AbstractIt has been suggested that reduced glutamate receptor expression protects glioma cells from glutamate toxicity. GluR2 is the critical subunit of the GluR2 subtype of AMPA glutamate receptors as this subunit determines the Ca2+ permeability of the receptor. The gene encoding the GluR2 subtype of AMPA receptors has been described as a target gene for the transcription repressor REST. However, we recently showed that the GluR2 gene is not regulated by REST in several neuronal and neuroendocrine cell lines, due to a repressive chromatin environment. Here, we show that the GluR2 gene has an open chromatin configuration in human glioma cells. Overexpression of REST reduced GluR2 mRNA levels while shRNA‐mediated depletion of REST or expression of a REST mutant, that contained a transcriptional activation domain, enhanced GluR2 expression. Incubation with trichostatin A (TSA), a histone deacetylase inhibitor, induced acetylation of histone 4 of the GluR2 locus in glioma cells, leading to an upregulation of GluR2 expression. Together, these data suggest that REST is responsible for the reduced expression of GluR2 in glioma cells. The transcription factor Sp1 additionally binds under physiological conditions to the GluR2 gene in human glioma cells and expression of a dominant‐negative mutant of Sp1 reduced expression of GluR2. Thus, the regulation via Sp1 represents a further control point for GluR2 expression in glioma cells. Together, we show that the GluR2 gene is embedded into an open chromatin configuration in glioma cells and expression of GluR2 is controlled by REST and Sp1. J. Cell. Biochem. 113: 528–543, 2012. © 2011 Wiley Periodicals, Inc.