Fig. 4: cPSAP^DAT mice are more vulnerable to AAV-α-synuclein-induced toxicity, while AAV-PSAP intranigral injection counteracts the toxicity by reducing p-Ser129 α-syn levels.

A Schematic representation of unilateral stereotaxic surgery (upper left) and timeline of experiments. B Quantification of apomorphine-induced net contralateral rotation (contralateral-ipsilateral) of AAV-α-syn injected mice with or without AAV-PSAP at 6w (gray), 10w (orange), and 14w (black). Each dot depicts one mouse. N = 8, 7, 8, 6, mice in four groups, respectively. Repeated measures (RM) two-way ANOVA with Bonferroni’s post hoc test was applied; *compared to 6w, ^# interactions. C Representative images of TH (orange), PSAP (magenta), and α-syn (cyan) immunofluorescent staining on postmortem substantia nigra sections of AAV-α-syn-injected mice with or without AAV-PSAP. Top panel, low-magnification (scale bar=1000 μm) images of the whole substantia nigra. Bottom panels, high-magnification (scale bar=50 μm) images of TH neurons. D Quantification of mean fluorescence intensity (MFI) of PSAP staining in TH-positive neurons of AAV-α-syn-injected WT and cPSAP^DAT mice. Black and red circles depict contralateral and ipsilateral PSAP immunoreactivity, respectively. N_WT = 8, N_cPSAP^DAT = 7. RM two-way ANOVA with Bonferroni’s post hoc test; *compared to contralateral, ^# compared to cPSAP^DAT. E Representative images of α-syn (red dots indicate the injection side) and TH immunohistochemical staining in striatal and substantia nigra sections, and DAT staining in striatal sections of AAV-α-syn-injected mice. Scale bar, 1 mm. F Densitometry analysis of ipsilateral TH immunoreactivity in the striatum and SNc, and ipsilateral DAT immunoreactivity in the striatum of all groups of mice. Values are normalized to the mean of their corresponding contralateral immunoreactivity. Each circle represents one mouse. N = 8, 8, 7, 7, 6, 6, 6, 6 mice in eightgroups, respectively. Two-way ANOVA with Fisher’s LSD post hoc test. G Representative images of p-Ser129 α-syn immunohistochemical staining (enhanced by nickel) in striatal sections of AAV-α-syn-injected mice. Scale bar, 25 μm. H Quantification of number of p-Ser129 α-syn accumulations (area ≥ 0.64 μm²) in striatal sections of AAV-α-syn-injected mice; N = 8, 8, 7, 7mice in four groups, respectively; One-way ANOVA with Bonferroni’s post hoc test. Data are presented as mean ± S.E.M. *^/#p < 0.05, **p < 0.01, ***p < 0.001, ****^/####p < 0.0001.