Coadministration of Flunarizine Prevents Tolerance to Nitrazpam in Mice
Authors: Suzuki Yasuhiro , Nagai Toshisaburo , Goto Megumi , Futagi Yasuyuki , Okada Shintaro 
Published: August 2009

Purpose: Benzodiazepines (BZDs) are the most potent drugs available for the emergency treatment of seizures. For the long‐term treatment of epilepsy, however, BZDs are often administered in combination with other antiepileptic drugs(s) (AEDs) because of the development of tolerance. In this study, the influence of concurrently administered AEDs [carbamazepine (CBZ), phenytoin (PHT), zonisamide (ZNS), vigabatrin (VGB), lamotrigine (LTG), or flunarizine (FNR)] on the tolerance to the anticonvulsant action of the BZDs was studied by using an animal tolerance mode. Methods: In the first experiment, adult male mice (ICR, 30–35 g) were given nitrazepam (NZP; 0.6 mg/kg, i.p.) twice daily (8:00 a.m. and 6:00 p.m.) for 0 (acute), 0.5, 1, 3, or 5 days before the test day. On the morning (8:00 a.m.) of the test day, all mice were given pentylenetetrazole (PTZ; 150 mg/kg, i.p.) 30 min after the injection of a single protective dose of NZP (0.6 mg/kg). Mice were then observed for 30 min for the development of unequivocal clonic convulsions (duration, <3 s). In the second experiment, mice were randomly assigned to one of eight groups depending on the coadministered AED. Mice received vehicle, NZP (0.6 mg/kg, i.p.) alone or NZP concurrent with one of the six AEDs (CBZ, 6.25 mg/kg; PHT, 10 mg/kg; ZNS, 20 mg/kg; VGB, 250 mg/kg: LTG, 15 mg/kg; and FNR, 10 mg/kg) twice daily for 5 days. A single dose of CBZ, PHT, ZNS, VGB, LTG, or FNR alone was ineffective against the clonic convulsions induced by PTZ (150 mg/kg). On the test day, several mice were randomly chosen from each group, and blood samples were collected to determine the concentration of NZP at 30 min after the protective injections of NZP. The remaining mice were tested as mentioned. Tolerance to the anticonvulsant action of NZP was considered to develop when the difference in seizure frequency was statistically significant (by the x2 test) between the non‐NZP treatment group (experiment 1, acute group: experiment 2, vehicle‐treated group) and another treatment group. Results: In mice treated with NZP alone, tolerance was evident even after the second dose (1 day) but did not increase further after 3 or 5 days of treatment; significant increases in seizure frequency (45–55%) were found in mice treated with NZP for 1, 3, or 5 days, as compared with the acute group (9.7%), but there were no significant differences between the duration of treatment (1, 3, or 5 days). Tolerance to the anticonvulsant action of NZP developed in mice treated with NZP plus CBZ, PHT, ZNS, VGB, or LTG. On the other hand, mice receiving NZP + FNR showed no tolerance; there was no significant difference in seizure frequency between the vehicle group (four of 21) and NZP + FNR group (four of 21), but a significant difference (p > 0.05) was observed between the NZP alone group (12 of 21) and NZP + FNR group. There were no significant differences in plasma NZP concentrations between three groups (vehicle, 560.8 ± 275.4 ng/ml; NZP alone, 398.6 ± 216.4 ng/ml; and NZP + FNR, 537.9 ± 130.2 ng/ml). Conclusions: These data suggest that coadministration of FNR but not CBZ, PHT, ZNS, VGB, or LTG may delay, if not prevent, the development of tolerance to the anticonvulsant action of BZDs.