Fig. 6
A53T α-synuclein over-expression in nigral dopaminergic neurons is associated with accumulation of polyubiquitinated conjugates. Adult wild-type rats received unilateral stereotaxic co-injection of AAV-UbG76V-GFP with AAV-A53T, AAV-Empty or AAV-TdTomato into the right SNpc. Animals were culled at 1 wpi. a Comparison of immunofluorescent staining with anti-tyrosine hydroxylase (TH; cyan), anti-polyubiquitinated conjugate (PolyUb; yellow), anti-α-synuclein (SYN; red) antibodies or native TdTomato fluorescence (TdTom; red) in SNpc at 1 wpi. Scale bar 500 μm (inset 50 μm). b Quantification of mean anti-polyubiquinated conjugate staining intensity in TH+ neurons of SNpc 1 week following ipsilateral AAV injection. A significant increase in levels of polyubiquitinated conjugates is observed in nigral dopaminergic neurons of animals expressing AAV-A53T α-synuclein/AAV-UbG76V-GFP, relative to AAV-Empty/AAV-UbG76V-GFP and AAV-TdTomato/AAV-UbG76V-GFP control groups (****p < 0.0001; one-way ANOVA with post-hoc Tukey tests; n = 3–5 per group). c Representative scatter plots displaying relationship between anti-synuclein and anti-polyubiquitinated conjugate staining intensities in individual TH+ neurons in SNpc 1 week after ipsilateral AAV-A53T α-synuclein/AAV-UbG76V-GFP administration. A significant positive correlation between levels of A53T α-synuclein and polyubiquitinated conjugates in nigral dopaminergic neurons is observed (Spearman correlation; individual r values as displayed for n = 4 animals; p < 0.0001)