Alpha-synuclein and its neurotoxic fragment inhibit dopamine uptake into rat striatal synaptosomes. Relationship to nitric oxide.
Alpha-synuclein (ASN), a 140-amino acid protein, is richly expressed in presynaptic terminals in the central nervous system, where it plays a role in synaptic vesicle function. However, if it is altered and accumulated it is involved in neurodegeneration as Parkinson's disease (PD). ASN contained 35-amino acid domain known as non-amyloid beta component of Alzheimer's disease amyloid (NAC) that is probably responsible for its aggregation and toxicity. Up till now the role of ASN in dopaminergic system function and in pathogenesis of PD is unknown. The aim of this study was to determine the effect of brain aging and the role of ASN and NAC peptide on striatal dopamine transporter (DAT) function. The study was carried out using radiochemical and spectrofluorimetrical determination. It was found that DAT activity assessed by measuring [3H]-dopamine (DA) uptake into striatal synaptosomes significantly decreased in 24-month-old rats comparing to 4-month-old. ASN and NAC peptide at 10 microM concentration inhibited DAT activity by 30%. Both molecules evoked intrasynaptosomal generation of reactive oxygen species measured by fluorogenic probe, 2'7'-dichlorofluorescin diacetate. In addition, ASN activated striatal cytosolic nitric oxide synthase (NOS) by 20%. Nitric oxide (NO) donor, sodium nitroprusside (SNP) (10 microM) and oxidative stress evoked by FeCl2 (25 microM) reduced [3H]DA uptake by 28 and 41%, respectively. Potent antioxidants: Trolox and 4-hydroxy-Tempo had no effect on DAT function but NOS inhibitor Nomega-nitro-L-arginine (100 microM), prevented ASN-evoked DAT down-regulation. These data indicated an important role of ASN in alteration of DA synaptic homeostasis, probably by NO mediated DAT alteration.
Adamczyk
Agata
A
Department of Cellular Signalling, Medical Research Centre, Polish Academy of Sciences, 5 Pawińskiego St., 02-106 Warsaw, Poland. agatazambrzycka@hotmail.com
Kaźmierczak
Anna
A
Strosznajder
Joanna B
JB
D000375:Aging / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004298:Dopamine / Q000378:metabolism*; D018765:Dopamine Uptake Inhibitors; D066298:In Vitro Techniques; D015227:Lipid Peroxidation / Q000187:drug effects; D008297:Male; D017072:Neostriatum / Q000187:drug effects / Q000378:metabolism*; D009498:Neurotoxins / Q000494:pharmacology*; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D052248:Nitric Oxide Synthase Type I / Q000378:metabolism; D009599:Nitroprusside / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D010446:Peptide Fragments / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*; D051844:alpha-Synuclein / Q000494:pharmacology*
D018765:Dopamine Uptake Inhibitors; D009498:Neurotoxins; D020030:Nitric Oxide Donors; D010446:Peptide Fragments; D051844:alpha-Synuclein; D009599:Nitroprusside; D009569:Nitric Oxide; D052248:Nitric Oxide Synthase Type I; D004298:Dopamine