Isoflurane enhances the expression and activity of glutamate transporter type 3 in C6 glioma cells

Background: Glutamate transporters play an important role in maintaining extracellular glutamate homeostasis. Volatile anesthetics have been shown to affect glutamate transporter activity acutely (within minutes after the exposure). It is not known whether volatile anesthetics affect the expression of glutamate transporters. Methods: Rat cultured C6 glioma cells that express excitatory amino acid transporter type 3 (EAAT3) were exposed to isoflurane at various concentrations (0.5-4.0%) or for different periods (1-24 h) at 37degrees. EAAT3 mRNA, proteins, and activity were quantified. Results: lsoflurane induced a time- and concentration-dependent increase in the mRNA and protein levels of EAAT3 in C6 cells. The maximal increase was induced by 2% isoflurane, and the cells incubated with 2% isoflurane for 3 and 7 h expressed the highest levels of EAAT3 mRNA and proteins, respectively. Similarly, glutamate uptake was higher in C6 cells exposed to 2% isoflurane for 7 h than in control cells. Actinomycin D and cycloheximide, inhibitors for mRNA and protein synthesis, respectively, did not affect the isoflurane-induced increase in EAAT3 mRNA and protein expression. Phorbol 12-myristate 13-acetate, a protein kinase C activator, also enhanced EAAT3 expression. The combination of 2% isoflurane and phorbol 12-myristate 13-acetate caused a higher level of EAAT3 expression than that induced by 2% isoflurane alone. Neither staurosporine, a protein kinase C inhibitor, nor wortmannin, a phosphatidylinositol 3 kinase inhibitor, inhibited the isoflurane-induced increase in EAAT3 expression. Conclusions: The results of this study suggest that isoflurane increases the expression and activity of EAAT3 by stabilizing EAAT3 mRNA and proteins via protein kinase C- and phosphatidylinositol 3 kinase-independent pathways.