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Formation of α-synuclein Lewy neurite-like aggregates in axons impedes the transport of distinct endosomes.

Volpicelli-Daley LA, Gamble KL, Schultheiss CE, Riddle DM, West AB, Lee VM - Mol. Biol. Cell (2014)

Bottom Line: Ultrastructural analyses and live imaging demonstrate that α-syn accumulations do not cause a generalized defect in axonal transport; the inclusions do not fill the axonal cytoplasm, disrupt the microtubule cytoskeleton, or affect the transport of synaptophysin or mitochondria.In addition, the TrkB receptor-associated signaling molecule pERK5 accumulates in α-syn aggregate-bearing neurons.These early effects of α-syn accumulations may predict points of intervention in the neurodegenerative process.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Behavioral Neurobiology, University of Alabama, Birmingham, Birmingham, AL 35294 Department of Pathology and Laboratory Medicine, Institute on Aging, and Center for Neurodegenerative Disease Research, University of Pennsylvania School of Medicine, Philadelphia, PA, 19104 volpicel@uab.edu.

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Normal transport of synaptophysin-GFP in neurons with α-syn aggregates. Primary hippocampal neurons were transfected with synaptophysin-GFP, treated with PBS or PFFs, and imaged 7 d later. Synaptophysin-GFP, number of particles analyzed, 2044 for PBS and 1944 for PFF (19 axons, PBS; 17 axons, PFF). (A) Top, images from movies captured every 1 s for 3 min; scale bar, 10 μm. Kymographs shown below the images were generated as visual representations of distance traveled over time. (B) Of the mobile particles, the percentages of anterograde and retrograde particles were also quantified. There were no significant differences between the PBS- and PFF-treated groups. (C) There were no significant differences in the mean number of synaptophysin-GFP particles per 50 μm of axonal membrane. In addition, there were no significant differences in the number of pauses (D) or reversals (E) between the two groups. A Poisson regression on velocities binned with 10 cut points was not statistically significant between PBS and PFF groups for anterograde synaptophysin-GFP velocities (Wald χ2 = 1.420, p = NS; F) or for retrograde synaptophysin-GFP velocities (Wald χ2 = 3.246, p = NS; G). (F, G) Right, median and interquartile ranges of the velocities of the mobile synaptophysin-GFP particles. The Mann–Whitney test did not produce significant differences for anterograde or retrograde velocities.
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Figure 3: Normal transport of synaptophysin-GFP in neurons with α-syn aggregates. Primary hippocampal neurons were transfected with synaptophysin-GFP, treated with PBS or PFFs, and imaged 7 d later. Synaptophysin-GFP, number of particles analyzed, 2044 for PBS and 1944 for PFF (19 axons, PBS; 17 axons, PFF). (A) Top, images from movies captured every 1 s for 3 min; scale bar, 10 μm. Kymographs shown below the images were generated as visual representations of distance traveled over time. (B) Of the mobile particles, the percentages of anterograde and retrograde particles were also quantified. There were no significant differences between the PBS- and PFF-treated groups. (C) There were no significant differences in the mean number of synaptophysin-GFP particles per 50 μm of axonal membrane. In addition, there were no significant differences in the number of pauses (D) or reversals (E) between the two groups. A Poisson regression on velocities binned with 10 cut points was not statistically significant between PBS and PFF groups for anterograde synaptophysin-GFP velocities (Wald χ2 = 1.420, p = NS; F) or for retrograde synaptophysin-GFP velocities (Wald χ2 = 3.246, p = NS; G). (F, G) Right, median and interquartile ranges of the velocities of the mobile synaptophysin-GFP particles. The Mann–Whitney test did not produce significant differences for anterograde or retrograde velocities.

Mentions: Synaptophysin-GFP traversed axons in punctate carriers (Figure 3A). The mobility of these particles appeared similar in control neurons that did not harbor α-syn pathology and PFF-treated neurons bearing axonal LN-like aggregates (PFF-treated; Supplemental Movie S2). There were no significant differences in the percentage of mobile particles between control neurons and neurons harboring α-syn inclusions (Figure 3B). In addition, there were no significant differences in the abundance of particles per 50 μm of axonal membrane (Figure 3C) or in the number of particle pauses or reversals (Figure 3, D and E). Because the velocities from the PBS- and PFF-treated groups did not fit a normal distribution, a Poisson regression analysis was performed on binned synaptophysin-GFP velocities. This analysis revealed no statistically significant differences between PBS and PFF groups for anterograde or retrograde transport (Figure 3, F and G). Plotting the velocities of mobile vesicles as the median and interquartile range also revealed no differences in the velocities of synaptophysin-GFP in control and α-syn aggregate–bearing axons.


