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首页> 外文期刊>Neurochemistry International: The International Journal for the Rapid Publication of Critical Reviews, Preliminary and Original Research Communications in Neurochemistry >Developmental expression and activity of high affinity glutamate transporters in rat cortical primary cultures.
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Developmental expression and activity of high affinity glutamate transporters in rat cortical primary cultures.

机译：高亲和力谷氨酸转运蛋白在大鼠皮层原代培养物中的发育表达和活性。

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摘要

The expression and activity of glutamate transporters (EAAC1, GLAST and GLT1) were examined during the development of cortical neuron-enriched cultures. Protein content and mitochondrial respiration both increased during the first 7 days, later stabilized and decreased from DIV14. Glutamate transport and extracellular concentration were relatively constant from DIV3 to 18. The kinetic parameters of glutamate transport were at DIV7: K(m)=19+/-3 microM and V(max)=1068+/-83 pmol/mg protein/min and at DIV14: K(m)=40.8+/-9.3 microM and V(max)=1060+/-235 pmol/mg protein/min. The shift in K(m) towards higher values suggest a more important participation of GLAST after DIV14. At DIV7 and 14, glutamate transport was poorly sensitive to dihydrokainate (DHK) suggesting a weak participation of GLT1 in glutamate transport. Western blot experiments and immunocytochemistry showed that EAAC1 was expressed by neurons whatever the stage of the culture. GLAST was found in astrocytes as soon as DIV3 and labeling increased during the development of the culture. There was little neuronal GLT1 immunoreactivity at DIV7, only detected by immunocytochemistry. From DIV10 to 18, an increasing astrocytic expression of GLT1 was observed, also detected by Western blotting. These results show that: (1) glutamate uptake remains stable all along the development of the cultures although the pattern of expression of the different transporters is changing, suggesting that glutamate transport is highly regulated; (2) neuronal EAAC1 may play a critical role during the early stages of the culture when it is expressed alone; and (3) the developmental expression pattern of glutamate transporters in cortical neuron-enriched cultures is quite similar to that observed in vivo during early postnatal development.
机译：在皮质神经元富集培养的发展过程中检查了谷氨酸转运蛋白（EAAC1，GLAST和GLT1）的表达和活性。蛋白质含量和线粒体呼吸在前7天均增加，随后稳定下来，并从DIV14开始下降。谷氨酸转运和细胞外浓度从DIV3到18相对恒定。谷氨酸转运的动力学参数为DIV7：K（m）= 19 +/- 3 microM和V（max）= 1068 +/- 83 pmol / mg蛋白质/最小和DIV14：K（m）= 40.8 +/- 9.3 microM和V（max）= 1060 +/- 235 pmol / mg蛋白质/分钟。 K（m）向更高值的转变表明DIV14之后GLAST的参与更为重要。在DIV7和14，谷氨酸转运对二氢海藻酸盐（DHK）的敏感性较弱，表明GLT1对谷氨酸转运的参与较弱。 Western印迹实验和免疫细胞化学表明，EAAC1在培养的任何阶段均由神经元表达。在培养过程中，DIV3和标记增加后，星形胶质细胞中就会发现GLAST。只有通过免疫细胞化学检测到，DIV7的神经元GLT1免疫反应很少。从DIV10到18，还观察到GLT1星形细胞表达的增加，也通过Western blotting检测到。这些结果表明：（1）尽管不同转运蛋白的表达方式发生了变化，但谷氨酸的摄取在整个培养过程中仍保持稳定，这表明谷氨酸的转运受到高度调节。 （2）当单独表达时，神经元EAAC1可能在培养的早期阶段起关键作用; （3）富含皮质神经元的培养物中谷氨酸转运蛋白的发育表达模式与出生后早期体内观察到的十分相似。

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