Early exposure of cultured hippocampal neurons to excitatory amino acids protects from later excitotoxicity
Early exposure of cultured hippocampal neurons to excitatory amino acids protects from later excitotoxicity
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DOI：
10.1016/j.ijdevneu.2009.11.002
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发表时间：
2010-04-01
影响因子：
1.8
通讯作者：
Segal, Menahem
中科院分区：
文献类型：
--
作者：
Friedman, Linda K.;Segal, Menahem
癫痫持续状态发生在出生后发育早期，保护CA1海马神经元，这是发育中的大脑中对癫痫诱导的损伤最敏感的区域。在此，我们在大鼠海马区的分离培养中建立了“两次打击”模型，以测试在相对抵抗期预先暴露于高浓度谷氨酸、N-甲基-D-天冬氨酸或alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic酸是否能防止神经元在神经元成熟后再次暴露于相同的化学物质后死亡，并变得非常容易受到兴奋性氨基酸的影响。不同剂量的谷氨酸、NMDA或AMPA在5DIV作用48h，再在14DIV作用5、15或30min。Neun免疫组织化学显示，早期暴露于谷氨酸(500 MU M)导致大约一半的神经元死亡(52+/-8.6%)，而一次暴露于14DIV后出现明显的枯竭(98+/-0.79%)。先用中剂量谷氨酸(200mM)刺激培养，7d后再用高剂量谷氨酸刺激，大量神经元存活下来(35.3+/-1.2%)。同样，预先暴露于最大剂量的NMDA(100微米)增加了第二次挑战后存活细胞的比例。相比之下，AMPA(100亩M)在早期或晚期施药后毒性相同，对第二次暴露没有保护作用。200 p，M谷氨酸染毒48h后，GluR1亚基表达明显下降(分别下降44.57+/-3.6%和45.07+/-3.69%)，而GluR2亚基表达下降较少(25.7+/-3.8%)。共聚焦显微镜显示，一次或两次NMDA暴露导致GluR2蛋白进一步下调，而相同神经元中的小白蛋白(PV)显着增加四倍以上。另一方面，首次暴露于500亩M谷氨酸后，钙结合蛋白(CB)免疫反应几乎消失。这些数据表明，早期、短暂地暴露于某些高剂量的EAA可以诱导长期的神经保护。GluR1/GluR2比率的改变以及特定钙结合蛋白的差异表达可能有助于这种神经保护。由爱思唯尔爱尔兰有限公司代表ISDN出版。
Status epilepticus occurring in early postnatal development protects CA1 hippocampal neurons, the region most sensitive to seizure-induced injury in the developing brain. Here, we developed a "two hit" model in dissociated cultures of the rat hippocampus to test whether pre-exposure of immature neurons to high concentrations of glutamate, N-methyl-D-aspartic acid (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) during a relatively resistant period prevents neurons from dying following a second exposure to the same chemicals after neurons mature and become highly vulnerable to excitatory amino acids (EAAs). Cultures were exposed to varied doses of glutamate, NMDA, or AMPA for 48 h at 5 DIV and again at 14 DIV for 5, 15, or 30 min. NeuN immunohistochemistry showed early exposure to glutamate (500 mu M) killed approximately half of the neurons (52 +/- 8.6%) compared to the marked depletion that occurs after one exposure at 14 DIV (98 +/- 0.79%). When cultures were first challenged with moderate doses of glutamate (200 mu M) followed by the high dose 7 days later, a significant population of neurons was spared (35.3 +/- 1.2%). Similarly, pre-exposure to maximal doses of NMDA (100 mu M) increased the proportion of surviving cells following the second challenge. In contrast, AMPA (100 mu M) was equally toxic after early or late applications and did not protect from the second exposure. GluR1 subunit expression was markedly decreased at 48 h after one or two exposures to 200 p,M glutamate (by 44.57 +/- 3.6%, 45.07 +/- 3.69%) whereas GluR2 subunit expression was reduced by a lesser amount (25.7 57 +/- 3.8%). Confocal microscopy showed that one or two exposures to NMDA caused GluR2 protein to downregulate even further whereas parvalbumin (PV) was dramatically increased in the same neurons by over four-fold. On the other hand, calbindin (CB) immunoreactivity was nearly absent after the first exposure to 500 mu M glutamate. These data indicate that early, transient exposure to certain EAAs at high doses can induce long-lasting neuroprotection. Alterations in the GluR1/GluR2 ratio as well as differential expression of specific calcium binding proteins may contribute to this neuroprotection. Published by Elsevier Ireland Ltd on behalf of ISDN.