The 71 kDa glutamate-binding protein is increased in cerebellar granule cells after chronic ethanol treatment
The 71 kDa glutamate-binding protein is increased in cerebellar granule cells after chronic ethanol treatment
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DOI：
10.1016/0169-328x(96)00021-6
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发表时间：
1996-07-01
期刊：
影响因子：
--
通讯作者：
Michaelis, EK
中科院分区：
文献类型：
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作者：
Hoffman, PL;Bhave, SV;Michaelis, EK
除了N-甲基-D-天冬氨酸(NMDA)受体蛋白NR1和NR2外，小脑颗粒细胞还表达另一种蛋白质复合体，该复合体含有NMDA受体特有的配体结合部位。后一种复合体中的蛋白质之一是71 kDa的谷氨酸结合蛋白(GBP)。为了确定GBP在NMDA反应中的作用，用与该蛋白的mRNA互补的反义寡核苷酸处理原代培养的小脑颗粒细胞。这种处理大大降低了GBP的mRNA和蛋白质水平，以及细胞对NMDA的反应，通过Fura-2荧光测量细胞内钙离子的增加。反义寡核苷酸处理不改变钙离子对KCI或海人藻酸的反应。先前研究表明，慢性酒精暴露可增加NMDA受体功能，并增加小脑颗粒细胞中NMDA受体通道阻滞剂地佐西平的结合部位密度。现在显示，细胞长期暴露在100 mM乙醇中，GBP的mRNA和蛋白质水平显著增加(分别为45%和100%)。乙醇处理不影响NR1或NR2A的mRNA水平，仅引起NR1蛋白水平的小幅上升(20%)，而导致NR2A蛋白的下降(30%)。虽然不能排除NMDA受体NR1/NR2亚基的作用，但这些结果与GBP参与慢性乙醇诱导的小脑颗粒细胞NMDA受体功能增强的假说是一致的。
Besides the N-methyl-D-aspartate (NMDA) receptor proteins NR1 and NR2, another complex of proteins which has been shown to contain ligand-binding sites characteristic of NMDA receptors is expressed in cerebellar granule cells. One of the proteins in the latter complex is the 71 kDa glutamate-binding protein (GBP). To determine the role of the GBP in the response to NMDA, primary cultures of cerebellar granule cells were treated with an antisense oligonucleotide complementary to mRNA for this protein. This treatment substantially reduced both mRNA and protein levels of the GBP, as well as the response of the cells to NMDA, measured as an increase in intracellular Ca2+ with fura-2 fluorescence. The antisense oligonucleotide treatment did not alter the Ca2+ responses to KCI or kainate. Chronic ethanol exposure has previously been shown to increase NMDA receptor function and the density of binding sites for the NMDA receptor channel blocker, dizocilpine, in cerebellar granule cells. Chronic exposure of the cells to 100 mM ethanol is now shown to result in significant increases in mRNA and protein levels for the GBP (45% and 100%, respectively). Ethanol treatment did not affect mRNA levels for NR1 or NR2A, caused only a small increase (20%) in protein levels for NR1, and resulted in a decrease (30%) in NR2A protein. Although a role of the NMDA receptor NR1/NR2 subunits cannot be ruled out, these results are compatible with the hypothesis of involvement of the GBP in the chronic ethanol-induced increase in NMDA receptor function in cerebellar, granule cells.