Mutations in synaptic NMDA receptors (NMDARs) are associated with epilepsy and neurodevelopmental disorders. The effects of several such mutations have been investigated in recombinantly-expressed NMDARs under conditions of steady-state activation. Such experiments provide only limited insight into how mutations affect NMDAR-mediated excitatory synaptic currents (EPSCs). The present study aimed to characterize the effects of the GluN2AN615K, GluN2BN615I and GluN2BV618G gain-of-function mutations on EPSCs mediated by diheteromeric GluN1/2A and GluN1/2B receptors and triheteromeric GluN1/2A/2B receptors, as these are the most abundant synaptic NMDARs in vivo. Subunit composition was controlled by studying 'artificial' synapses formed between cultured neurons (which provide presynaptic terminals) and HEK293 cells that express the NMDAR subunits of interest plus the synapse-promoting molecule, neuroligin-1B. When incorporated into diheteromeric receptors, all three mutations ablated voltage-dependent Mg2+ block of EPSCs, as previously shown. In addition, we were surprised to find that increasing external Mg2+ from 0 to 1 mM strongly enhanced the magnitude of EPSCs mediated by mutant diheteromers. In contrast, triheteromeric receptors exhibited normal voltage-dependent Mg2+ block. The GluN2AN615K mutation also slowed the decay of GluN1/2A/2B- but not GluN1/2A-mediated EPSCs. The GluN2BN615I mutation enhanced the magnitude of both GluN1/2B- and GluN1/2A/2B-mediated EPSCs. The GluN2BV618G mutation enhanced the magnitude of both GluN1/2B- and GluN1/2A/2B-mediated EPSCs, although these effects were partly compensated by a faster EPSC decay rate. The mutations also diminished the potency of the anti-epileptic pore-blocker, memantine, thus explaining the lack of memantine efficacy in patients with GluN2BN615I or GluN2BV618G mutations. Given these effects, the three mutations would be expected to enhance the cation influx rate and thereby contribute to epilepsy phenotypes.

中文翻译：

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GluN2A和GluN2B功能获得性癫痫突变对二异聚和三异聚NMDA受体介导的突触电流的影响。

突触NMDA受体（NMDARs）的突变与癫痫和神经发育障碍有关。已经在稳态激活条件下在重组表达的NMDAR中研究了几种此类突变的作用。此类实验仅对突变如何影响NMDAR介导的兴奋性突触电流（EPSC）提供了有限的见识。本研究旨在表征GluN2AN615K，GluN2BN615I和GluN2BV618G功能获得性突变对由二聚体GluN1 / 2A和GluN1 / 2B受体以及三聚体GluN1 / 2A / 2B受体介导的EPSC的影响。体内NMDAR。通过研究“人工”来控制亚基组成 在培养的神经元（提供突触前末端）与表达感兴趣的NMDAR亚基以及促进突触的分子Neuroligin-1B的HEK293细胞之间形成突触。如前所述，当掺入二聚体受体中时，所有三个突变均会消除EPSC的电压依赖性Mg2 +阻滞。此外，我们惊讶地发现，将外部Mg2 +从0增加到1 mM可以大大增强由突变二异聚体介导的EPSC的强度。相反，三异聚体受体表现出正常的电压依赖性Mg2 +阻滞。GluN2AN615K突变也减慢了GluN1 / 2A / 2B-的衰减，但没有减缓GluN1 / 2A介导的EPSC的衰减。GluN2BN615I突变增强了GluN1 / 2B和GluN1 / 2A / 2B介导的EPSC的大小。GluN2BV618G突变增强了GluN1 / 2B和GluN1 / 2A / 2B介导的EPSC的强度，尽管这些影响被更快的EPSC衰减速率所补偿。突变还降低了抗癫痫的毛孔阻断剂美金刚的效力，从而解释了在具有GluN2BN615I或GluN2BV618G突变的患者中美金刚缺乏疗效。考虑到这些影响，可以预期这三个突变会增加阳离子流入速度，从而有助于癫痫的表型。