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目的探讨钙蛋白酶拮抗剂Calpeptin对锰干扰突触体囊泡融合的保护作用。方法将24只小鼠随机分成4组,每组6只。对照组(腹腔注射0.9%氯化钠),低、高剂量染锰组(腹腔注射25μmol/kg、100μmol/kg氯化锰),Calpeptin预处理组(皮下注射Calpeptin 100μg/kg,30 min后腹腔注射100μmol/kg氯化锰),注射容量为2 ml/kg,每周5次,共4周。处死小鼠后分离基底核,制备突触体,测量小鼠基底核内锰含量,神经细胞内钙离子浓度和钙蛋白活力,突触体SNARE复合物及其相关蛋白的变化和囊泡融合情况。结果与对照组比较,高剂量染锰组小鼠基底核锰含量,神经细胞内钙离子浓度和钙蛋白酶活力显著增加;SNAP25蛋白表达下降,并出现裂解碎片,SNARE复合物形成减少,FM1-43荧光强度的下降;Calpeptin预处理可以明显抑制神经细胞内钙蛋白酶活力,缓解钙蛋白酶对SNAP25的裂解,100 k Da SNARE复合物也明显增多,而且SNARE复合物介导的突触囊泡融合有所上升。结论钙蛋白酶拮抗剂Calpeptin可以对锰干扰突触体囊泡融合起到有效的保护作用。
Objective To investigate the protective effect of calpain antagonist calpeptin on manganese interference synaptosome vesicle fusion. Methods Twenty-four mice were randomly divided into 4 groups with 6 mice in each group. The rats in the control group received intraperitoneal injection of 0.9% sodium chloride, and those in the low and high dose groups received intraperitoneal injection of 25 μmol / kg and 100 μmol / kg manganese chloride. Calpeptin preconditioning group (subcutaneous injection of Calpeptin 100 μg / kg, Injection of 100 μmol / kg manganese chloride), injection volume of 2 ml / kg, 5 times a week for 4 weeks. After the mice were sacrificed, the basal ganglia were separated and the synaptosomes were prepared. The contents of manganese in basal ganglia, intracellular calcium concentration and calpain activity, changes of synaptic SNARE complex and related proteins and vesicle fusion were measured . Results Compared with the control group, the levels of Mn in basal ganglia, intracellular Ca2 + and calpain activity in the high dose Mn2 + -treated group were significantly increased. The expression of SNAP25 protein was decreased and the fragmentation was observed. The formation of SNARE complex was reduced. The expression of FM1-43 Fluorescence intensity decreased; Calpeptin pretreatment can significantly inhibit the activity of calpain in nerve cells to ease calpain SNAP25 cleavage, 100 kDa SNARE complex also increased significantly, and SNARE complex-mediated synaptic vesicle fusion rise. Conclusions Calpain antagonist calpeptin can effectively protect manganese from interfering synaptosomal vesicle fusion.