Yokota M, Tani E, Tsubuki S, Yamaura I, Nakagaki I, Hori S, Saido T C
Department of Neurosurgery, Hyogo College of Medicine, Mukogawacho 1-1, Nishinomiya, Hyogo 663, Japan.
Brain Res. 1999 Feb 20;819(1-2):8-14. doi: 10.1016/s0006-8993(98)01334-1.
Transient forebrain ischemia induces activation of calpain and proteolysis of a neuronal cytoskeleton, fodrin, in gerbil hippocampus. This phenomenon precedes delayed neuronal death in hippocampal CA1 neurons. We examined effects of a calpain inhibitor on delayed neuronal death after transient forebrain ischemia. In gerbils, a selective calpain inhibitor entrapped in liposome was given transvenously and 30 min later, 5-min forebrain ischemia was produced by occlusion of both common carotid arteries. On day 7, CA1 neuronal damage was examined in the hippocampal slices stained with cresyl violet. Calpain-induced proteolysis of fodrin was also examined by immunohistochemistry and immunoblot. Additionally, to assure entrapment of the inhibitor by CA1 neurons, the inhibitor-liposome complex was labeled with FITC and given to gerbils. Fluorescence in the hippocampal slices was examined by confocal laser scanning microscope. Selective CA1 neuronal damage induced by forebrain ischemia was prevented by administration of the inhibitor in a dose-dependent manner. Calpain-induced proteolysis of fodrin was also extinguished by the calpain inhibitor in a dose-dependent manner. Bright fluorescence of the FITC-labeled inhibitor was observed in the CA1 neurons. The data show an important role of calpain in the development of the ischemic delayed neuronal death. Calpain seems to produce neuronal damage by degrading neuronal cytoskeleton. Our data also show a palliative effect of the calpain inhibitor on the neurotoxic damage, which offers a new and potent treatment of transient forebrain cerebral ischemia.
短暂性前脑缺血可诱导沙鼠海马体中钙蛋白酶的激活以及神经元细胞骨架蛋白（血影蛋白）的蛋白水解。这种现象先于海马体CA1神经元的迟发性神经元死亡出现。我们研究了一种钙蛋白酶抑制剂对短暂性前脑缺血后迟发性神经元死亡的影响。在沙鼠中，静脉注射包裹在脂质体中的选择性钙蛋白酶抑制剂，30分钟后，通过结扎双侧颈总动脉造成5分钟的前脑缺血。在第7天，对经甲酚紫染色的海马体切片进行CA1神经元损伤检查。还通过免疫组织化学和免疫印迹法检测钙蛋白酶诱导的血影蛋白水解情况。此外，为确保CA1神经元捕获抑制剂，将抑制剂-脂质体复合物用异硫氰酸荧光素（FITC）标记后给予沙鼠。通过共聚焦激光扫描显微镜检查海马体切片中的荧光。前脑缺血诱导的选择性CA1神经元损伤通过给予抑制剂以剂量依赖的方式得到预防。钙蛋白酶抑制剂也以剂量依赖的方式消除了钙蛋白酶诱导的血影蛋白水解。在CA1神经元中观察到FITC标记的抑制剂发出明亮荧光。数据表明钙蛋白酶在缺血性迟发性神经元死亡的发生过程中起重要作用。钙蛋白酶似乎通过降解神经元细胞骨架来造成神经元损伤。我们的数据还显示了钙蛋白酶抑制剂对神经毒性损伤的缓解作用，这为短暂性前脑缺血提供了一种新的有效治疗方法。
