Disruption of dopamine homeostasis underlies selective neurodegeneration mediated by alpha-synuclein.
A key challenge in Parkinson's disease research is to understand mechanisms underlying selective degeneration of dopaminergic neurons mediated by genetic factors such as alpha-synuclein (alpha-Syn). The present study examined whether dopamine (DA)-dependent oxidative stress underlies alpha-Syn-mediated neurodegeneration using Drosophila primary neuronal cultures. Green fluorescent protein (GFP) was used to identify live dopaminergic neurons in primary cultures prepared on a marked photoetched coverslip, which allowed us to repeatedly access preidentified dopaminergic neurons at different time points in a non-invasive manner. This live tracking of GFP-marked dopaminergic neurons revealed age-dependent neurodegeneration mediated by a mutant human alpha-Syn (A30P). Degeneration was rescued when alpha-Syn neuronal cultures were incubated with 1 mm glutathione from Day 3 after culturing. Furthermore, depletion of cytoplasmic DA by 100 microm alpha-methyl-p-tyrosine completely rescued the early stage of alpha-Syn-mediated dopaminergic cell loss, demonstrating that DA plays a major role in oxidative stress-dependent neurodegeneration mediated by alpha-Syn. In contrast, overexpression of a Drosophila tyrosine hydroxylase gene (dTH1) alone caused DA neurodegeneration by enhanced DA synthesis in the cytoplasm. Age-dependent dopaminergic cell loss was comparable in alpha-Syn vs dTH1-overexpressed neuronal cultures, indicating that increased DA levels in the cytoplasm is a critical change downstream of mutant alpha-Syn function. Finally, overexpression of a Drosophila vesicular monoamine transporter rescued alpha-Syn-mediated neurodegeneration through enhanced sequestration of cytoplasmic DA into synaptic vesicles, further indicating that a main cause of selective neurodegeneration is alpha-Syn-induced disruption of DA homeostasis. All of these results demonstrate that elevated cytoplasmic DA is a main factor underlying the early stage of alpha-Syn-mediated neurodegeneration.
Park
Soon S
SS
Neuroscience Program, Department of Biological Sciences, Ohio University, Athens, OH 45701, USA.
Schulz
Emily M
EM
Lee
Daewoo
D
D000409:Alanine / Q000235:genetics; D000818:Animals; D030801:Animals, Genetically Modified; D002478:Cells, Cultured; D004298:Dopamine / Q000378:metabolism*; D004330:Drosophila; D004625:Embryo, Nonmammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006706:Homeostasis / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007211:Indoles; D009154:Mutation / Q000502:physiology; D009410:Nerve Degeneration / Q000139:chemically induced / Q000188:drug therapy / Q000378:metabolism*; D011392:Proline / Q000235:genetics; D017447:Receptors, Dopamine D1 / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D050493:Vesicular Monoamine Transport Proteins / Q000378:metabolism; D019805:alpha-Methyltyrosine / Q000494:pharmacology; D051844:alpha-Synuclein / Q000235:genetics / Q000378:metabolism*
D004791:Enzyme Inhibitors; D007211:Indoles; D017447:Receptors, Dopamine D1; D050493:Vesicular Monoamine Transport Proteins; D051844:alpha-Synuclein; D049452:Green Fluorescent Proteins; C007293:DAPI; D019805:alpha-Methyltyrosine; D011392:Proline; D014446:Tyrosine 3-Monooxygenase; D005978:Glutathione; D000409:Alanine; D004298:Dopamine