Motor deficits and brain pathology in the Parkinson's disease mouse model hA53Ttg

Abstract

Background: Parkinson's disease (PD) is a debilitating neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons and the accumulation of α-synuclein (α-syn) aggregates. The A53T missense point mutation occurs in autosomal dominant familial PD and has been found to promote the aggregation of α-syn. To investigate the role of the A53T mutation in PD, researchers have developed various mouse models with this mutation.

Objective: We therefore conducted a comprehensive characterization of the tg(THY1-SNCA*A53T)M53Sud mouse model (hA53Ttg mice) for its motor and pathological features.

Methods: hA53Ttg mice were tested for motor impairments in a series of motor tests at 2, 4 or 6 months of age. Human α-syn and α-syn pSer129, as well as GFAP and Iba1 signal were labeled and quantified in the cortex, hippocampus, and brainstem. Neurofilament light chain (NF-L) levels were measured in the cerebrospinal fluid (CSF) and plasma. Ex vivo analyses were performed at the age of 2, 4, 6, and 10 months.

Results: Behavioral tests revealed early muscle weakness and motor impairments that progressed with age. Immunohistochemical analyses demonstrated elevated levels of human α-syn and α-syn pSer129 in all evaluated brain regions. α-syn pSer129 labeling further revealed fiber-like structures in the cortex of older animals. Neuroinflammation was observed in an age-dependent manner. Biochemical evaluation revealed elevated NF-L levels in the plasma and CSF. Overall, our findings highlight the value of hA53Ttg mice in modeling PD-associated pathologies that closely resemble those observed in PD patients.

Conclusion: Our results thus suggest that hA53Ttg mice are a useful tool for studying the underlying mechanisms of PD.

Keywords: Parkinson’s disease; motor skills; neurodegeneration; neuroinflammation; transgenic mouse; α-synuclein.

Figure 1

Decreased muscle strength and motor coordination in hA53Ttg mice. The wire hanging test (A) shows a reduced latency to fall while the beam walk test shows an increased number of slips (B) and an increased time (C) needed to traverse the 13 mm square beam in hA53Ttg animals in comparison to ntg controls; n = 23–24 per group. Mean ± SEM; Two-way ANOVA followed by Bonferroni’s post hoc test; #p < 0.05, ##p < 0.01, ***/###p < 0.001. *Differences between genotypes; #differences between age groups.

Figure 2

Performers and non-performers in the beam walk test. Number of animals excluded from the beam walk test evaluation due to a fall from the beam (A). Rows display the number of excluded animals for each trial while columns display the number of excluded animals per group. The bottom row contains the total number of animals tested per group. Analysis of performance showed that less hA53Ttg animals were able to traverse the beam at any age compared to ntg mice (B). Number of animals in percent; n = 23–24 per group. Fisher’s exact test; ***p < 0.001.

Figure 3

Motor coordination and orofacial muscle impairment in hA53Ttg mice. Analysis of animals showed a reduced latency to fall in the RotaRod test (A) and a reduced number of bites per episode in the pasta gnawing test (B) starting at 4 months of age in hA53Ttg animals compared to ntg mice; n = 23–24 per group. Mean ± SEM; two-way ANOVA followed by Bonferroni’s post hoc test; **/##p < 0.01, ***/###p < 0.001. *Differences between genotypes; #differences between age groups.

Figure 4

Human and phosphorylated α-synuclein in different brain regions. (A) Labeling of α-syn pSer129 (green) and hα-syn (red) in the cortex of hA53Ttg mice at the age of 4 and 10 months; cell nuclei were labeled with DAPI (blue). Mice at 4 months display α-syn pSer129 labeling which is restricted to nuclear and somatic location in the cortex, whereas the nuclear/somatic pSer129 α-syn labeling becomes weaker in 10 months old mice. However, note the appearance of high intensity signal which resembles a more fiber-like pattern at this age. Single channel images were taken at the position indicated by the rectangle in the left overview image. Increased immunoreactive (IR) area of human α-synuclein (hα-syn) in the cortex (B), hippocampus (C) and brainstem (D) in hA53Ttg animals already at 2 months of age compared to ntg littermates. IR area of α-synuclein phosphorylated at serine 129 (α-syn pSer129) in the cortex (E), hippocampus (F) and brainstem (G); n = 7–9 per group. Mean ± SEM; two-way ANOVA followed by Bonferroni’s post hoc test; */#p < 0.05, **/##p > 0.01, ***/###p < 0.001. *Differences between genotypes; #differences between age groups.

Figure 5

Neuroinflammation in the brainstem. (A) Labeling of pSer129 α-syn (green), GFAP (red), and Iba1 (white) shows striking differences between individual animals of the same age and genotype; cell nuclei were labelled with DAPI (blue). “Weak pathology” mice display little labeling for all three markers, whereas a large amount of pSer129 α-syn and associated gliosis is evident in the brainstem of “strong pathology” mice. Images thus support a high within-group variation of hA53Ttg mice at the age of 10 months. Single channel images were taken at the position indicated by the rectangle in the left overview image. Arrows point towards areas with strong pathology in the cortex. Increased immunoreactive (IR) area of ionized calcium binding adaptor molecule 1 (Iba1; B) and glial fibrillary acidic protein (GFAP; C) in the brainstem of 10-months-old hA53Ttg animals compared to ntg littermates; n = 7–9 per group. Mean ± SEM; two-way ANOVA followed by Bonferroni’s post hoc test; ***/###p < 0.001. *Differences between genotypes; #differences between age groups.

Figure 6

Quantification of NF-L levels in the plasma of hA53Ttg mice and correlation with α-syn pSer129. (A) Neurofilament-light chain (NF-L) levels in pg/mL in the plasma of hA53Ttg and ntg littermates at 2–10 months of age; n = 8 per group. Mean ± SEM; two-way ANOVA followed by Bonferroni’s post hoc test; ***/###p < 0.001. (B) High correlation between plasma NF-L levels and immunoreactive (IR) area of α-syn pSer129 in the brainstem of 10-months-old hA53Ttg mice. *Differences between genotypes; #differences between age groups.