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Nigral proteasome inhibition in mice leads to motor and non-motor deficits and increased expression of Ser129 phosphorylated α-synuclein.

Bentea E, Van der Perren A, Van Liefferinge J, El Arfani A, Albertini G, Demuyser T, Merckx E, Michotte Y, Smolders I, Baekelandt V, Massie A - Front Behav Neurosci (2015)

Bottom Line: Lactacystin, a potent inhibitor of the proteasome, was previously delivered to the nigrostriatal pathway of rodents to model nigrostriatal degeneration.Although lactacystin-treated animals develop parkinsonian motor impairment, it is currently unknown whether they also develop non-motor symptoms characteristic of this disorder.We studied the degree of neurodegeneration and the behavioral phenotype 1 and 3 weeks after lactacystin lesion both in terms of motor impairment, as well as non-motor symptoms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Biotechnology and Molecular Biology, Center for Neurosciences, Vrije Universiteit Brussel Brussels, Belgium.

ABSTRACT
Parkinson's disease is a neurodegenerative disorder characterized by motor and non-motor disturbances. Various pathogenic pathways drive disease progression including oxidative stress, mitochondrial dysfunction, α-synuclein aggregation and impairment of protein degradation systems. Dysfunction of the ubiquitin-proteasome system in the substantia nigra of Parkinson's disease patients is believed to be one of the causes of protein aggregation and cell death associated with this disorder. Lactacystin, a potent inhibitor of the proteasome, was previously delivered to the nigrostriatal pathway of rodents to model nigrostriatal degeneration. Although lactacystin-treated animals develop parkinsonian motor impairment, it is currently unknown whether they also develop non-motor symptoms characteristic of this disorder. In order to further describe the proteasome inhibition model of Parkinson's disease, we characterized the unilateral lactacystin model, performed by stereotaxic injection of the toxin in the substantia nigra of mice. We studied the degree of neurodegeneration and the behavioral phenotype 1 and 3 weeks after lactacystin lesion both in terms of motor impairment, as well as non-motor symptoms. We report that unilateral administration of 3 μg lactacystin to the substantia nigra of mice leads to partial (~40%) dopaminergic cell loss and concurrent striatal dopamine depletion, accompanied by increased expression of Ser129-phosphorylated α-synuclein. Behavioral characterization of the model revealed parkinsonian motor impairment, as well as signs of non-motor disturbances resembling early stage Parkinson's disease including sensitive and somatosensory deficits, anxiety-like behavior, and perseverative behavior. The consistent finding of good face validity, together with relevant construct validity, warrant a further evaluation of proteasome inhibition models of Parkinson's disease in pre-clinical research and validation of therapeutic targets.

No MeSH data available.


Related in: MedlinePlus

Accumulation of α-synuclein and S129-P α-synuclein in the SN of mice receiving 3 μg LAC. Immunohistochemical analyses revealed an increase in S129-P α-synuclein (A,C) and α-synuclein (B,D) immunoreactivity in the SN of LAC-treated mice. High magnification photomicrographs of the ipsilateral SNr in (A,B) demonstrate the presence of S129-P α-synuclein and α-synuclein immunoreactive fibers (C,D respectively). Data are presented as percentage increase in S129-P α-synuclein (C) or α-synuclein (D) optical density compared to the intact side (mean ± s.e.m.). ***p < 0.001, **p < 0.01 (Two-Way ANOVA), ###p < 0.001, #p < 0.05 (Bonferroni post-hoc vs. sham). Sample size indicated in the figure. LAC lactacystin, OD optical density, S129-P S129-phosphorylated, SN substantia nigra. Scale bar 400 μm (A,B), 50 μm (C,D).
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Figure 2: Accumulation of α-synuclein and S129-P α-synuclein in the SN of mice receiving 3 μg LAC. Immunohistochemical analyses revealed an increase in S129-P α-synuclein (A,C) and α-synuclein (B,D) immunoreactivity in the SN of LAC-treated mice. High magnification photomicrographs of the ipsilateral SNr in (A,B) demonstrate the presence of S129-P α-synuclein and α-synuclein immunoreactive fibers (C,D respectively). Data are presented as percentage increase in S129-P α-synuclein (C) or α-synuclein (D) optical density compared to the intact side (mean ± s.e.m.). ***p < 0.001, **p < 0.01 (Two-Way ANOVA), ###p < 0.001, #p < 0.05 (Bonferroni post-hoc vs. sham). Sample size indicated in the figure. LAC lactacystin, OD optical density, S129-P S129-phosphorylated, SN substantia nigra. Scale bar 400 μm (A,B), 50 μm (C,D).

