Regulation of N-Methyl-D-aspartate Receptors by Calpain in Cortical Neurons*
Regulation of N-Methyl-D-aspartate Receptors by Calpain in Cortical Neurons*
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DOI：
10.1074/jbc.m501603200
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发表时间：
2005-06
影响因子：
4.8
通讯作者：
Hai‐yan Wu;E. Yuen;Yun-fei Lu;M. Matsushita;H. Matsui;Zhen Yan;K. Tomizawa
中科院分区：
文献类型：
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作者：
Hai‐yan Wu;E. Yuen;Yun-fei Lu;M. Matsushita;H. Matsui;Zhen Yan;K. Tomizawa
N-甲基-d-天冬氨酸 (NMDA) 受体是一种对钙具有高度渗透性的阳离子通道，在控制神经元的正常和病理功能中发挥着关键作用。钙通过 NMDA 受体 (NMDAR) 进入可导致 Ca2+ 依赖性蛋白酶、钙蛋白酶的激活。在这里，我们研究了钙蛋白酶在 NMDAR 通道功能调节中的参与。经过 NMDA 或谷氨酸长时间（5 分钟）处理后，在急性分离和培养的皮质锥体神经元中，全细胞 NMDAR 介导的电流显着降低。选择性钙蛋白酶抑制剂的浴应用可阻断 NMDAR 电流的下调。细胞内注射特定的钙蛋白酶抑制肽也消除了长期 NMDA 治疗引起的 NMDAR 电流下调。相反，动力抑制肽对 NMDAR 电流的抑制没有影响，表明动力/网格蛋白介导的 NMDAR 内化在此过程中不参与。免疫印迹分析表明，在用谷氨酸处理的培养的皮层神经元中，NMDAR 的 NR2A 和 NR2B 亚基明显降解，并且钙蛋白酶抑制剂可以阻止 NR2 亚基的降解。综上所述，我们的结果表明，神经元中 NMDAR 的长期激活会激活钙蛋白酶，而激活的钙蛋白酶又会下调 NMDAR 的功能，这提供了一种针对伴随缺血和中风的 NMDAR 过度刺激的神经保护机制。
The N-methyl-d-aspartate (NMDA) receptor is a cation channel highly permeable to calcium and plays critical roles in governing normal and pathologic functions in neurons. Calcium entry through NMDA receptors (NMDARs) can lead to the activation of the Ca2+-dependent protease, calpain. Here we investigated the involvement of calpain in regulation of NMDAR channel function. After prolonged (5-min) treatment with NMDA or glutamate, the whole-cell NMDAR-mediated current was significantly reduced in both acutely dissociated and cultured cortical pyramidal neurons. The down-regulation of NMDAR current was blocked by bath application of selective calpain inhibitors. Intracellular injection of a specific calpain inhibitory peptide also eliminated the down-regulation of NMDAR current induced by prolonged NMDA treatment. In contrast, dynamin inhibitory peptide had no effect on the depression of NMDAR current, suggesting the lack of involvement of dynamin/clathrin-mediated NMDAR internalization in this process. Immunoblotting analysis showed that the NR2A and NR2B subunits of NMDARs were markedly degraded in cultured cortical neurons treated with glutamate, and the degradation of NR2 subunits was prevented by calpain inhibitors. Taken together, our results suggest that prolonged activation of NMDARs in neurons activates calpain, and activated calpain in turn down-regulates the function of NMDARs, which provides a neuroprotective mechanism against NMDAR overstimulation accompanying ischemia and stroke.