Accumulating evidence supports that activation of inflammatory pathways is a crucial factor contributing to the pathogenesis of seizures. In particular, the activation of interleukin-1 beta (IL-1β) system exerts proconvulsant effects in a large variety of seizure models. Myeloid differentiation factor 88 (MyD88) is a critical adaptor protein in the signaling cascade elicited by IL-1β. The present study aimed to investigate the expression pattern of MyD88 in rat models of seizures and in patients with refractory temporal lobe epilepsy (TLE), and to study the role of MyD88 in epileptic seizures. Our results revealed that MyD88 was up-regulated in the hippocampus of rats in the lithium-pilocarpine model of acute seizures. Importantly, MyD88 overexpression was also significantly present in the brain from chronic epileptic rats and the temporal neocortex specimens from drug-resistant TLE patients. In the acute seizure model, both the behavioral and electrographic seizure activities were record and analyzed in rats for 90 min, starting immediately after pilocarpine injection. ST2825, a synthetic MyD88 inhibitor, was administered intracerebroventricularly (2.5–5.0–10 μg in 2 μl) 20 min before pilocarpine injection. We found that ST2825 at doses of 5 μg and 10 μg significantly inhibited the pilocarpine-induced behavioral and electrographic seizures. Moreover, 10 μg ST2825 prevented the proconvulsant actions of IL-1β. As previous evidence suggested that IL-1β proconvulsant effects was mediated by enhancing the phosphorylation level of the NR2B subunit of N-methyl-d-aspartate (NMDA) receptor, we then probed whether this molecular was involved in the effect of the pharmacological inhibition. Our results revealed that 10 μg ST2825 markedly reversed the increased Tyr1472-phosphorylation of the NR2B subunit of NMDA receptor observed in the proconvulsant conditions of IL-1β and in seizures induced by pilocarpine alone. These findings indicate that altered expression of MyD88 might contribute to the pathogenesis of seizures and targeting of this adaptor protein might represent a novel therapeutic strategy to suppress seizure activities.