Audinat E, Lambolez B, Rossier J, Crépel F
Laboratoire de Neurobiologie et Neuropharmacologie du Développement, Centre National de la Recherche Scientifique, Université Paris-Sud, Orsay, France.
Eur J Neurosci. 1994 Dec 1;6(12):1792-800. doi: 10.1111/j.1460-9568.1994.tb00572.x.
The glutamate receptor channels of the N-methyl-D-aspartate (NMDA) subtype are composed of different subunits named NR1 and NR2A-D. These subunits can combine in different oligomers with diverging properties and their expression is developmentally regulated. We have used rat cerebellar slice cultures to test the involvement of bioelectrical activity and synaptic transmission in the changes in NR2A-C expression observed in developing granule cells. A correlation between the functional properties of the NMDA receptors and expression of the NR2A-C mRNAs was obtained in single granule cells by coupling patch-clamp recording and reverse transcription followed by polymerase chain reaction. Granule cells grown under standard culture conditions expressed mainly NR2A mRNA when examined after 15-40 days in vitro. Consistent with this observation, their responses to NMDA were only weakly reduced by 3 microM ifenprodil, a non-competitive antagonist which discriminates between NR2A and NR2B subunits in expression systems. In cerebellar cultures chronically exposed to tetrodotoxin to eliminate spontaneous electrical activity, granule cells maintained a predominant expression of NR2B subunits and their responses to NMDA were largely inhibited by 3 microM ifenprodil. These results provide evidence that the expression of the NR2A and B subunits is regulated through an activity-dependent mechanism leading to the formation of NMDA receptors with different pharmacological properties. Finally, the NR2C subunit, abundantly expressed in vivo by adult granule cells, was only rarely detected in slice cultures, even when excitatory synapses were formed between granule cells and fibres originating from co-cultured brainstem explants. These data suggest that the induction of NR2C expression observed in vivo requires an additional factor(s) that remains to be identified.
N-甲基-D-天冬氨酸（NMDA）亚型的谷氨酸受体通道由名为NR1和NR2A-D的不同亚基组成。这些亚基可以组合成具有不同特性的不同寡聚体，并且它们的表达受发育调控。我们使用大鼠小脑切片培养物来测试生物电活动和突触传递在发育中的颗粒细胞中观察到的NR2A-C表达变化中的作用。通过结合膜片钳记录和逆转录随后进行聚合酶链反应，在单个颗粒细胞中获得了NMDA受体的功能特性与NR2A-C mRNA表达之间的相关性。在体外培养15 - 40天后检查时，在标准培养条件下生长的颗粒细胞主要表达NR2A mRNA。与该观察结果一致，它们对NMDA的反应仅被3 microMifenprodil（一种在表达系统中区分NR2A和NR2B亚基的非竞争性拮抗剂）微弱降低。在长期暴露于河豚毒素以消除自发电活动的小脑培养物中，颗粒细胞维持NR2B亚基的主要表达，并且它们对NMDA的反应被3 microMifenprodil大大抑制。这些结果提供了证据，表明NR2A和B亚基的表达通过一种活性依赖机制进行调节，导致形成具有不同药理学特性的NMDA受体。最后，NR2C亚基在成年颗粒细胞体内大量表达，但即使在颗粒细胞与来自共培养脑干外植体的纤维之间形成兴奋性突触时，在切片培养物中也很少检测到。这些数据表明，在体内观察到的NR2C表达的诱导需要一个有待确定的额外因素。
