Fig. 1: Overexpression of aSyn is induced at hippocampal neurons by infection with haSyn lentivirus.

A–C Primary hippocampal continental cultures from rat were infected with lentivirus mediating the expression of either eGFP alone or haSyn and eGFP. A Representative immunoblots of total aSyn, and eGFP levels. B 2.2-fold increase in total aSyn levels in haSyn-infected neurons (n = 6 independent cultures). β-actin was used as a loading control. C Representative images of continental hippocampal neurons infected with eGFP or haSyn lentivirus and immunolabelled using anti-MAP2 and corresponding 3D-tracing. Scale bar = 100 μm. D Cell death assessed by ToxiLight assay in rat primary neurons transduced with lentiviral encoding for haSyn for up to DIV21 (n = 7 independent cultures). haSyn transduction alter the total dendritic length (E), apical dendritic length (F) and basal dendritic length (G) at DIV 21 (n = 17–23/3 neurons/independent cultures). H Sholl analysis of aSyn-overexpressed neurons showed reduced number of intersections (n = 17–23/3 neurons/independent cultures). I–P Representative images of immunocytochemistry of continental hippocampal neurons infected with eGFP or haSyn lentivirus and labelled for aSyn (I, M), syntaxin (J, N), and MAP2 (K, O). Overlays are shown (L, P). Scale bar = 10 μm. Quantification of aSyn synaptic mean fluorescence intensity (Q), synaptic area (R), and synaptic volume (S) of neurons infected with haSyn lentivirus compared to neurons infected with eGFP (n = 6–10/1 neurons/independent culture). (T) Quantification of Syntaxin-positive puncta of neurons infected with haSyn lentivirus compared to neurons infected with eGFP (n = 6-10/1 neurons/independent culture). All data are expressed as mean ± SEM; Student’s t-test; * p-value ≤ 0.05; ** p-value ≤ 0.01; *** p-value ≤ 0.001.