Pharmacological profiling of conventional antiseizure medicines against seizures and kindling acquisition in female mice

Abstract

Rationale: Historical ASM discovery has primarily relied on demonstration of efficacy in clinically validated acute seizure models evoked in male, wild-type rodents (e.g., maximal electroshock, subcutaneous pentylenetetrazol, or kindling). However, little work has been done to establish ASM response profiles in female rodents with acute or chronic seizures. Therefore, this thesis aimed to build on our previously established female C57BL/6J studies (Lehmann & Barker-Haliski 2023) to fill this knowledge gap and to define the acute median effective dose (ED50) of 4 FDA-approved ASMs (levetiracetam (LEV), carbamazepine (CBZ), valproic acid (VPA), and phenobarbital (PB)) in female outbred CF-1 mice using the 6 Hz 32 mA model of focal seizures. Further, no compound has yet been found to prevent the development of epilepsy so we propose a moderate-throughput anti-epileptogenesis drug screening paradigm for early compound profiling via repeated administration of anticonvulsant doses of VPA to assess the potential to prevent the formation of chronic hyperexcitable neuronal network or modify the severity of behavioral comorbidities of epilepsy in male versus female mice using the corneal kindled mouse (CKM) model of chronic seizures. Methods: Female outbred CF-1 mice were used to quantify of an ED50 of candidate ASMs (n=8 mice/dose) in a 6 Hz 32 mA test. Motor impairment was assessed via performance on a fixed speed rotarod. To establish a moderate-throughput, early-stage disease modification screening paradigm, male (n=40) and female (n=45) CF-1 mice underwent 60 Hz corneal kindling, a model of chronic network hyperexcitability, for 3-4 weeks in the presence or absence of twice daily VPA (150 mg/kg, intraperitoneal – i.p.) or vehicle administration. Mice were monitored twice daily for the presence and severity of evoked behavioral seizures until all animals met the kindled seizure endpoint. Following the kindling acquisition period, all CKM were challenged in an open field test to quantify the extent to which ASM administration during the kindling process modified anxiety-like behavioral comorbidities of epilepsy. Results: The calculated ED50 [and 95% confidence intervals] in female CF-1 mice for VPA (100 mg/kg [60.5-139]), CBZ (14.0 mg/kg [8.77-23.1]), LEV (14.3 mg/kg [0.591-36.0]) and PB (18.3 mg/kg [6.8-30.4]) were consistent with our previously reported values in the 6 Hz test in inbred female C57BL/6J mice (Lehmann and Barker-Haliski 2023). Notably, CBZ was poorly tolerated in CF-1 female mice: 87.5% of female mice were impaired on a rotarod at the highest dose tested. No other motor impairment was noted. In both male and female corneal kindling acquisition studies, VPA prevented kindled seizure presentation. The number of CKM reaching the fully kindled state was lower in both female (2/10) and male (2/11) mice than in VEH-treated female (8/10) and male (14/15) mice (male: X2 value = 18.57, p < 0.001; female: X2 value = 12.77, p < 0.001). However, repeated VPA administration did not significantly impact CKM activity in the center of an open field versus the activity of VEH-treated mice, suggesting no disease modifying effect of VPA administration on the anxiety-like behavioral comorbidity of epilepsy. Conclusion: This study establishes the anticonvulsant properties of four prototypical ASMs in wild-type outbred female CF-1 mice to inform future preclinical epilepsy studies using novel investigational compounds. This work also establishes the feasibility of a disease-modification drug screening paradigm for epilepsy in male and female mice using the corneal kindled mouse model and open field test of anxiety-like behaviors, a known clinical comorbidity of epilepsy. Altogether, this work offers a strategy on which to rapidly assess the anticonvulsant and disease modifying potential of candidate investigational treatments for the symptomatic management, and possibly prevention, of epilepsy.