Alpha-synuclein deficient mice are resistant to toxin-induced multiple system atrophy

Abstract

Multiple systems atrophy (MSA) is a neurodegenerative disorder characterized by oligodendrocytic accumulations of alpha-synuclein (alphasyn). Oxidative stress is a key mechanism proposed to underlie MSA pathology. To address the role of alphasyn modifications, over and above general oxidative modifications, this study examined the effects of 3-nitropropionic acid (3NP) administration, a technique used to model MSA, in knock-out mice lacking alphasyn (alphasynKO). Although susceptible to 3NP-induced oxidative stress, alphasynKO mice display reduced neuronal loss and dendritic pathology. The alphasynKO mice are resistant to 3NP-induced motor deficits and display attenuated loss of tyrosine hydroxylase and dopamine transporter striatal immunoreactivity. The results suggest that deficits in MSA are not due to general oxidative protein modification but in addition may be related to specific alphasyn modifications.

Fig. 1

Analysis of oxidative stress, alpha-synuclein (α-syn) levels and behavioral deficits upon 3-nitropropionic acid (3NP) administration. Oxyblot analysis of global protein oxidation levels in non-transgenic littermates (NTg), knock-out mice lacking αsyn (αsynKO), and myelin basic protein (MBP)-hαsyn Tg mice (a), analyzed in (b). Assessment of mitochondrial complex II (Mitc Cmplx II) activity levels after 3NP administration in the NTg, αsynKO, and MBP-hαsyn Tg mice (c). Immunoblot analysis of total αsyn levels, using two different antibodies and nitrotyrosine levels 3NP administration in the NTg, αsynKO, and MBP-hαsyn Tg mice (d), quantitative analysis (e–g). Motor behavior was assessed using the pole test (h). Mice were placed at the top of a vertical pole and total time to descend (T-total) was measured. *Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between saline and 3NP-treated members of the same genotype (saline-treated NTg vs. 3NP-treated NTg mice). **Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between mice of different genotype (saline-treated NTg vs. saline-treated αsynKO mice). Arb., arbitrary.

Fig. 2

Immunohistochemical analysis of alpha-synuclein (α-syn) and dopamine markers in non-transgenic littermates (NTg), knock-out mice lacking αsyn (αsynKO), and myelin basic protein (MBP)-hαsyn Tg mice after 3-nitropropionic acid (3NP) administration. Immunohistochemical analysis was performed to examine the number of αsyn immunoreactive cells in the striatum of vehicle-treated and 3NP-treated NTg mice (a and b), αsynKO mice (c and d) and MBP-hαsyn mice (e and f), analyzed in (g). To assess the effect of 3NP administration on dopamine markers immunohistochemical analysis was performed to examine the number of tyrosine hydroxylase (TH) and dopamine transporter (DAT) immunoreactive cells in the striatum of vehicle-treated and 3NP-treated NTg mice (h, i, o, and p, respectively), αsynKO mice (j, k, q, and r, respectively) and MBP-hαsyn mice (l, m, s, and t, respectively), analyzed in (n) and (u). Scale bar = 50 μM. *Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between saline and 3NP-treated members of the same genotype (saline-treated NTg vs. 3NP-treated NTg mice). **Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between mice of different genotype (saline-treated NTg vs. saline-treated αsynKO mice).

Fig. 3

The knock-out mice lacking αsyn (αsynKO) display attenuated neuropathological deficits after 3-nitropropionic acid (3NP) administration. Immunohistochemical analysis of neuronal marker (NeuN) positive cells in the striatum was conducted in order to investigate the effects of 3NP administration on neuronal density in saline-treated and 3NP treated non-transgenic littermates (NTg) mice (a and b), αsynKO mice (c and d) and myelin basic protein (MBP)-hαsyn Tg mice (e and f), analyzed (g). Dendritic pathology was assessed by the percentage of microtubule-associated protein-2 (MAP2) immunoreactive area of neutrophil in the striatum of saline-treated and 3NP treated NTg mice (h and i), αsynKO mice (j and k) and MBP-hαsyn Tg mice (l and m), analyzed in (n). Astrogliosis after 3NP administration was assessed through glial fibrillary acidic protein (GFAP) immunoreactivity in the striatum of saline-treated and 3NP treated NTg mice (o and p), αsynKO mice (q and r), and MBP-hαsyn Tg mice (s and t), analyzed (u). Scale bar (a–f and o–t) = 50 μM, scale bar (h–m) = 20 μM. *Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between saline and 3NP-treated members of the same genotype (saline-treated NTg vs. 3NP-treated NTg mice). **Significant difference (P < 0.05, one-way analysis of variance and post hoc Fisher's test) between mice of different genotype (saline-treated NTg vs. saline-treated αsynKO mice). Arb., arbitrary.