Objective:

To identify a suitable diazepam (DZP) dosing strategy in rats for subsequent subchronic in vivo efficacy studies.

Background:

DZP elimination pharmacokinetics are substantially faster in rats than humans, complicating subchronic and chronic studies in rats. Previous rat studies suggest that repeat doses of intraperitoneal (IP) DZP 20 mg/kg result in sustained DZP accumulation in serum and cerebrospinal fluid but impair neurologic function, measured by righting reflex. 

Design/Methods:

Adult male Sprague Dawley rats (~400 g) were administered 3 repeat doses of IP DZP (0.75, 1.5, 3, or 6 mg/kg) 1 hour apart. A separate group received only a single 3-mg/kg dose. Righting reflex was evaluated. Blood was collected at 10 minutes and 1, 3, and 6 hours after the final injection. Brain samples were collected at 1, 3, or 6 hours. Dose-dependent changes in plasma and brain DZP concentrations were evaluated.

Results:

Repeat dosing of DZP (0.75–3 mg/kg) resulted in a dose-dependent increase in plasma and brain DZP levels up to 6 hours. A single 6-mg/kg dose resulted in substantial impairment of righting reflex and was not studied further. Brain-to-plasma ratios suggest a modest accumulation of DZP in the brain as long as 6 hours (3 mg/kg: 38.3±34.6 ng/g vs 23.8±18.4 ng/mL). Brain accumulation of DZP at 6 hours was greater after repeat doses vs a single dose (repeat: 38.2±34.6 ng/g vs single: 6.1±3.4 ng/g). 

Conclusions:

In this rat model, a repeat-dosing paradigm of DZP ≤3 mg/kg resulted in sustained DZP levels in plasma and brain. Accumulation of DZP in rat brain vs plasma suggests that DZP may be long-acting at the site of action. This repeat-dosing paradigm for DZP in rats mimics plasma drug concentrations seen in humans, offering a preclinical tool to study the impact of benzodiazepine rescue therapy on seizure-cluster biology.