Expression of mRNAs encoding subunits of the N-methyl-D-aspartate receptor in cultured cortical neurons.
Expression of mRNAs encoding subunits of the N-methyl-D-aspartate receptor in cultured cortical neurons.
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发表时间：
1994-05
影响因子：
3.6
通讯作者：
J. Zhong;S. Russell;D. B. Pritchett;P. Molinoff;K. Williams
中科院分区：
文献类型：
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作者：
J. Zhong;S. Russell;D. B. Pritchett;P. Molinoff;K. Williams
在原代培养的皮层神经元中检测了编码n -甲基- d -天冬氨酸（NMDA）受体亚基的mrna的表达。从胚胎第17天的大鼠新皮层制备培养物。在这个发育年龄，NR1、NR2A、NR2B和NR2C mRNA的水平很低或检测不到。NR1 mRNA的表达在第1 ~ 21天逐渐升高。NR2A mRNA的表达量在第1 ~ 7天没有变化，但在第7 ~ 21天有所增加。相比之下，NR2B mRNA水平在第1天至第7天之间升高，第7天之后几乎没有进一步的变化。NR2B mRNA水平比NR2A mRNA水平高出约4倍。通过配体结合试验，还发现随着培养时间的推移，对伊芬地尔具有低亲和力的NMDA受体的比例也在增加。将细胞维持在低浓度血清培养基中，可以减少或阻止对伊芬普罗地尔低亲和力受体表达的增加以及NR1和NR2A mrna的增加。结果与NR2A亚基在天然NMDA受体中的包含是其对伊芬普罗地尔低亲和力的原因的假设是一致的。在培养的神经元中，缺乏5'（氨基末端）插入的NR1的剪接变体被发现是NR1的主要形式。含有5'插入的变异只占NR1 mRNA总数的一小部分（<或= 5%），并且它们的比例不随培养时间的变化而改变。培养神经元中NMDA受体特性和亚基mRNA表达的时间依赖性变化与发育中的大鼠脑中观察到的变化相似。因此，体内发生的NMDA受体表达的发育序列部分保留在体外维持的神经元中。
The expression of mRNAs encoding subunits of the N-methyl-D-aspartate (NMDA) receptor was examined in cortical neurons maintained in primary culture. Cultures were prepared from embryonic day 17 rat neocortex. At this developmental age, levels of NR1, NR2A, NR2B, and NR2C mRNA were low or undetectable. Expression of NR1 mRNA increased progressively between days 1 and 21 in vitro. The amount of NR2A mRNA did not change between days 1 and 7 but increased between days 7 and 21. In contrast, levels of NR2B mRNA increased between days 1 and 7, with little further change after day 7. The level of NR2B mRNA was approximately 4-fold higher than that of NR2A mRNA in 21-day cultures. Using ligand binding assays, the proportion of NMDA receptors having a low affinity for ifenprodil was also found to increase over time in culture. The increase in the expression of receptors having a low affinity for ifenprodil and the increase in NR1 and NR2A mRNAs were reduced or prevented by maintaining cells in medium with a low concentration of serum. The results are consistent with the hypothesis that inclusion of the NR2A subunit in native NMDA receptors is responsible for their low affinity for ifenprodil. Splice variants of NR1 lacking the 5' (amino-terminal) insert were found to be the predominant forms of NR1 in cultured neurons. Variants containing the 5' insert represented only a small (< or = 5%) fraction of total NR1 mRNA, and their proportion was not altered as a function of time in culture. Time-dependent changes in the properties of NMDA receptors and in the expression of subunit mRNA occurring in cultured neurons are similar to changes observed in developing rat brain. Thus, the developmental sequence of NMDA receptor expression that occurs in vivo is partially retained in neurons maintained in vitro.