Abstracts

Rationale: The molecular and cellular events responsible for the selective vulnerability of hippocampal neurons to pilocarpine-induced seizure activity are not completely understood. In the pilocarpine model of temporal lobe epilepsy (TLE) there is a marked involvement of excitotoxic neuronal injury (Mello et al., Epilepsia 1993; 34[6]: 985-995). Reactive oxygen species (ROS) production has been considered to be a part of the mechanisms involved with the glutamatergic excitotoxicity both in vitro and in vivo (Bonfoco et al., PNAS 1995; 92[16]:7162-7166), and it is well known that in the pilocarpine model of TLE there is an increase in the generation of ROS. However, the interaction between ROS generation and the neurodegeneration observed in that model is unknown. Therefore, the aim of this study is to evaluate the role of ROS in neuronal death in the hippocampus of rats submitted to the pilocarpine-induced status epilepticus (SE) during the acute phase. Methods: Male Wistar rats were first treated with methylscopolamine (s.c., 1 mg/kg) to limit the peripheral effects of the convulsant. SE was induced by injecting pilocarpine hydrochloride (i.p., 360 mg/kg) 30 minutes after methylscopolamine administration. Control animals received the methylscopolamine treatment and saline instead of pilocarpine. The animals were sacrificed 3h, 6h, 12h and 24h after the induction of SE and their brains were processed for detection of neurodegeneration using Fluoro-Jade B staining and for detection of ROS generation by means of dihydroethidine (DHE) fluorescence probe in CA1, CA3 and DG areas of the hippocampus. Data were analyzed using ANOVA followed by Tukey’s test and values of p<0.05 were accepted as significant. Results: After 12h of the beginning of the SE, a 16-fold and 11-fold increase was observed in neuronal death in CA1 and CA3 areas, respectively; in addition, an intense pattern of neurodegeneration was observed in these same areas 24h after the SE onset. This pattern of neuronal death was also seen in the DG in the same periods mentioned above (69-fold increase after 12h of SE and 145-fold increase after 24h of SE). In relation to the generation of ROS, no alterations were observed in the CA1 area of the hippocampus in the groups submitted to SE. An increase of 48%, when compared to the control, was detected in the CA3 area of the animals analyzed 6h after the onset of SE. All groups evaluated presented increase in the generation of ROS in the DG area when compared to the control animals (3h of SE: 25%; 6h of SE: 41%; 12h of SE: 31% and 24h of SE: 29%). Conclusions: The results showed an increase in the generation of ROS and more abundant neuronal death during the acute phase of the pilocarpine model of epilepsy. The data also revealed that the generation of ROS precedes the neuronal death in the CA3 and DG areas of the hippocampus of animals submitted to SE. Taken together, these preliminary results suggest that ROS could be involved in the neuronal damage induced by pilocarpine during SE. Financial Support: FAPESP and CNPq (Brazil).