Dose-Dependent Induction of Differential Seizure Phenotypes by Pilocarpine in Rats: Considerations for Translational Potential

Abstract

Background and Objectives: Pilocarpine is used in experimental studies for testing antiepileptic drugs, but further characterization of this model is essential for its usage in testing novel drugs. The aim of our study was to study the behavioral and EEG characteristics of acute seizures caused by different doses of pilocarpine in rats. Materials and Methods: Male Wistar rats were treated with a single intraperitoneal dose of 100 mg/kg (P100), 200 mg/kg (P200), or 300 mg/kg (P300) of pilocarpine, and epileptiform behavior and EEG changes followed within 4 h. Results: The intensity and the duration of seizures were significantly higher in P300 vs. the P200 and P100 groups, with status epilepticus dominating in P300 and self-limiting tonic-clonic seizures in the P200 group. The seizure grade was significantly higher in P200 vs. the P100 group only during the first hour after pilocarpine application. The latency of seizures was significantly shorter in P300 and P200 compared with P100 group. Conclusions: Pilocarpine (200 mg/kg) can be used as a suitable model for the initial screening of potential anti-seizure medications, while at a dose of 300 mg/kg, it can be used for study of the mechanisms of epileptogenesis.

Keywords: EEG; behavioral changes; pilocarpine; rats; seizures.

Figure 1

The incidence of seizures (%) caused by a single intraperitoneal administration of pilocarpine at a dose of 100 mg/kg (P100), 200 mg/kg (P200), or 300 mg/kg (P300). The exact probability test was used to assess the significance of the difference (p = 0.6458).

Figure 2

The intensity of seizures induced by a single administration of pilocarpine in groups P100, P200, and P300 within 4 h after treatment. The intensity was assessed according to the descriptive scale and expressed as a grade from 0 to 4: 0—no seizures; 1—head nodding and clonic jaw twitching; 2—myoclonic body jerks (hot plate reaction), bilateral forelimb clonus with full rearing (Kangaroo position); 3—progression to generalized clonic convulsions followed by tonic extension of fore and hind limbs and tail; 4—prolonged severe tonic–clonic convulsions lasting over 15 s (status epilepticus) or frequent repeated episodes of clonic convulsions for an extended period of time (over 5 min). The significance of the difference was estimated by Friedman with Dunn–Bonferroni post-hoc test (* p < 0.05 vs. P100, # p < 0.05 vs. P200, ** p < 0.001 vs. P100, ## p < 0.001 vs. P200). For detailed information, see Figure 1.

Figure 3

Latency period (s) of seizures caused by a single administration of pilocarpine in the P100, P200, and P300 groups. Latency period is defined as a time between pilocarpine administration and the first seizure sign. The significance of the difference was estimated by Kruskall–Wallis with Mann–Whitney post-hoc test (* p < 0.05 vs. P100). For detailed information, see Figure 1.

Figure 4

Representative EEG findings in pilocarpine-treated groups obtained in the first hour after pilocarpine treatment from the parietal cortex. (A) Note activity without signs of epileptiform discharges. Basal EEG activity in the beta band (14–30 Hz) corresponding to the physiological behavior before pilocarpine administration. (B) Intermittent paroxysmal polyspikes and spike–wave complexes in the P100 group. (C) Epileptiform spike bursts of variable amplitude lasting from 2.2 to 14.1s in the P200 group and high voltage spikes with amplitudes of more than 300–500 μV. (D) High-voltage, high-frequency polyspike burst activity combined with intermittent spike–wave complexes lasting for up to 200 s, corresponding to status epilepticus. For detailed information, see Figure 1.