Recent studies have shown that exposure to sevoflurane in developing brains causes neuronal apoptosis and cognitive dysfunction. “Necroptosis” is a novel pathway of necrosis. We introduced the caspase-specific inhibitor Z-VAD in addition to the receptor-interacting protein kinase 1 (RIPK1) inhibitor Nec-1, to ascertain the existence and importance of necroptosis. Sprague–Dawley rat pups postnatal day 7 were randomly assigned into one of five groups: control, sevoflurane + Z-VAD, sevoflurane + Nec-1, sevoflurane + Z-VAD + Nec-1 and 3% sevoflurane group. Neuronal apoptosis was evaluated by hematoxylin and eosin staining. The MTT assay was performed to evaluate cell viability. Immunofluorescence was employed to measure expression of RIPK1 and RIPK3. Western blots showing expression of RIPK1, RIPK3 and phosphorylation of mixed lineage kinase domain-like (p-MLKL) were used to explore the role of necroptosis. Binding of RIPK1/RIPK3 was detected via co-immunoprecipitation. Finally, the Morris water maze test was used to determine cognitive function. Exposure to 3% sevoflurane for 6 h induced neurotoxicity and inhibited cell viability. Neuron viability was low in the SEV, SEV + Z-VAD and SEV + Nec-1 groups. The study revealed that RIPK1 and RIPK3 protein expression increased significantly, but there was no significant differences between the SEV and SEV + Z-VAD groups. The expression of p-MLKL significantly increased in the SEV and SEV + Z-VAD groups, but not in the SEV + Nec-1 group or SEV + Z-VAD + Nec-1 group compared to the control group. Co-immunoprecipitation results showed that sevoflurane exposure enhanced binding of RIPK1/RIPK3 protein significantly. Blockade of apoptosis and necroptosis alleviated sevoflurane-induced cognitive impairment. Sevoflurane exposure elicited neurotoxicity within neonatal hippocampal neurons and tissues. Blockade of apoptosis or necroptosis alone did not attenuate sevoflurane-induced neurotoxicity (SIN). RIPK1/RIPK3-mediated necroptosis was involved in SIN in hippocampal neurons. SIN could be attenuated only by inhibiting both apoptosis and necroptosis.

最近的研究表明，发育中的大脑接触七氟烷会导致神经元凋亡和认知功能障碍。“坏死性凋亡”是一种新的坏死途径。除了受体相互作用蛋白激酶 1 (RIPK1) 抑制剂 Nec-1 之外，我们还引入了 caspase 特异性抑制剂 Z-VAD，以确定坏死性凋亡的存在和重要性。出生后第 7 天的 Sprague-Dawley 大鼠幼崽被随机分为五组之一：对照组、七氟烷 + Z-VAD、七氟烷 + Nec-1、七氟烷 + Z-VAD + Nec-1 和 3% 七氟烷组。通过苏木精和伊红染色评估神经元凋亡。进行MTT测定以评估细胞活力。采用免疫荧光法测定 RIPK1 和 RIPK3 的表达。使用显示 RIPK1、RIPK3 表达和混合谱系激酶结构域样 (p-MLKL) 磷酸化的蛋白质印迹来探讨坏死性凋亡的作用。通过免疫共沉淀检测 RIPK1/RIPK3 的结合。最后，使用莫里斯水迷宫测试来确定认知功能。暴露于 3% 七氟醚 6 小时会诱导神经毒性并抑制细胞活力。SEV、SEV + Z-VAD 和 SEV + Nec-1 组的神经元活力较低。研究显示，RIPK1和RIPK3蛋白表达显着增加，但SEV组和SEV+Z-VAD组之间没有显着差异。与对照组相比，SEV和SEV+Z-VAD组中p-MLKL的表达显着增加，但SEV+Nec-1组或SEV+Z-VAD+Nec-1组中p-MLKL的表达不显着增加。免疫共沉淀结果显示七氟醚暴露显着增强了 RIPK1/RIPK3 蛋白的结合。阻断细胞凋亡和坏死性凋亡可减轻七氟烷引起的认知障碍。七氟醚暴露会引起新生儿海马神经元和组织内的神经毒性。单独阻断细胞凋亡或坏死性凋亡并不能减轻七氟烷诱导的神经毒性（SIN）。RIPK1/RIPK3 介导的坏死性凋亡参与海马神经元的 SIN。SIN只能通过抑制细胞凋亡和坏死性凋亡来减弱。