Fig. 6

Interaction of α-syn and EAAT2 following RBC-EV exposure. a Schematic representation demonstrating the proximity ligation (PL) assay using antibodies against EAAT2 and human α-syn. b Representative images of cultured astrocytes containing EAAT2/α-syn complexes (proximity ligation products) treated with DiI-labeled RBC-EVs. White arrows indicate DiI-labeled RBC-EVs overlapping with proximity ligation products, white arrow heads indicate DiI-labeled RBC-EVs not overlapping with proximity ligation products (Scale bar, 10 μm). c Quantification analysis of percentage of astrocytes containing EAAT2/α-syn complexes (means + S.E.M; n = 3; ****p < 0.0001 by One-way ANOVA test). Note that the presence of dynasore eliminated the RBC-EV-induced increase of EAAT2/α-syn complexes. d Western blot analysis of EAAT2 (E2) immunoprecipitates (IP) from the lysates of A53T mouse brain performed with antibodies against EAAT2 and human α-syn (211). IP with control nonimmune rabbit immunoglobulins (IgG) served as control. e Quantification analysis of oligomeric α-syn/EAAT2 co-localization in cortex (CTX), striatum (STR), midbrain (MIDB) and cerebellum (CERE) (means + S.E.M; n = 5 independent animals were used in each group; ****p < 0.0001 by One-way ANOVA test)