Laboratorio de Neurobiología Celular, Departamento de Neurociencias, Centro de Investigacion en Medicina Aplicada, 31008 Pamplona, Spain, and Département des Neurosciences Fondamentales, Université de Genève, Faculté de Médecine, Centre Médical Universitaire, 1211 Genève 4, Switzerland.
Département des Neurosciences Fondamentales, Université de Genève, Faculté de Médecine, Centre Médical Universitaire, 1211 Genève 4, Switzerland.
J Neurosci. 2014 Jul 9;34(28):9213-21. doi: 10.1523/JNEUROSCI.5183-13.2014.
Synaptic rearrangements during critical periods of postnatal brain development rely on the correct formation, strengthening, and elimination of synapses and associated dendritic spines to form functional networks. The correct balance of these processes is thought to be regulated by synapse-specific changes in the subunit composition of NMDA-type glutamate receptors (NMDARs). Among these, the nonconventional NMDAR subunit GluN3A has been suggested to play a role as a molecular brake in synaptic maturation. We tested here this hypothesis using confocal time-lapse imaging in rat hippocampal organotypic slices and assessed the role of GluN3A-containing NMDARs on spine dynamics. We found that overexpressing GluN3A reduced spine density over time, increased spine elimination, and decreased spine stability. The effect of GluN3A overexpression could be further enhanced by using an endocytosis-deficient GluN3A mutant and reproduced by silencing the adaptor protein PACSIN1, which prevents the endocytosis of endogenous GluN3A. Conversely, silencing of GluN3A reduced spine elimination and favored spine stability. Moreover, reexpression of GluN3A in more mature tissue reinstated an increased spine pruning and a low spine stability. Mechanistically, the decreased stability in GluN3A overexpressing neurons could be linked to a failure of plasticity-inducing protocols to selectively stabilize spines and was dependent on the ability of GluN3A to bind the postsynaptic scaffold GIT1. Together, these data provide strong evidence that GluN3A prevents the activity-dependent stabilization of synapses thereby promoting spine pruning, and suggest that GluN3A expression operates as a molecular signal for controlling the extent and timing of synapse maturation.
在出生后大脑发育的关键期，突触的重新排列依赖于突触和相关树突棘的正确形成、增强和消除，以形成功能性网络。这些过程的正确平衡被认为是受 NMDA 型谷氨酸受体 (NMDAR) 亚基组成的突触特异性变化调节的。在这些亚基中，非典型的 NMDAR 亚基 GluN3A 被认为在突触成熟中起分子制动的作用。我们在这里使用大鼠海马器官型切片的共聚焦延时成像来检验这一假说，并评估含有 GluN3A 的 NMDAR 在棘突动力学中的作用。我们发现，过表达 GluN3A 会随着时间的推移降低棘突密度，增加棘突消除，并降低棘突稳定性。使用内吞作用缺陷的 GluN3A 突变体进一步增强 GluN3A 的过表达作用，并通过沉默衔接蛋白 PACSIN1 来复制该作用，PACSIN1 可阻止内源性 GluN3A 的内吞作用。相反，沉默 GluN3A 可减少棘突消除并有利于棘突稳定性。此外，在更成熟的组织中重新表达 GluN3A 会恢复增加的棘突修剪和低棘突稳定性。从机制上讲，过表达 GluN3A 的神经元稳定性降低可能与诱导可塑性的方案不能选择性地稳定棘突有关，并且依赖于 GluN3A 结合突触后支架 GIT1 的能力。总之，这些数据提供了强有力的证据表明，GluN3A 可防止突触的活动依赖性稳定，从而促进棘突修剪，并表明 GluN3A 表达作为控制突触成熟程度和时间的分子信号发挥作用。
