FMRP Mediates Chronic Ethanol-Induced Changes in NMDA, Kv4.2, and KChIP3 Expression in the Hippocampus

Background: Exposure to chronic ethanol (EtOH) results in changes in the expression of proteins that regulate neuronal excitability. This study examined whether chronic EtOH alters the hippocampal expression and function of fragile X mental retardation protein (FMRP) and the role of FMRP in the modulation of chronic EtOH-induced changes in the expression of NMDA receptors and Kv4.2 channels. Methods: For in vivo studies, C57BL/6J mice underwent a chronic intermittent EtOH (CIE) vapor exposure procedure. After CIE, hippocampal tissue was collected and subjected to immunoblot blot analysis of NMDA receptor subunits (GluN1, GluN2B), Kv4.2, and its accessory protein KChIP3. For in vitro studies, hippocampal slice cultures were exposed to 75 mM EtOH for 8 days. Following EtOH exposure, mRNAs bound to FMRP was measured. In a separate set of studies, cultures were exposed to an inhibitor of S6K1 (PF-4708671 [PF], 6 mu M) in order to assess whether EtOH-induced homeostatic changes in protein expression depend upon changes in FMRP activity. Results: Immunoblot blot analysis revealed increases in GluN1 and GluN2B but reductions in Kv4.2 and KChIP3. Analysis of mRNAs bound to FMRP revealed a similar bidirectional change observed as reduction of GluN2B and increase in Kv4.2 and KChIP3 mRNA transcripts. Analysis of FMRP further revealed that while chronic EtOH did not alter the expression of FMRP, it significantly increased phosphorylation of FMRP at the S499 residue that is known to critically regulate its activity. Inhibition of S6K1 prevented the chronic EtOH-induced increase in phospho-FMRP and changes in NMDA subunits, Kv4.2, and KChIP3. In contrast, PF had no effect in the absence of alcohol, indicating it was specific for the chronic EtOH-induced changes. Conclusions: These findings demonstrate that chronic EtOH exposure enhances translational control of plasticity-related proteins by FMRP, and that S6K1 and FMRP activities are required for expression of chronic EtOH-induced homeostatic plasticity at glutamatergic synapses in the hippocampus.