Fig. 1: Human α-synuclein resulted in motor learning deficits associated with cortical hyperexcitability.

a Upper, the design of adeno-associated virus (AAV)- human synapsin (hSyn)-human α-synuclein (SNCA)-mCherry viral vector. Lower, Schematic diagram of generating mouse synucleinopathy model by expressing SNCA gene into bilateral primary motor cortex (M1) of adult male mice under the help of an AAV vector carrying hSyn promoter. Three weeks later, behavioral test batteries were performed. b Representative blotting bands of total synuclein and phosphorylated synuclein in M1 extracts. c Quantification for the relative expression of total (left) and phosphorylated synuclein (right). Two-sample unpaired t-test, t(8) = 6.967, P = 0.0001 for synuclein; t(8) = 3.907, P = 0.005 for p-synuclein. N = 5 mice in each group. d Upper panels: coronal section of mouse M1 after SNCA overexpression. AAV-SNCA was mostly expressed within neurons (NeuN+). Lower panels: cortical expression of synuclein and phosphorylated synuclein were observed. Scale bar, 100 μm or 20 μm (enlarged panels in lower right). e Total distance traveled in the open field arena was unaffected after SNCA overexpression. Two-sample unpaired t-test, t(14) = 0.3807, P = 0.7091. f SNCA-overexpressing mice had remarkably impaired motor performance on the accelerating rotarod. Two-way analysis of variance (ANOVA) with respect to group factor, F(1126) = 55.81, P < 0.0001. Sidak’s multiple comparison test of post hoc comparison at each time point. g SNCA group spent longer time to accomplish the vertical climbing task. Two-way ANOVA with respect to group factor, F(142) = 4.553, P = 0.0387. Sidak’s multiple comparison test of post hoc comparison at each time point. h Worsening performance on the walking beam in mutant mice. F(139) = 13.55, P = 0.0007. Sidak’s multiple comparison test of post hoc comparison at each time point. N = 8 mice in each group in (e–h). i Patch-clamp recording traces of layer 5 pyramidal neurons (L5 PNs) showed potentiated miniature excitatory postsynaptic currents (mEPSCs) in SNCA group. j SNCA group had higher amplitude of mEPSCs. Non-parametric Mann–Whitney test U = 6, P = 0.018. k Increased frequency of mEPSCs was observed under synucleinopathy. Non-parametric Mann–Whitney test U = 6, P = 0.017. l Elevated number of action potentials (AP) in SNCA group under fixed injection currents. Two-way ANOVA with respect to group factor, F(1104) = 4.530, P = 0.0357. m Higher potentials of afterhyperpolarization (AHP). Two-sample unpaired t-test, t(13) = 3.817, P = 0.0021. n Higher rheobase values in SNCA group. Two-sample unpaired t-test, t(13) = 3.026, P = 0.0097. o Decreased resting membrane potential (RMP) in SNCA group. Two-sample unpaired t-test, t(13) = 4.181, P = 0.0011. N = 8 and 7 neurons from 3 animals in mCherry and SNCA group, respectively, in (j–o). ns, no significant difference, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. All data were presented as mean ± SEM.