Characteristics of murine phenotypes with signs of epilepsy obtained through ENU mutagenesis

Abstract

Epilepsy is a chronic neurological disorder marked by recurring seizures and associated dysfunctions of motor, sensory, autonomic, and mental functions resulting from excessive electrical activity of neurons. Identification and characterization of the mutations that cause this pathology are essential for understanding the mechanisms that control epileptogenesis. The aim of the study is to characterize the S8-3 mutant mice strain, which shows induced epileptiform activity in response to audiogenic stimulation. Mutant strains of mice were acquired through induced chemical mutagenesis using N-ethyl-N-nitrosourea (ENU). Twenty-nine male mice were administered three rounds of ENU injections at a dose of 90 mg/kg during the study. On day 21 after birth (P21), identification and selection of mouse mutants with an elevated inclination towards epileptic seizures were conducted using the Krushinsky scale, which considered the intensity of audiogenic seizures. Strains with a recessive mutation were created by selecting animals that exhibited the aberrant phenotype for the second time. The abnormal phenotype was confirmed in the G5 generation, and basic behavioral phenotyping was performed to characterize the resulting epileptic lines. Correct grammar, spelling, and punctuation. This included assessing memory, learning ability, motor reactions, and emotional status. In vitro experiments assessed spontaneous calcium activity with primary neuronal cultures of the cerebral cortex isolated from newborn mice. The Ca2+ indicator used was Oregon Green 488 BAPTA-1 AM. Upon screening 60 strains of mice for sensitivity to audiogenic stimulation, 12 strains exhibiting epileptiform activity were observed. Subsequently, the S8-3 group was selected for further research, as its offspring (G3) showed a higher frequency of the aberrant phenotype in comparison to the other groups. Results from behavioral studies comparing S5-1 mice to the control hybrid animal group displayed a higher intensity of the acoustic startle response. The open field tests revealed that the motor activity of S8-3 strain mice was higher than that of the control group, based on the average distance traveled, and their anxiety level was lower, as indicated by fewer rears, urinary points, and boluses. When assessing cognitive functions through the CPAR test, mutant individuals exhibited high learning ability. In vitro experiments showed an increase in the frequency of spontaneous calcium events in primary cell cultures of the cerebral cortex isolated from S8-3 mice.