Department of Physiology, School of Medicine, Teikyo University, Tokyo 173-8605, Japan.
Proc Natl Acad Sci U S A. 2010 Aug 24;107(34):15252-7. doi: 10.1073/pnas.0906551107. Epub 2010 Aug 9.
The GluN2B (GluRepsilon2/NR2B) and GluN2A (GluRepsilon1/NR2A) NMDA receptor (NMDAR) subtypes have been differentially implicated in activity-dependent synaptic plasticity. However, little is known about the respective contributions made by these two subtypes to developmental plasticity, in part because studies of GluN2B KO [Grin2b(-/-) (2b(-/-))] mice are hampered by early neonatal mortality. We previously used in vitro slice cocultures of rodent cerebral cortex (Cx) and spinal cord (SpC) to show that corticospinal (CS) synapses, once present throughout the SpC, are eliminated from the ventral side during development in an NMDAR-dependent manner. To study subtype specificity of NMDAR in this developmental plasticity, we cocultured Cx and SpC slices derived from postnatal day 0 (P0) animals with different genotypes [2b(-/-), Grin2a(-/-) (2a(-/-)), or WT mice]. The distribution of CS synapses was studied electrophysiologically and with a voltage-sensitive dye. Synapse elimination on the ventral side was blocked in WT(Cx)-2b(-/-)(SpC) pairs but not in WT(Cx)-2a(-/-)(SpC) or 2b(-/-)(Cx)-WT(SpC) pairs. CS axonal regression was also observed through live imaging of CS axons labeled with enhanced yellow fluorescent protein (EYFP) through exo utero electroporation. These findings suggest that postsynaptic GluN2B is selectively involved in CS synapse elimination. In addition, the elimination was not blocked in 2a(-/-) SpC slices, where Ca(2+) entry through GluN2B-mediated CS synaptic currents was reduced to the same level as in 2b(-/-) slices, suggesting that the differential effect of GluN2B and GluN2A in CS synapse elimination might not be explained based solely on greater Ca(2+) entry through GluN2B-containing channels.
GluN2B（GluRepsilon2/NR2B）和 GluN2A（GluRepsilon1/NR2A）N-甲基-D-天冬氨酸受体（NMDAR）亚基在活性依赖型突触可塑性中被不同地牵连。然而，对于这两种亚型对发育可塑性的各自贡献，人们知之甚少，部分原因是 GluN2B KO [Grin2b(-/-)（2b(-/-)）] 小鼠的研究受到早期新生鼠死亡率的阻碍。我们之前使用体外鼠大脑皮层（Cx）和脊髓（SpC）切片共培养，显示在发育过程中，CS 突触一旦存在于整个 SpC 中，就会以 NMDAR 依赖性的方式从腹侧被消除。为了研究这种发育可塑性中 NMDAR 的亚型特异性，我们使用来自出生后第 0 天（P0）的动物的 Cx 和 SpC 切片进行共培养，这些动物具有不同的基因型 [2b(-/-)、Grin2a(-/-)（2a(-/-)）或 WT 小鼠]。通过电生理学和电压敏感染料研究 CS 突触的分布。CS 突触的消除在 WT(Cx)-2b(-/-)(SpC) 对中被阻断，但在 WT(Cx)-2a(-/-)(SpC) 或 2b(-/-)(Cx)-WT(SpC) 对中未被阻断。通过通过胚胎外电穿孔用增强型黄色荧光蛋白（EYFP）标记 CS 轴突进行活细胞成像，也观察到 CS 轴突的退化。这些发现表明，突触后 GluN2B 选择性参与 CS 突触的消除。此外，在 2a(-/-) SpC 切片中，CS 突触电流通过 GluN2B 介导的 Ca(2+) 内流减少到与 2b(-/-) 切片相同的水平，消除并未被阻断，这表明 CS 突触消除中 GluN2B 和 GluN2A 的差异效应可能不仅仅基于通过含有 GluN2B 的通道的更大 Ca(2+) 内流来解释。
