Induction of epileptic seizures

To induce sustained epileptic seizures, 8-week-old male C57BL/6 N mice (CLEA) were injected intraperitoneally with pilocarpine (280–300 mg/kg in 0.9% NaCl) as described previously⁵⁵. To minimize peripheral side effects of pilocarpine, mice were injected intraperitoneally with scopolamine methyl nitrate (2 mg/kg) 30 min prior to pilocarpine administration. Control mice were treated with scopolamine methyl nitrate alone. Seizures were classified according to the previously-published reports^{56, 57} with slight modifications⁵⁵. Status epilepticus was defined as sustained constant motor seizures of stage 3 or greater, which was terminated by injection of 30–40 mg/kg pentobarbital 4.5 h after the onset of status epilepticus. After status epilepticus, all animals were given soaked rodent food and 2 ml of Lactec D subcutaneously. Of pilocarpine-treated mice, only those that reached stage 6 at least once and exhibited status epilepticus (stage 3.5) for 4.5 h⁵⁵ were used for this study. The experiments were performed 1 to 4 weeks after the administration of pilocarpine.