Fig. 2. Syntaxin 1A specifically inhibits EAAC1-mediated glutamate transport. In C6 glioma cells, the transfection of Syn1A and specific Syn1A siRNA (Syn1A siRNA) are indicated (A-D). In HEK293 cells, Syn1A are co-transfected pairwise with EAAC1, EAAT4 and GLT1; EAAC1 is co-transfected pairwise with syntaxin 4, SNAP-25 and VAMP-2 as indicated (E-G). (A) Syntaxin 1A significantly inhibits EAAC1 transport activity in C6 glioma cells. ^**P<0.01 compared with the cells transfected with ß-gal (n=4). (B) Kinetics analysis of glutamate transport in C6 glioma cells transfected with Syn1A. (Left panel) Saturation analysis of glutamate transport shows that Syn1A decreases transport maximal velocity compared with the cells transfected with ß-gal, whereas K_m is nearly identical. (Right panel) Eadie-Hofstee transformations of these data (n=4). (C) Specific siRNA of Syn1A increases EAAC1 transport activity in C6 glioma cells. ^**P<0.01 compared with the cells transfected with non-specific siRNA (NS siRNA) (n=5). (D) Kinetics analysis of glutamate transport in C6 glioma cells transfected with Syn1A siRNA. (Left panel) Saturation analysis shows that Syn1A siRNA increases EAAC1 transport maximal velocity compared with the cells transfected with NS siRNA, whereas K_m is nearly identical. (Right panel) Eadie-Hofstee transformations of these data (n=5). (E) EAAC1 transport activity is decreased progressively with increasing amounts of Syn1A plasmid. The HEK293 cells are triple-transfected with EAAC1, Syn1A, and ß-gal expression plasmids as indicated. ^**P<0.01 compared with control (n=3). (F) Syn1A has no effect on the transport activity of EAAT4 or GLT1 (n=3). (G) The transport activity of EAAC1 is not regulated by syntaxin 4, SNAP-25 or VAMP-2 (n=3).