The lethal expression of the GluR2flip/GluR4flip AMPA receptor in HEK293 cells

alpha -amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) -type glutamat e receptors play a critical role in excitotoxicity associated with cerebral hypoxia, ischaemia and other acute brain insults. AMPA receptors are compo sed of GluR1-GluR4 subunits in homomeric and heteromeric assemblies, formin g nonselective cation channels. In addition, each subunit has alternative s plice variants, flip and flop forms. Heterologous expression studies showed that the AMPA receptor channels exhibit diverse properties depending on su bunit/variant composition. For example, the absence of the GluR2 subunit ma kes AMPA receptor assemblies Ca2+-permeable. Excitotoxicity induced by acti vating AMPA receptor channels has been linked to excessive Ca2+ influx thro ugh the GluR2-lacking channels. Here we demonstrate that coexpression of th e AMPA receptor GluR2flip and GluR4flip subunits exerts a lethal effect on HEK293 cells, whereas no lethal activity is observed in other homomeric or heteromeric combinations of AMPA receptor subunits. Patch clamp recordings and Ca2+ imaging analyses have revealed that this GluR2flip/GluR4flip recep tor exhibits a low Ca2+ permeability. This subunit combination, however, sh owed prolonged Na+ influx following AMPA stimulation, even in the absence o f cyclothiazide, which attenuates AMPA receptor desensitization. Furthermor e, the GluR2flip/GluR4flip-mediated lethality was potentiated by the interr uption of cellular Na+ extrusion mechanisms using ouabain or benzamil. Thes e observations suggest that the GluR2flip/GluR4flip receptor-mediated excit otoxicity is attributed to Na+ overload, but not Ca2+ influx.