Increased ambient glutamate concentration alters the expression of NMDA receptor subunits in cerebellar granule neurons

Effects of prolonged (48 h) inhibition of glutamate reuptake on the relativ e abundance of mRNAs coding for N-methyl-D-aspartate (NMDA) receptor subuni ts, and the expression of corresponding proteins were investigated in prima ry cultures of rat cerebellar granule neurons. In cells exposed to the glut amate transport blocker, L-trans-pyrrolidine-2,4-dicarboxylate (PDC), the e xpression of the C1 exon-positive NR1 mRNA variant was reduced by about 40% whereas, the expression of Cl-negative mRNA was increased leading to signi ficant reduction of the +Cl/-C1 ratio. The expression of the Nl-negative NR 1 variants was slightly reduced following exposure to PDC, indicating that increased medium levels of glutamate changed the relative abundance of NR1 splice-variant expression but did not reduce the overall NR1 transcription. Expression of NR2A and NR2B mRNAs was 40-50% lower in PDC-treated cells as compared to control. Immunoblot experiments revealed that PDC exposure red uced the expression of NR1 and all NR2 proteins with NR2A and NR2B proteins being reduced to a greater extent than NRL The overall decrease in NMDA re ceptor subunit protein expression was considerably more pronounced than the reduction of their corresponding mRNAs, suggesting involvement of a post-t ranscriptional regulation. Our data support the hypothesis that functional activity and number of NMDA receptors are regulated by strength of the glut amatergic input. Thus, reduced glutamate uptake resulting in increased conc entration of ambient glutamate initiate a series of adaptive responses mani fested as a gradual down-regulation of the functional activity and expressi on of NMDA receptors. (C) 2001 Elsevier Science Ltd. All rights reserved.