Abstracts

Rationale: Inflammatory pathways activated by seizures or brain damage, or by lipopolysaccharide (LPS) which mimics systemic infection, contribute to neuronal hyperexcitability. LPS via activation of Toll-like Receptor 4 (TLR4) lowers seizure threshold. TLR4 is also activated during tissue injury by damage-associated endogenous molecules, among which the high mobility group box 1 (HMGB1) that is a nuclear non-histone DNA-binding protein that facilitates gene transcription. HMGB1 also exerts proinflammatory activity upon its extracellular release in disease conditions. We studied whether the activation of TLR4-signalling by HMGB1 contributes to seizure activity. Methods: Seizures were induced by intrahippocampal injection of kainic acid (KA) in mice and they were monitored and quantified by EEG analysis. TLR4 and HMGB1 expression was investigated in the hippocampus by immunohistochemistry, 1h and 3h after KA. HMGB1 was injected before KA to study its effect on seizures; TLR4 was blocked using selective antagonists or mutant mice with a defective TLR4 signalling; BoxA (competitive antagonist of HMGB1 receptors) was used to block endogenous HMGB1’s effects; ifenprodil was used to block NR2B-containing NMDA receptors. Results: HMGB1 expression was increased steadily in astrocytes but transiently in microglia after seizures onset; TLR4 was steadily expressed by neurons and astrocytes. HMGB1 was proconvulsant since it increased 2-fold the number and duration of seizures and reduced 2-fold the onset time of seizures. TLR4 antagonists and BoxA significantly reduced the frequency and the total time in seizures and delayed seizures onset by 2-fold. Mutant mice with impaired TLR4 signalling showed reduced seizure activity and a delay in the onset of seizures. HMGB1 did not increase seizures in TLR4 mutant mice. Ifenprodil was able to fully prevent the proconvulsant activity of HMGB1. Conclusions: The increased expression of HMGB1 and TLR4 in the hippocampus suggests the activation of a Toll-like receptor signalling pathway during seizures. HMGB1 has proconvulsant activity which depends on TLR4 and NR2B receptor activation. As previously described for IL-1β/IL-1R1 signalling (Balosso et al, Brain, 2008, 131(Pt 12):3256-65), this Toll-like receptor dependent pathway contributes to neuronal hyperexcitability resulting in enhanced seizure activity. Pharmacological blockade of TLR4 or inactivation of its endogenous ligands may represent an anticonvulsant strategy.