Mentions: Immunohistochemistry revealed that intranigral administration of LAC led to an increase in α-synuclein (Figures 2B,D) and S129-P α-synuclein (Figures 2A,C) immunoreactivity at both 1 and 3 weeks post-lesion, with no signs of progression of pathology between the two time points. Densitometric analyses revealed an increase in α-synuclein immunoreactivity in the ipsilateral SN with ~50% [treatment factor: F(1, 12) = 40.20, p < 0.001], that could be observed at both 1 week (p < 0.001) and 3 weeks (p < 0.05) post-lesion (Figure 2D, Table 1). Furthermore, we could document an increase in S129-P α-synuclein immunoreactivity in the ipsilateral SN with ~150% [treatment factor: F(1, 12) = 13.23, p < 0.01], at both 1 week (p < 0.05) and 3 weeks (p < 0.05) post-surgery (Figure 2C, Table 1). The increased expression of α-synuclein and S129-P α-synuclein could be observed throughout the entire rostro-caudal extent of the SN, and did not affect neighboring regions, such as the VTA (Supplementary Figure 1). Furthermore, the increased immunoreactivity seemed to be observed mainly in the SNr, where it presented as neuropil (Figures 2C,D). Confocal fluorescent double labeling experiments also indicated increased S129-P α-synuclein expression in SNc neurons, as S129-P α-synuclein was seen to accumulate in TH+ (Figure 3B) and NeuN+ (Figure 3C) nigral neurons, but not GFAP+ astrocytes (Figure 3A). Accumulated S129-P α-synuclein showed both nuclear (Figure 3B), and cytoplasmic (Figure 3C) subcellular distribution.


Nigral proteasome inhibition in mice leads to motor and non-motor deficits and increased expression of Ser129 phosphorylated α-synuclein.

Bentea E, Van der Perren A, Van Liefferinge J, El Arfani A, Albertini G, Demuyser T, Merckx E, Michotte Y, Smolders I, Baekelandt V, Massie A - Front Behav Neurosci (2015)

Accumulation of α-synuclein and S129-P α-synuclein in the SN of mice receiving 3 μg LAC. Immunohistochemical analyses revealed an increase in S129-P α-synuclein (A,C) and α-synuclein (B,D) immunoreactivity in the SN of LAC-treated mice. High magnification photomicrographs of the ipsilateral SNr in (A,B) demonstrate the presence of S129-P α-synuclein and α-synuclein immunoreactive fibers (C,D respectively). Data are presented as percentage increase in S129-P α-synuclein (C) or α-synuclein (D) optical density compared to the intact side (mean ± s.e.m.). ***p < 0.001, **p < 0.01 (Two-Way ANOVA), ###p < 0.001, #p < 0.05 (Bonferroni post-hoc vs. sham). Sample size indicated in the figure. LAC lactacystin, OD optical density, S129-P S129-phosphorylated, SN substantia nigra. Scale bar 400 μm (A,B), 50 μm (C,D).
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Related In: Results  -  Collection

