Ravikumar Manish, Durairaj Brindha, Uvarajan Deenathayalan
Department of Biochemistry, PSG College of Arts & Science, Coimbatore, Tamil Nadu, India.
Mol Neurobiol. 2025 Apr;62(4):4534-4544. doi: 10.1007/s12035-024-04571-0. Epub 2024 Oct 28.
Epilepsy is a common neurological disorder affecting around 70 million people worldwide. Despite significant research and advancements in pharmaceutical therapies, the exact mechanisms underlying epileptogenesis remain unclear. As a result, current antiepileptic drug treatments are ineffective for approximately 30% of patients, providing only symptomatic relief. The epileptic process is influenced by the signaling of tumor necrosis factor alpha (TNFα), which affects neuronal excitability. The TNFα/TNFR1 signaling pathway and the role of RIPK1 in initiating inflammatory cell death pathways are well studied in human disorders. Dysregulation of RIPK1 is linked to inflammation and neurodegenerative diseases. Necrostatin-1 (Nec-1) selectively inhibits RIPK1 in various pathological conditions. The current study aimed to investigate the anticonvulsant properties of Nec-1 (a selective RIPK1 inhibitor) in PTZ-induced seizures in zebrafish larvae. Before the onset of seizures, zebrafish were treated with Nec-1 (15 µM) for 24 h at 6 days post-fertilization (dpf), followed by exposure to 15 mM of PTZ for 30 min. Behavioral assessments were conducted to observe changes in locomotor activity. Additionally, c-Fos expression was measured as an indicator of neuronal activation and analyzed mRNA levels of the astrocyte activation marker (GFAP) along with various inflammatory cytokines. Western blot analysis was conducted to evaluate GABA receptor expression. Pretreatment with Nec-1 restored normal behavior and reversed the expression of c-Fos in zebrafish larvae induced by PTZ. Additionally, Nec-1 reduced the elevated mRNA expression of inflammatory cytokines and significantly suppressed TNF/TNFR1 signaling, thereby inhibiting the enhanced internalization of GABA receptors. Our findings indicate that Nec-1 reduced the severity of PTZ-induced seizures in zebrafish by regulating behavior changes, suppressing inflammatory mediators, and enhancing the expression of GABA receptors, suggesting potential anticonvulsant properties.
癫痫是一种常见的神经系统疾病，全球约有7000万人受其影响。尽管在药物治疗方面进行了大量研究并取得了进展，但癫痫发生的确切机制仍不清楚。因此，目前的抗癫痫药物治疗对约30%的患者无效，只能提供症状缓解。癫痫过程受肿瘤坏死因子α（TNFα）信号传导的影响，TNFα会影响神经元兴奋性。TNFα/TNFR1信号通路以及RIPK1在启动炎症细胞死亡途径中的作用在人类疾病中得到了充分研究。RIPK1的失调与炎症和神经退行性疾病有关。Necrostatin-1（Nec-1）在各种病理条件下选择性抑制RIPK1。本研究旨在探讨Nec-1（一种选择性RIPK1抑制剂）对斑马鱼幼体戊四氮诱导癫痫发作的抗惊厥特性。在癫痫发作开始前，在受精后6天（dpf）用Nec-1（15µM）处理斑马鱼24小时，然后暴露于15mM戊四氮中30分钟。进行行为评估以观察运动活动的变化。此外，测量c-Fos表达作为神经元激活的指标，并分析星形胶质细胞激活标志物（GFAP）以及各种炎症细胞因子的mRNA水平。进行蛋白质免疫印迹分析以评估GABA受体表达。用Nec-1预处理可恢复斑马鱼幼体的正常行为，并逆转由戊四氮诱导的c-Fos表达。此外，Nec-1降低了炎症细胞因子升高的mRNA表达，并显著抑制TNF/TNFR1信号传导，从而抑制GABA受体增强的内化。我们的研究结果表明，Nec-1通过调节行为变化、抑制炎症介质和增强GABA受体表达来降低斑马鱼中戊四氮诱导癫痫发作的严重程度，提示其具有潜在的抗惊厥特性。