Formation of α-synuclein Lewy neurite-like aggregates in axons impedes the transport of distinct endosomes.

Volpicelli-Daley LA, Gamble KL, Schultheiss CE, Riddle DM, West AB, Lee VM - Mol. Biol. Cell (2014)

Normal transport of synaptophysin-GFP in neurons with α-syn aggregates. Primary hippocampal neurons were transfected with synaptophysin-GFP, treated with PBS or PFFs, and imaged 7 d later. Synaptophysin-GFP, number of particles analyzed, 2044 for PBS and 1944 for PFF (19 axons, PBS; 17 axons, PFF). (A) Top, images from movies captured every 1 s for 3 min; scale bar, 10 μm. Kymographs shown below the images were generated as visual representations of distance traveled over time. (B) Of the mobile particles, the percentages of anterograde and retrograde particles were also quantified. There were no significant differences between the PBS- and PFF-treated groups. (C) There were no significant differences in the mean number of synaptophysin-GFP particles per 50 μm of axonal membrane. In addition, there were no significant differences in the number of pauses (D) or reversals (E) between the two groups. A Poisson regression on velocities binned with 10 cut points was not statistically significant between PBS and PFF groups for anterograde synaptophysin-GFP velocities (Wald χ2 = 1.420, p = NS; F) or for retrograde synaptophysin-GFP velocities (Wald χ2 = 3.246, p = NS; G). (F, G) Right, median and interquartile ranges of the velocities of the mobile synaptophysin-GFP particles. The Mann–Whitney test did not produce significant differences for anterograde or retrograde velocities.
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Figure 3: Normal transport of synaptophysin-GFP in neurons with α-syn aggregates. Primary hippocampal neurons were transfected with synaptophysin-GFP, treated with PBS or PFFs, and imaged 7 d later. Synaptophysin-GFP, number of particles analyzed, 2044 for PBS and 1944 for PFF (19 axons, PBS; 17 axons, PFF). (A) Top, images from movies captured every 1 s for 3 min; scale bar, 10 μm. Kymographs shown below the images were generated as visual representations of distance traveled over time. (B) Of the mobile particles, the percentages of anterograde and retrograde particles were also quantified. There were no significant differences between the PBS- and PFF-treated groups. (C) There were no significant differences in the mean number of synaptophysin-GFP particles per 50 μm of axonal membrane. In addition, there were no significant differences in the number of pauses (D) or reversals (E) between the two groups. A Poisson regression on velocities binned with 10 cut points was not statistically significant between PBS and PFF groups for anterograde synaptophysin-GFP velocities (Wald χ2 = 1.420, p = NS; F) or for retrograde synaptophysin-GFP velocities (Wald χ2 = 3.246, p = NS; G). (F, G) Right, median and interquartile ranges of the velocities of the mobile synaptophysin-GFP particles. The Mann–Whitney test did not produce significant differences for anterograde or retrograde velocities.
Mentions: Synaptophysin-GFP traversed axons in punctate carriers (Figure 3A). The mobility of these particles appeared similar in control neurons that did not harbor α-syn pathology and PFF-treated neurons bearing axonal LN-like aggregates (PFF-treated; Supplemental Movie S2). There were no significant differences in the percentage of mobile particles between control neurons and neurons harboring α-syn inclusions (Figure 3B). In addition, there were no significant differences in the abundance of particles per 50 μm of axonal membrane (Figure 3C) or in the number of particle pauses or reversals (Figure 3, D and E). Because the velocities from the PBS- and PFF-treated groups did not fit a normal distribution, a Poisson regression analysis was performed on binned synaptophysin-GFP velocities. This analysis revealed no statistically significant differences between PBS and PFF groups for anterograde or retrograde transport (Figure 3, F and G). Plotting the velocities of mobile vesicles as the median and interquartile range also revealed no differences in the velocities of synaptophysin-GFP in control and α-syn aggregate–bearing axons.

Bottom Line: Ultrastructural analyses and live imaging demonstrate that α-syn accumulations do not cause a generalized defect in axonal transport; the inclusions do not fill the axonal cytoplasm, disrupt the microtubule cytoskeleton, or affect the transport of synaptophysin or mitochondria.In addition, the TrkB receptor-associated signaling molecule pERK5 accumulates in α-syn aggregate-bearing neurons.These early effects of α-syn accumulations may predict points of intervention in the neurodegenerative process.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Behavioral Neurobiology, University of Alabama, Birmingham, Birmingham, AL 35294 Department of Pathology and Laboratory Medicine, Institute on Aging, and Center for Neurodegenerative Disease Research, University of Pennsylvania School of Medicine, Philadelphia, PA, 19104 volpicel@uab.edu.

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Related in: MedlinePlus