Abstracts

Rationale: Interleukin (IL)-1beta and its signal-transducing receptor (IL-1R1) are chronically overexpressed in glia and neurons in epileptic tissue from rodents and from patients with drug resistant epilepsies. IL-1beta contributes to the generation and maintenance of seizures in experimental models by increasing NMDA receptor-mediated neuronal excitability. We investigated the intracellular pathway that could be involved in the proconvulsant effects of IL-1beta in the mouse hippocampus. Methods: Seizures induced by intrahippocampal (ih) injection of kainic acid (KA) were monitored and quantified for 3h by EEG analysis in freely-moving adult male C57BL6 mice. IL-1-induced sphingomyelinase-dependent ceramide and activation of Src-kinase signaling were investigated pharmacologically by using systemic or ih administration of selective compounds that act along this pathway. Protein levels were measured in hippocampal homogenates by western blot (WB) analysis, 1 h after KA administration in additional groups. Results: C2-ceramide (40-80 µg/kg, ih), a cell permeable analog of ceramide mimicked the proconvulsant effects of 1ng IL-1beta by increasing 2- and 3-fold respectively the number and total duration of KA-seizures. 3-O-methylsphingomyelin (3µg/0.5µl, ih), a selective inhibitor of sphingomyelinase, and ifenprodil (1mg/kg, intraperitoneally), a NR2B-selective NMDA antagonist, did not affect seizures when given alone but prevented the proconvulsant effect of IL-1beta when coadministered with this cytokine. IL-1beta or KA similarly increased by 20% and 50% the phosphorylated form of NR2B (NR2B-P) and Src kinase (Src-P) respectively compared to vehicle-treated mice. In IL-1beta+KA-treated mice, NR2B-P and Src-P were increased by 50% and 110% respectively compared to vehicle-treated mice. Conclusions: A sphingomyelinase-dependent pathway activated by IL-1beta during seizures leads to the production of ceramide which in turn induces Src-dependent phosphorylation of NR2B subunit of NMDA receptor. This pathway is involved in the proconvulsant actions of IL-1beta and may be targeted to attain fast anti-inflammatory effects and seizure inhibition.