# Intranasal Diazepam Outperforms Midazolam and Lorazepam in Early Seizure Control in Pilocarpine-Induced Mice Model

## TL;DR

This study finds that intranasal diazepam is more effective than midazolam and lorazepam for early seizure control in a mouse model of epilepsy.

## Contribution

The study is the first to compare intranasal benzodiazepines in a pilocarpine-induced seizure model and identifies diazepam as the most effective.

## Key findings

- Intranasal diazepam showed the highest brain-to-plasma ratio and best seizure control at 10 minutes.
- Diazepam reduced TNF-α mRNA and protein levels more effectively than midazolam and lorazepam.
- All three drugs had similar brain binding and receptor affinity, but diazepam had the lowest plasma protein binding.

## Abstract

Epilepsy is a central nervous system disease characterized by the sudden onset of seizures, loss of consciousness, or confusion. In recent years, many non-intravenous routes of administration of benzodiazepines have been developed for seizure control, with intranasal administration being an attractive route of choice. However, for such a route of administration, there is a lack of evidence on the choice of the epilepsy drug. This study aims to compare the intranasal formulations of three first-line drugs for seizure control, namely midazolam, diazepam, and lorazepam, via an ideal intranasal treatment. A pilocarpine-induced seizure model in mice was used to compare drug efficacy. The three drugs were administered intranasally to 36 C57 mice at a single dose of 1 mg/kg, followed by inducing the seizures via intraperitoneal injection of pilocarpine. The subsequent seizure scores were observed for either 10–100 min. After sacrificing the mice at 10/100 minutes post-dosing, whole brain tissue and plasma were collected to analyze the drug concentrations as well as brain-to-plasma concentration ratios. In addition, effects of these intranasally delivered drugs on neuroinflammation-associated molecules were monitored and compared via the mRNA levels and/or protein expression of GABARα1, TNF-α, and IL-1β in the cortex and hippocampus. Additionally, drug bindings to plasma and brain tissue obtained using the ultrafiltration method were compared, and drug binding affinities towards the GABAA receptor, as determined via the computer docking technique, were also compared. Among the three tested drugs, our results suggested that intranasal diazepam, with the highest brain-to-plasma ratio, was the best in seizure control at 10 min. Although all three drugs showed good stability, similar brain binding and receptor binding affinity, diazepam demonstrated the greatest efficacy in reducing TNF-α mRNA and protein levels, and lowest plasma protein binding, which could contribute to its higher brain-to-plasma ratio and better acute epilepsy control compared to the other two drugs. Our pilot in vivo experiments in the pilocarpine-induced mice seizure model for the first time demonstrated that intranasally administered benzodiazepines are effective for seizure control at the early stage, with intranasally delivered diazepam being the most potent one.

The online version contains supplementary material available at 10.1007/s11064-026-04720-y.