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Figure 2: Accumulation of α-synuclein and S129-P α-synuclein in the SN of mice receiving 3 μg LAC. Immunohistochemical analyses revealed an increase in S129-P α-synuclein (A,C) and α-synuclein (B,D) immunoreactivity in the SN of LAC-treated mice. High magnification photomicrographs of the ipsilateral SNr in (A,B) demonstrate the presence of S129-P α-synuclein and α-synuclein immunoreactive fibers (C,D respectively). Data are presented as percentage increase in S129-P α-synuclein (C) or α-synuclein (D) optical density compared to the intact side (mean ± s.e.m.). ***p < 0.001, **p < 0.01 (Two-Way ANOVA), ###p < 0.001, #p < 0.05 (Bonferroni post-hoc vs. sham). Sample size indicated in the figure. LAC lactacystin, OD optical density, S129-P S129-phosphorylated, SN substantia nigra. Scale bar 400 μm (A,B), 50 μm (C,D).
Mentions: Immunohistochemistry revealed that intranigral administration of LAC led to an increase in α-synuclein (Figures 2B,D) and S129-P α-synuclein (Figures 2A,C) immunoreactivity at both 1 and 3 weeks post-lesion, with no signs of progression of pathology between the two time points. Densitometric analyses revealed an increase in α-synuclein immunoreactivity in the ipsilateral SN with ~50% [treatment factor: F(1, 12) = 40.20, p < 0.001], that could be observed at both 1 week (p < 0.001) and 3 weeks (p < 0.05) post-lesion (Figure 2D, Table 1). Furthermore, we could document an increase in S129-P α-synuclein immunoreactivity in the ipsilateral SN with ~150% [treatment factor: F(1, 12) = 13.23, p < 0.01], at both 1 week (p < 0.05) and 3 weeks (p < 0.05) post-surgery (Figure 2C, Table 1). The increased expression of α-synuclein and S129-P α-synuclein could be observed throughout the entire rostro-caudal extent of the SN, and did not affect neighboring regions, such as the VTA (Supplementary Figure 1). Furthermore, the increased immunoreactivity seemed to be observed mainly in the SNr, where it presented as neuropil (Figures 2C,D). Confocal fluorescent double labeling experiments also indicated increased S129-P α-synuclein expression in SNc neurons, as S129-P α-synuclein was seen to accumulate in TH+ (Figure 3B) and NeuN+ (Figure 3C) nigral neurons, but not GFAP+ astrocytes (Figure 3A). Accumulated S129-P α-synuclein showed both nuclear (Figure 3B), and cytoplasmic (Figure 3C) subcellular distribution.

Bottom Line: Lactacystin, a potent inhibitor of the proteasome, was previously delivered to the nigrostriatal pathway of rodents to model nigrostriatal degeneration.Although lactacystin-treated animals develop parkinsonian motor impairment, it is currently unknown whether they also develop non-motor symptoms characteristic of this disorder.We studied the degree of neurodegeneration and the behavioral phenotype 1 and 3 weeks after lactacystin lesion both in terms of motor impairment, as well as non-motor symptoms.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Biotechnology and Molecular Biology, Center for Neurosciences, Vrije Universiteit Brussel Brussels, Belgium.

ABSTRACT
Parkinson's disease is a neurodegenerative disorder characterized by motor and non-motor disturbances. Various pathogenic pathways drive disease progression including oxidative stress, mitochondrial dysfunction, α-synuclein aggregation and impairment of protein degradation systems. Dysfunction of the ubiquitin-proteasome system in the substantia nigra of Parkinson's disease patients is believed to be one of the causes of protein aggregation and cell death associated with this disorder. Lactacystin, a potent inhibitor of the proteasome, was previously delivered to the nigrostriatal pathway of rodents to model nigrostriatal degeneration. Although lactacystin-treated animals develop parkinsonian motor impairment, it is currently unknown whether they also develop non-motor symptoms characteristic of this disorder. In order to further describe the proteasome inhibition model of Parkinson's disease, we characterized the unilateral lactacystin model, performed by stereotaxic injection of the toxin in the substantia nigra of mice. We studied the degree of neurodegeneration and the behavioral phenotype 1 and 3 weeks after lactacystin lesion both in terms of motor impairment, as well as non-motor symptoms. We report that unilateral administration of 3 μg lactacystin to the substantia nigra of mice leads to partial (~40%) dopaminergic cell loss and concurrent striatal dopamine depletion, accompanied by increased expression of Ser129-phosphorylated α-synuclein. Behavioral characterization of the model revealed parkinsonian motor impairment, as well as signs of non-motor disturbances resembling early stage Parkinson's disease including sensitive and somatosensory deficits, anxiety-like behavior, and perseverative behavior. The consistent finding of good face validity, together with relevant construct validity, warrant a further evaluation of proteasome inhibition models of Parkinson's disease in pre-clinical research and validation of therapeutic targets.

No MeSH data available.


Related in: MedlinePlus