Figure 3.

Expression of AQP proteins in HEK-293 cells induces EAAT2 protein expression. DNA is stained blue (A and B). (A) EAAT1 (red) is expressed constitutively, regardless of transfection. (B) EAAT2 (red) is localized in the plasma membrane of cells expressing AQP4 or AQP5 (green), but is undetectable in cells transfected with GFP-vector lacking an AQP cDNA. Bars, 10 μm. (C) For RT-PCR, all three cell lines express EAAT2 mRNA, regardless of AQP expression. Markers indicate basepair reference standards. (D) The Western blot shows that EAAT2 protein is up-regulated in cells transfected with AQP4 or AQP5, but not in GFP-vector–transfected cells. (E) EAAT1 protein and actin are expressed constitutively. Markers indicate kilodalton reference standards (D and E). (F) In glutamate transport experiments, GFP-AQP4 cells take up approximately threefold more l-[³H]glutamate than vector-transfected cells (note: Na⁺ dependence). All experiments were performed a minimum of two times. The error bars represent the standard error of six and four replicates, respectively.