Article

α-Synuclein mediates alterations in membrane conductance: A potential role for α-synuclein oligomers in cell vulnerability

Department of Neuroscience, Georgetown University Medical Center, Washington, DC 20057, USA.
European Journal of Neuroscience (Impact Factor: 3.67). 07/2010; 32(1):10-7. DOI: 10.1111/j.1460-9568.2010.07266.x
Source: PubMed

ABSTRACT alpha-Synuclein has been linked to the pathogenesis of Parkinson's disease and other synucleinopathies through its propensity to form toxic oligomers. The exact mechanism for oligomeric synuclein-directed cell vulnerability has not been fully elucidated, but one hypothesis portends the formation of synuclein-containing pores within cell membranes leading to leak channel-mediated calcium influx and subsequent cell death. Here we demonstrate synuclein-induced formation of sodium dodecyl sulfate-stable oligomers, intracellular synuclein-positive aggregates, alterations in membrane conductance reminiscent of leak channels and subsequent cytotoxicity in a dopaminergic-like cell line. Furthermore we demonstrate that the synuclein-induced membrane conductance changes are blocked by direct extracellular application of an anti-synuclein antibody. The work presented here confirms that synuclein overexpression leads to membrane conductance changes and demonstrates for the first time through antibody-blocking studies that synuclein plays a direct role in the formation of leak channels.

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    • "the presence of extracellular a-syn, suggesting that the neurotoxic effects of a-syn might also occur in the extracellular space through unknown mechanisms (El-Agnaf et al. 2003, 2006; Paleologou et al. 2009; Tokuda et al. 2010; Bidinosti et al. 2012). Previous studies have shown that a-syn alters membrane permeability through the formation of ion permeable pores (Volles and Lansbury 2002, 2003; Quist et al. 2005; Tsigelny et al. 2007, 2012; Zakharov et al. 2007; Kostka et al. 2008; Kim et al. 2009; Feng et al. 2010; van Rooijen et al. 2010; Schmidt et al. 2012). Although this is an attractive idea to explain part of the associated neurotoxicity of a-syn, it has never been demonstrated to occur in neuronal membranes exposed directly to extracellular a-syn oligomers (Tsigelny et al. 2012; Lashuel et al. 2013). "
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    ABSTRACT: It has been postulated that the accumulation of extracellular α-synuclein (α-syn) might alter the neuronal membrane by formation of "pore-like structures" that will lead to alterations in ionic homeostasis. However, this has never been demonstrated to occur in brain neuronal plasma membranes. In this study, we show that α-syn oligomers rapidly associate to hippocampal membranes in a punctate fashion, resulting in increased membrane conductance (5 fold over control) and the influx of both calcium and a fluorescent glucose analogue. The enhancement in intracellular calcium (1.7 fold over control) caused a large increase in the frequency of synaptic transmission (2.5 fold over control), calcium transients (3 fold over control) and synaptic vesicle release. Both primary hippocampal and dissociated nigral neurons showed rapid increases in membrane conductance by α-syn oligomers. In addition, we show here that α-syn caused synaptotoxic failure associated to a decrease in SV2, a membrane protein of synaptic vesicles associated to neurotransmitter release. In conclusion, extracellular α-syn oligomers facilitates the perforation of the neuronal plasma membrane, thus explaining, in part, the synaptotoxicity observed in neurodegenerative diseases characterized by its extracellular accumulation. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Journal of Neurochemistry 02/2015; 132(6). DOI:10.1111/jnc.13060 · 4.24 Impact Factor
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    • "Considering the ability of αsyn to bind to membranes characterized by different lipid compositions [57], a " scaffolding effect " may be envisioned, partially due to the localization of αsyn oligomers at the numerous membranous structures present in the cell cytoplasm and at the plasma membrane. This interpretation is supported by a model for αsyn aggregation where the early αsyn oligomers are formed not only in the cell cytoplasm but also at the membranes [32] [58], which can lead to calcium leakage [59] and to synaptic vesicles homeostasis disruption in neurons [60]. Another explanation for this pattern is the sequestration of the toxic oligomeric species by lysosomes. "
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    ABSTRACT: Background Alpha-synuclein oligomerization is associated to Parkinson's disease etiopathogenesis. The study of alpha-synuclein oligomerization properties in live cell and the definition of their effects on cellular viability are among fields expected to provide the knowledge required to unravel the mechanism(s) of toxicity that lead to the disease. Methods We used Number and Brightness method, which is a method based on fluorescence fluctuation analysis, to monitor alpha-synuclein tagged with EGFP aggregation in living SH-SY5Y cells. The presence of alpha-synuclein oligomers detected with this method was associated with intracellular structure conditions, evaluated by fluorescence confocal imaging. Results Cells overexpressing alpha-synuclein-EGFP present a heterogeneous ensemble of oligomers constituted by less than 10 monomers, when the protein approaches a threshold concentration value of about 90 nM in the cell cytoplasm. We show that the oligomeric species are partially sequestered by lysosomes and that the mitochondria morphology is altered in cells presenting oligomers, suggesting that these mitochondria may be dysfunctional. Conclusions We showed that alpha-synuclein overexpression in SH-SY5Y causes the formation of alpha-synuclein oligomeric species, whose presence is associated with mitochondrial fragmentation and autophagic-lysosomal pathway activation in live cells. General significance The unique capability provided by the Number and Brightness analysis to study alpha-synuclein oligomers distribution and properties, and the study their association to intracellular components in single live cells is important to forward our understanding of the molecular mechanisms Parkinson’s disease and it may be of general significance when applied to the study of other aggregating proteins in cellular models.
    Biochimica et Biophysica Acta (BBA) - General Subjects 06/2014; 1840(6). DOI:10.1016/j.bbagen.2014.02.013 · 3.83 Impact Factor
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    • "ls when present in protofibrils and / or oligomers , fibrils and large aggregates forms . Recent studies have determined the presence of oligomeric Syn , Syn - immunopositive aggregates , localized Syn , and alterations in membrane conductance which imply that leak channels may result in subsequent cytotoxicity in a dopaminergic - like cell line ( Feng et al . , 2010 ) . Furthermore , it can impair the viability of adjacent cells and ultimately trigger neuronal death . This dynamic rela - tionship is an important component of PD pathogenesis . Autophagy also participates in the turnover of mitochondria , whose dysfunction represents an important pathogenic mechanism of cell death in PD . The product"
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    ABSTRACT: Neurodegenerative diseases, such as Alzheimer's disease Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis, share a common cellular and molecular pathogenetic mechanism involving aberrant misfolded protein or peptide aggregation and deposition. Autophagy represents a major route for degradation of aggregated cellular proteins and dysfunctional organelles. Emerging studies have demonstrated that up-regulation of autophagy can lead to decreased levels of these toxic aggregate-prone proteins, and is beneficial in the context of aging and various models of neurodegenerative diseases. Understanding the signaling pathways involved in the regulation of autophagy is crucial to the development of strategies for therapy. This review will discuss the cellular and molecular mechanisms of autophagy and its important role in the pathogenesis of aging and neurodegenerative diseases, and the ongoing drug discovery strategies for therapeutic modulation.
    Neurobiology of aging 11/2013; 35(5). DOI:10.1016/j.neurobiolaging.2013.11.019 · 4.85 Impact Factor
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