Synaptic Structure Laboratory, Instituto de Investigación en Discapacidades Neurológicas (IDINE), Departamento de Ciencias Médicas, Facultad de Medicina, Universidad Castilla-La Mancha, Campus Biosanitario, Albacete, Spain.
Centro de Biología Molecular Severo Ochoa, CSIC-UAM, Madrid, Spain.
Brain Pathol. 2023 Jan;33(1):e13115. doi: 10.1111/bpa.13115. Epub 2022 Sep 4.
N-methyl-d-aspartate receptors (NMDARs) are pivotal players in the synaptic transmission and synaptic plasticity underlying learning and memory. Accordingly, dysfunction of NMDARs has been implicated in the pathophysiology of Alzheimer disease (AD). Here, we used histoblot and sodium dodecylsulphate-digested freeze-fracture replica labelling (SDS-FRL) techniques to investigate the expression and subcellular localisation of GluN1, the obligatory subunit of NMDARs, in the hippocampus of P301S mice. Histoblots showed that GluN1 expression was significantly reduced in the hippocampus of P301S mice in a laminar-specific manner at 10 months of age but was unaltered at 3 months. Using the SDS-FRL technique, excitatory synapses and extrasynaptic sites on spines of pyramidal cells and interneuron dendrites were analysed throughout all dendritic layers in the CA1 field. Our ultrastructural approach revealed a high density of GluN1 in synaptic sites and a substantially lower density at extrasynaptic sites. Labelling density for GluN1 in excitatory synapses established on spines was significantly reduced in P301S mice, compared with age-matched wild-type mice, in the stratum oriens (so), stratum radiatum (sr) and stratum lacunosum-moleculare (slm). Density for synaptic GluN1 on interneuron dendrites was significantly reduced in P301S mice in the so and sr but unaltered in the slm. Labelling density for GluN1 at extrasynaptic sites showed no significant differences in pyramidal cells, and only increased density in the interneuron dendrites of the sr. This differential alteration of synaptic versus extrasynaptic NMDARs supports the notion that the progressive accumulation of phospho-tau is associated with changes in NMDARs, in the absence of amyloid-β pathology, and may be involved in the mechanisms causing abnormal network activity of the hippocampal circuit.
N-甲基-D-天冬氨酸受体（NMDARs）是突触传递和学习记忆所必需的突触可塑性的关键参与者。因此，NMDAR 的功能障碍与阿尔茨海默病（AD）的病理生理学有关。在这里，我们使用组织印迹和十二烷基硫酸钠消化冷冻断裂复制标记（SDS-FRL）技术研究了 P301S 小鼠海马中 NMDAR 必需亚基 GluN1 的表达和亚细胞定位。组织印迹显示，在 10 个月大时，P301S 小鼠的海马中 GluN1 的表达呈层特异性显著降低，但在 3 个月时未改变。使用 SDS-FRL 技术，分析了 CA1 场中所有树突层中锥体和中间神经元树突上的兴奋性突触和突触外部位的棘突上的 GluN1 表达。我们的超微结构方法显示，突触部位的 GluN1 标记密度很高，而突触外部位的标记密度则明显较低。与年龄匹配的野生型小鼠相比，P301S 小鼠在棘突上建立的兴奋性突触的 GluN1 标记密度显著降低，在 so、sr 和 slm 层中。在 so 和 sr 层中，P301S 小鼠中兴奋性突触上的 GluN1 标记密度在中间神经元树突上显著降低，但在 slm 层中没有改变。在锥体细胞中，突触外部位的 GluN1 标记密度没有显著差异，而在 sr 层中仅增加了中间神经元树突上的 GluN1 标记密度。这种突触与突触外 NMDAR 之间的差异改变支持这样的观点，即磷酸化 tau 的逐渐积累与 NMDAR 的变化有关，而与淀粉样蛋白-β 病理无关，并且可能与海马回路异常网络活动的机制有关。
