Dysfunction of neuronal nitric oxide synthase contributes to neurotoxicity, which triggers cell death in various neuropathological diseases, including epilepsy. Studies have shown that inhibition of neuronal nitric oxide synthase activity increases the epilepsy threshold, that is, has an anticonvulsant effect. However, the exact role and potential mechanism of neuronal nitric oxide synthase in seizures are still unclear. In this study, we performed RNA sequencing, functional enrichment analysis, and weighted gene coexpression network analysis of the hippocampus of tremor rats, a rat model of genetic epilepsy. We found damaged hippocampal mitochondria and abnormal succinate dehydrogenase level and Na⁺-K⁺-ATPase activity. In addition, we used a pilocarpine-induced N2a cell model to mimic epileptic injury. After application of neuronal nitric oxide synthase inhibitor 7-nitroindazole, changes in malondialdehyde, lactate dehydrogenase and superoxide dismutase, which are associated with oxidative stress, were reversed, and the increase in reactive oxygen species level was reversed by 7-nitroindazole or reactive oxygen species inhibitor N-acetylcysteine. Application of 7-nitroindazole or N-acetylcysteine downregulated the expression of caspase-3 and cytochrome c and reversed the apoptosis of epileptic cells. Furthermore, 7-nitroindazole or N-acetylcysteine downregulated the abnormally high expression of NLRP3, gasdermin-D, interleukin-1β and interleukin-18. This indicated that 7-nitroindazole and N-acetylcysteine each reversed epileptic cell death. Taken together, our findings suggest that the neuronal nitric oxide synthase/reactive oxygen species pathway is involved in pyroptosis of epileptic cells, and inhibiting neuronal nitric oxide synthase activity or its induced oxidative stress may play a neuroprotective role in epilepsy.

神经元一氧化氮合酶的功能障碍会导致神经毒性，从而引发各种神经病理疾病（包括癫痫）中的细胞死亡。研究表明，抑制神经元一氧化氮合酶活性可提高癫痫阈值，即具有抗惊厥作用。然而，神经元一氧化氮合酶在癫痫发作中的确切作用和潜在机制仍不清楚。在这项研究中，我们对震颤大鼠（一种遗传性癫痫大鼠模型）的海马体进行了 RNA 测序、功能富集分析和加权基因共表达网络分析。我们发现受损的海马线粒体和异常的琥珀酸脱氢酶水平和 Na ⁺ -K ⁺-ATP酶活性。此外，我们使用毛果芸香碱诱导的 N2a 细胞模型来模拟癫痫损伤。应用神经元一氧化氮合酶抑制剂7-硝基吲唑后，与氧化应激相关的丙二醛、乳酸脱氢酶和超氧化物歧化酶的变化被逆转，活性氧水平的升高被7-硝基吲唑或活性氧逆转N-乙酰半胱氨酸抑制剂。应用7-硝基吲唑或N-乙酰半胱氨酸可下调caspase-3和细胞色素c的表达，逆转癫痫细胞的凋亡。此外，7-硝基吲唑或 N-乙酰半胱氨酸下调 NLRP3、gasdermin-D、interleukin-1β 和 interleukin-18 的异常高表达。这表明 7-硝基吲唑和 N-乙酰半胱氨酸各自逆转了癫痫细胞死亡。