Department of Anesthesiology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, 510120, China; Guangdong Province Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, 510120, China.
Department of Anesthesiology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, 510120, China.
Toxicol Lett. 2021 May 1;341:68-79. doi: 10.1016/j.toxlet.2021.01.023. Epub 2021 Feb 3.
General anesthetics such as sevoflurane interfere with dendritic development and synaptogenesis, resulting in cognitive impairment. The collapsin response mediator protein2 (CRMP2) plays important roles in dendritic development and synaptic plasticity and its phosphorylation is regulated by cycline dependent kinase-5 (Cdk5) and glycogen synthase kinase-3β (GSK-3β). Here we investigated whether Cdk5/CRMP2 or GSK-3β/CRMP2 pathway is involved in sevoflurane-induced developmental neurotoxicity.
Rats at postnatal day 7 (PND7) were i.p. injected with Cdk5 inhibitor roscovitine, GSK-3β inhibitor SB415286 or saline 20 min. before exposure to 2.8% sevoflurane for 4 h. Western-blotting was applied to measure the expression of Cdk5/CRMP2 and GSK-3β/CRMP2 pathway proteins in the hippocampus 6 h after the sevoflurane exposure. When rats grew to adolescence (from PND25), they were tested for open-field and contextual fear conditioning, and then long term potentiation (LTP) from hippocampal slices was recorded, and morphology of pyramidal neuron was examined by Golgi staining and synaptic plasticity-related proteins expression in hippocampus were measured by western-blotting. In another batch of experiment, siRNA-CRMP2 or vehicle control was injected into hippocampus on PND5.
Sevoflurane activated Cdk5/CRMP2 and GSK-3β/CRMP2 pathways in the hippocampus of neonatal rats, reduced dendritic length, branches and the density of dendritic spine in pyramidal neurons. It also reduced the expressions of PSD-95, drebrin and synaptophysin in hippocampus, impaired memory ability of rats and inhibited LTP in hippocampal slices. All the impairment effects by sevoflurane were attenuated by pretreatment with inhibitor of Cdk5 or GSK-3β. Furthermore, rat transfected with siRNA-CRMP2 eliminated the neuroprotective effects of Cdk5 or GSK-3β blocker in neurobehavioral and LTP tests.
Cdk5/CRMP2 and GSK-3β/CRMP2 pathways participate in sevoflurane-induced dendritic development abnormalities and cognitive dysfunction in developing rats.
七氟醚等全身麻醉药会干扰树突发育和突触形成，导致认知障碍。 collapsin 反应介质蛋白 2（CRMP2）在树突发育和突触可塑性中发挥重要作用，其磷酸化受细胞周期依赖性激酶 5（Cdk5）和糖原合成酶激酶 3β（GSK-3β）调节。本研究旨在探讨 Cdk5/CRMP2 或 GSK-3β/CRMP2 通路是否参与七氟醚诱导的发育性神经毒性。
7 日龄（PND7）大鼠腹腔注射 Cdk5 抑制剂 roscovitine、GSK-3β 抑制剂 SB415286 或生理盐水 20 min.，然后暴露于 2.8%七氟醚中 4 h。Western-blotting 用于测量七氟醚暴露后 6 h 海马中 Cdk5/CRMP2 和 GSK-3β/CRMP2 通路蛋白的表达。当大鼠进入青春期（PND25 时），进行旷场和情境恐惧条件反射测试，然后记录海马切片长时程增强（LTP），通过 Golgi 染色观察锥体神经元形态，Western-blotting 测量海马中突触可塑性相关蛋白的表达。在另一批实验中，PND5 时向海马内注射 siRNA-CRMP2 或载体对照。
七氟醚激活新生大鼠海马中的 Cdk5/CRMP2 和 GSK-3β/CRMP2 通路，减少锥体神经元树突长度、分支和树突棘密度。它还降低了海马中 PSD-95、drebrin 和突触小体蛋白的表达，损害了大鼠的记忆能力并抑制了海马切片中的 LTP。Cdk5 或 GSK-3β 抑制剂预处理可减轻七氟醚引起的所有损伤作用。此外，转染 siRNA-CRMP2 的大鼠消除了 Cdk5 或 GSK-3β 阻滞剂在神经行为和 LTP 测试中的神经保护作用。
Cdk5/CRMP2 和 GSK-3β/CRMP2 通路参与七氟醚诱导的发育期大鼠树突发育异常和认知功能障碍。
