Autophagy Inhibitor 3-Methyladenine Improves Learning Ability And Its Underlying Mechanism In A Neonatal Rat Model Of Recurrent Seizures

Object: To explore the effects of 3-Methyladenine on neonatal seizure-induced cognitive deficit and its underlying mechanism.Methods: Ninety Sprague-Dawley neonatal rats(postnatal days 6,P6)were randomly divided into three groups:the control group (n=30),the recurrent-seizure group (RS group,n=30) and the 3-MA treated-seizure group (3-MA group,n=30).The volatile agent flurothyl was used to induce seizure attack.Starting at postnatal day 6(P6),rats were exposed to 45 flurothyl-induced seizures over a 9-day period of time.Control rats were placed into the container for an equal amount of time to their counterpart without exposure to flurothyl.Rats in the 3-MA group were pretreated with 3-Methyladenine before exposured to flurothyl.At 1.5h,3h,6h and 24h after the last convulsion, Beclin1 and LC3 protein level in hippocampus and cerebral cortex were detected by western blot method.Morris water maze test were performed to assess learning and memory ability during P31-P37.All the date was analyzed using the software SPSS 16.0.Results: 1.Western blot analysis (1) The ratio of LC3II to LC3I in hippocampus and cerebral cortex of RS group was significantly increased than that of contral group at 1.5h,3h,6h and 24h(p<0.05).The ratio of LC3II to LC3I in hippocampus and cerebral cortex of 3-MA group was significantly decreased than that of RS group(p<0.05). (2) The level of Beclin1 protein in hippocampus and cerebral cortex of RS group were significantly higher than that of control at 1.5h,3h,6h and 24h(p<0.05).The level of Beclin1 protein in hippocampus of 3-MA group was significantly lower than that of RS group(p<0.05).The level of Beclin1 protein in cerebral cortex of 3-MA group was significantly lower than that of RS group(p<0.05).2.Morris water maze test (1) The average latencies of all rats were reduced by degree from D1 to D5.The escape latency was significantly longer in rats of RS group than that of the control at D5(p<0.05).The escape latency was significantly shorter in rats of 3-MA group than that of RS group at D5(p<0.05).However,there were no significantly difference between 3-MA and control groups (p>0.05).(2) As far as search strategy is concerned ,rats of RS group performed worse than those in control and 3-MA groups at D5,with statistically significant difference (p<0.05). However,the performance of rats between the 3-MA and control groups had no significantly difference.(3) The frequency of passing through the platform quadrant in rats had no significantly difference among the three groups(p>0.05).Conclusion: 1. The protein level of Beclin1 and LC3 in hippocampus and cerebral cortex were significantly increased in the early period after flurothyl-induced neonatal seizures which could cause long-term effects on learning ability.2. Autophagy inhibitor 3-MA that was used before seizures could significantly reduced the increased expression of LC3 and Beclin1 in hippocampus and cerebral cortex.Moreover ,it could significantly improve the learning ability following recurrent neonatal seizure-induced cognitive deficit.