Abstracts

Rationale: The contribution of glia, and the associated inflammatory processes, to the pathogenesis of seizures has been demonstrated in experimental models, and is strongly supported by clinical evidence. These cells are activated by epileptogenic injuries or seizures to release proinflammatory mediators that increase neuronal excitability, thus promoting the establishment of a seizure generating network. A novel mechanism for efficient upstream control of pathologic brain inflammation in epilepsy is represented by the identification of micro(mi)RNAs acting as crucial post-transcriptional regulators of the innate immune response. Our goal was to study the anti-inflammatory and anti-ictogenic properties of miR146a which modulates the activation of the IL-1R/Toll-like receptor (TLR) pathway. This proinflammatory pathway has a demonstrated pro-ictogenic role in experimental models and is upregulated in human epileptic foci. We studied whether alterations of miR146a brain levels (i) affect the extent of brain inflammation in vivo; (ii) modify seizures induced acutely in mice.Methods: C57BL/6 adult mice were chronically implanted with bilateral hippocampal electrodes and unilateral injection cannula for intrahippocampal delivery of 7 ng kainate to induce acute recurrent seizures for ~90 min. Seizures were monitored and quantified by EEG analysis. Brain inflammation was induced in a separate group of mice by intracerebroventricular (icv) injection of 12.5 g/1 l lipopolysaccharide (LPS), a TLR4 agonist. Pre-miR146a (mimic), LNA-anti-miR146a (antagomir) or their respective scrambled controls were injected (1 g/1 l,icv) twice a day for 7 days, either alone or before kainate or LPS. qRT-PCR was used to measure IL-1 , TNF- , TLRs and miR146a levels in the hippocampus.Results: LPS increased TNF- , TLR4 and IL-1 mRNA in the hippocampus 24 h after injection, and these effects were prevented by the antagomir. The antagomir also prevented the LPS-induction of miR146a, and increased per se hippocampal cytokine mRNA levels while decreasing miR146a. miR146a knockdown induced by antagomir increased kainate induced seizures in mice. On the contrary, miR146a mimic induced an average 8-fold increase of hippocampal miR146a (n=8; p<0.01 vs control by Student s t test) and decreased by ~3-fold the number of seizures and time in seizures in kainate-injected mice (n=8 mice each group, p<0.01 vs control by Student s t test). Conclusions: Alterations of miR146a brain level modify seizures in mice, and modulate IL-1R/TLR mediated brain inflammation. Our data suggest that miR146a is induced in the brain as a feed-back anti-inflammatory mechanism for maintaining the homeostatic role of inflammation. During seizures, however, this mechanism is inefficient. Boosting this endogenous defence mechanism may represent an effective strategy to prevent the pathologic outcomes associated with uncontrolled brain inflammation, thus inhibiting seizures.