, suggesting a neuroprotective mechanism is lost in dopaminergic neurons during the disease development. Recent research has suggested another brain cell type, astrocytes, might play a key role in PD pathology, through direct or indirect contact with neurons. As there is currently no cure or treatment which slows down disease progression, understanding the role of astrocytes might provide a key therapeutic target. Unhealthy astrocytes could be detrimental to dopaminergic neurons, or healthy astrocytes might provide a protective mechanism. A multiple-hit hypothesis may suggest that the loss of a protective mechanism in the dopaminergic neurons is the first ‘hit’ in PD pathology, and loss of support from astrocytes is a second ‘hit’.
A model for a form of young-onset PD, resulting from mutations in the gene ATP13A2 (PARK9) has been established in the NLD lab in both induced Pluripotent Stem Cell-derived dopaminergic neurons and astrocytes. We were able to establish an indirect coculture of astrocytes and neurons by transferring astrocyte media to neurons. Neuronal health and morphology was assessed after maturation using immunocytochemistry and Sholl analysis. there was an increase in a lysosomal protein (called LAMP2) expression in PARK9-deficient (knockout of 2 exons leading to truncated protein) neurons receiving media from healthy astrocytes. We also observed a decrease in cell viability between the wildtype neurons and the PARK9-deficient dopaminergic neurons and increased levels of α-synuclein protein aggregation in PARK9-deficient dopaminergic neurons. Increases in neuronal complexity were observed in the astrocyte media treatment groups. A method to grow astrocytes and dopaminergic neurons together in vitro for future study about the direct contact of the two cell types was optimised utilising cell culture inserts.
Increasing neuronal complexity may allude to a compensatory mechanism facilitated by astrocytes during disease progression. The increase in LAMP2 expression in PARK9-deficient neurons treated with healthy astrocyte media suggests a potential protective mechanism from indirect contact, but it may be rendered redundant by the deficiency in PARK9. These neurons still had a decreased cell viability and increased expression of α-synuclein, suggesting that direct contact with astrocytes is needed to rescue dopaminergic neurons from degeneration.