Post-exposure administration of diazepam combined with soluble epoxide hydrolase inhibition stops seizures and modulates neuroinflammation in a murine model of acute TETS intoxication

Abstract

Tetramethylenedisulfotetramine (TETS) is a potent convulsant poison for which there is currently no approved antidote. The convulsant action of TETS is thought to be mediated by inhibition of type A gamma-aminobutyric acid receptor (GABAAR) function. We, therefore, investigated the effects of post-exposure administration of diazepam, a GABAAR positive allosteric modulator, on seizure activity, death and neuroinflammation in adult male Swiss mice injected with a lethal dose of TETS (0.15 mg/kg, ip). Administration of a high dose of diazepam (5 mg/kg, ip) immediately following the second clonic seizure (approximately 20 min post-TETS injection) effectively prevented progression to tonic seizures and death. However, this treatment did not prevent persistent reactive astrogliosis and microglial activation, as determined by GFAP and Iba-1 immunoreactivity and microglial cell morphology. Inhibition of soluble epoxide hydrolase (sEH) has been shown to exert potent anti-inflammatory effects and to increase survival in mice intoxicated with other GABAAR antagonists. The sEH inhibitor TUPS (1 mg/kg, ip) administered immediately after the second clonic seizure did not protect TETS-intoxicated animals from tonic seizures or death. Combined administration of diazepam (5 mg/kg, ip) and TUPS (1 mg/kg, ip, starting 1 h after diazepam and repeated every 24 h) prevented TETS-induced lethality and influenced signs of neuroinflammation in some brain regions. Significantly decreased microglial activation and enhanced reactive astrogliosis were observed in the hippocampus, with no changes in the cortex. Combining an agent that targets specific anti-inflammatory mechanisms with a traditional antiseizure drug may enhance treatment outcome in TETS intoxication.

Combined treatment with diazepam and allopregnanolone reverses tetramethylenedisulfotetramine (TETS)-induced calcium dysregulation in cultured neurons and protects TETS-intoxicated mice against lethal seizures

“…To determine whether these in vitro observations are predictive of in vivo results, we assessed the efficacy of pretreatment with diazepam and allopregnanolone singly or in combination in preventing death of mice intoxicated with a lethal dose of TETS. The lethal dose of TETS was 0.15 mg/kg i.p., as determined previously (Zolkowska et al, 2012; Vito et al, 2014), and doses of diazepam and allopregnanolone that were ineffective or partially effective when administered singly were determined in pilot dose-range finding studies (data not shown). Mice were treated (i.p.)…”

“…(Vito et al, 2014). When administered singly or in combination, diazepam or allopregnanolone at 0.03 mg/kg i.p.…”

“…In our initial studies using high dose diazepam to rescue TETS-intoxicated mice (Vito et al, 2014), we observed a pronounced sedative/anesthetic response following diazepam treatment. Treatment with a lower dose of diazepam in combination with a low dose of allopregnanolone did not seem to elicit this response.…”

“…Experimental evidence suggests that benzodiazepines provide some protection against TETS-induced seizures. For example, in mice, treatment with a high (5 mg/kg) dose of diazepam following exposure to a lethal dose of TETS reduced seizure manifestations and prevented lethality (Shakarjian et al, 2012; Vito et al, 2014), but did not eliminate ictal activity in the electroencephalogram (Shakarjian et al, 2012) or mitigate neuroinflammatory responses (Vito et al, 2014). Benzodiazepines are the standard-of-care treatment for acute seizures and high doses of diazepam or another benzodiazepine are often administered to humans experiencing TETS-induced seizures (Whitlow et al, 2005).…”

Rapid Throughput Analysis Demonstrates that Chemicals with Distinct Seizurogenic Mechanisms Differentially Alter Ca2+ Dynamics in Networks Formed by Hippocampal Neurons in Culture

“…In mice, intraperitoneal injections of TETS or PTX cause similar sequences of immobility, myoclonic body jerks, clonic seizures of the forelimbs and/or hindlimbs, tonic seizures (falling on the side followed by forelimb tonic contraction and hindlimb tonic extension), and eventually death (Zolkowska et al, 2012). Systematic TETS exposures that are not fatal produce transient gliosis (2-3 days postexposure) in both the hippocampus and cortex without evidence of cellular injury and neurodegeneration (Zolkowska et al, 2012;Vito et al, 2014). In contrast, administration of KA to rats induces a distinct progression of symptoms including staring episodes, head bobbing, numerous wet dog shakes, and isolated limbic motor seizures that increase in frequency, eventually leading to status epilepticus (Scerrati et al, 1986), which resemble the clinic features of human temporal lobe epilepsy (Ben-Ari, 1985).…”