Rapid subcellular redistribution of Bax precedes caspase-3 and endonuclease activation during excitotoxic neuronal apoptosis in rat brain
Rapid subcellular redistribution of Bax precedes caspase-3 and endonuclease activation during excitotoxic neuronal apoptosis in rat brain
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DOI：
10.1089/08977150260190410
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发表时间：
2002-07-01
影响因子：
4.2
通讯作者：
Martin, LJ
中科院分区：
文献类型：
--
作者：
Lok, J;Martin, LJ
神经元凋亡是由谷氨酸受体兴奋性毒性和相关损伤（包括创伤和缺氧缺血）在新生啮齿动物脑中显著诱导的。然而，这种神经变性的分子机制尚不清楚。我们测试了这一假设，即在新生大鼠纹状体兴奋性神经元凋亡的进展过程中，促凋亡蛋白Bax的亚细胞分布的变化先于下游凋亡效应机制的激活，如caspase-3裂解和核酸内切酶激活。将红藻氨酸（4 nmol）注入麻醉的7日龄大鼠纹状体，并在损伤后2、6、12和24 h处死动物。对照组为年龄匹配、注射溶媒或未处理大鼠。脑切片中经超微结构证实的纹状体神经元凋亡计数在24 h最高。纹状体组织进行显微解剖和分馏成胞质，线粒体，和核丰富的车厢。免疫印迹表明，Bax易位从胞质组分的线粒体组分，最大易位2小时的情况下，线粒体积累的变化。在6和24 h之间，在胞质和线粒体组分中裂解的caspase-3水平逐渐增加。如免疫定位所示，裂解的caspase-3在凋亡的纹状体神经元中积累。DNA的核粒间片段化与半胱天冬酶-3切割一致。我们的结论是，Bax的快速易位到线粒体前半胱天冬酶-3和核酸内切酶激活在新生大鼠脑兴奋性毒性神经元凋亡，并在谷氨酸受体激活后2小时内发生这种死亡级联反应的启动。
Neuronal apoptosis is induced prominently in the newborn rodent brain by glutamate receptor excitotoxicity and related insults, including trauma and hypoxia-ischemia. However, the molecular mechanisms of this neurodegeneration are unclear. We tested the hypothesis that changes in the subcellular distribution of the proapoptotic protein Bax precede the activation of downstream apoptosis-effector mechanisms such as caspase-3 cleavage and endonuclease activation during the progression of excitotoxic neuronal apoptosis in the striatum of newborn rat. Kainic acid (4 nmol) was injected into striatum of anesthetized 7-day-old rats, and the animals were killed at 2, 6, 12, and 24 h postinsult. Controls were age-matched, vehicle-injected, or naive rats. Counts of ultrastructurally confirmed striatal neuron apoptosis in brain sections were highest at 24 h. Striatal tissue was microdissected and fractionated into cytosolic, mitochondrial-, and nuclear-enriched compartments. Immunoblots showed that Bax translocates from the cytosol fraction to the mitochondrial fraction, with maximal translocation by 2 h in the absence of changes in mitochondrial accumulation. Cleaved caspase-3 levels increase progressively in both cytosolic and mitochondrial fractions between 6 and 24 h. Cleaved caspase-3 accumulates in apoptotic striatal neurons as shown by immunolocalization. Internucleosornal fragmentation of DNA coincides with caspase-3 cleavage. We conclude that rapid translocation of Bax to mitochondria precedes caspase-3 and endonuclease activation during excitotoxic neuronal apoptosis in newborn rat brain and that initiation of this death cascade occurs within 2 h after glutamate receptor activation.