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Fig. 4 | Acta Neuropathologica Communications

Fig. 4

From: Organotypic slice culture model demonstrates inter-neuronal spreading of alpha-synuclein aggregates

Fig. 4

Application II. Demonstrating that phosphorylation of S129 on α-syn is not a prerequisite for seeding α-syn aggregation or trans-synaptic spreading in hippocampal slices. a Experimental setup with establishment of neuronal expression of either WT- or non-phosphorylatable S129G-α-syn in α-syn KO slices prior to initiation of templated α-syn aggregation by injection of S129A PFFs. b Validation of virally mediated WT- and non-phosphorylatable S129G-α-syn expression in α-syn KO slices using antibodies against total and pS129-α-syn (11A5). c Expression of WT α-syn supports establishment of MJF-14- and pS129-positive (11A5) aggregate pathology in the DG region following PFF injection at DG. Magnified panels show axonal aggregates (i) and cell body inclusions (ii) at DG. Scale bar: 50 μm, i: 20 μm, ii: 5 μm. d MJF-14- and pS129 positive (11A5) pathology spreads to the CA1 region within 7 dpi. Scale bar: 50 μm. e Expression of S129G-α-syn supports establishment of MJF-14-positive/pS129-negative aggregate pathology in the DG, present in axons (i, arrows) and cell bodies (i, arrowheads). Scale bar: 50 μm, i: 20 μm. f The non-phosphorylated MJF-14-positive aggregate pathology spreads to the CA1 region within 7 dpi. Scale bar: 50 μm. Western blot in b is representative of 3 independent experiments, while images in c-f are illustrative of 3–5 experiments with 21–30 slices in total per condition

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