Vollum Institute, Oregon Health & Science University, Portland, Oregon 97239, USA.
J Neurosci. 2013 May 22;33(21):9150-60. doi: 10.1523/JNEUROSCI.0829-13.2013.
NMDA receptors are composed of two GluN1 (N1) and two GluN2 (N2) subunits. Constituent N2 subunits control the pharmacological and kinetic characteristics of the receptor. NMDA receptors in hippocampal or cortical neurons are often thought of as diheteromeric, meaning that they contain only one type of N2 subunit. However, triheteromeric receptors with more than one type of N2 subunit also have been reported, and the relative contribution of diheteromeric and triheteromeric NMDA receptors at synapses has been difficult to assess. Because wild-type hippocampal principal neurons express N1, N2A, and N2B, we used cultured hippocampal principal neurons from N2A and N2B knock-out mice as templates for diheteromeric synaptic receptors. However, summation of N1/N2B and N1/N2A EPSCs could not account for the deactivation kinetics of wild-type EPSCs. To make a quantitative estimate of NMDA receptor subtypes at wild-type synapses, we used the deactivation kinetics and the effects of the competitive antagonist NVP-AAM077. Our results indicate that three types of NMDA receptors contribute to wild-type EPSCs, with at least two-thirds being triheteromeric receptors. Functional isolation of synaptic triheteromeric receptors revealed deactivation kinetics and pharmacology that were distinct from either diheteromeric receptor subtype. Because of differences in open probability, synaptic triheteromeric receptors outnumbered N1/N2A receptors by 5.8 to 1 and N1/N2B receptors by 3.2 to 1. Our results suggest that triheteromeric NMDA receptors must either be preferentially assembled or preferentially localized at synapses.
NMDA 受体由两个 GluN1（N1）和两个 GluN2（N2）亚基组成。组成 N2 亚基控制受体的药理学和动力学特性。海马体或皮质神经元中的 NMDA 受体通常被认为是二聚体，这意味着它们只包含一种类型的 N2 亚基。然而，也有报道称存在含有多种 N2 亚基的三聚体受体，而且突触中二聚体和三聚体 NMDA 受体的相对贡献一直难以评估。由于野生型海马体主神经元表达 N1、N2A 和 N2B，我们使用 N2A 和 N2B 敲除小鼠的培养海马体主神经元作为二聚体突触受体的模板。然而，N1/N2B 和 N1/N2A EPSC 的总和不能解释野生型 EPSC 的失活动力学。为了对野生型突触中 NMDA 受体亚型进行定量估计，我们使用了失活动力学和竞争性拮抗剂 NVP-AAM077 的作用。我们的结果表明，三种类型的 NMDA 受体都有助于产生野生型 EPSC，其中至少有三分之二是三聚体受体。突触三聚体受体的功能分离揭示了与任何二聚体受体亚型都不同的失活动力学和药理学。由于开放概率的差异，突触三聚体受体的数量比 N1/N2A 受体多 5.8 比 1，比 N1/N2B 受体多 3.2 比 1。我们的结果表明，三聚体 NMDA 受体必须优先组装或优先定位于突触。
