Probing the putative α7 nAChR/NMDAR complex in human and murine cortex and hippocampus: Different degrees of complex formation in healthy and Alzheimer brain tissue

α7 nicotinic acetylcholine receptors (nAChRs) and N-methyl-D-aspartate receptors (NMDARs) are key mediators of central cholinergic and glutamatergic neurotransmission, respectively. In addition to numerous well-established functional interactions between α7 nAChRs and NMDARs, the two receptors have been proposed to form a multimeric complex, and in the present study we have investigated this putative α7 nAChR/NMDAR assembly in human and murine brain tissues. By α-bungarotoxin (BGT) affinity purification, α7 and NMDAR subunits were co-purified from human and murine cortical and hippocampal homogenates, substantiating the notion that the receptors are parts of a multimeric complex in the human and rodent brain. Interestingly, the ratios between GluN1 and α7 levels in BGT pull-downs from cortical homogenates from Alzheimer’s disease (AD) brains were significantly lower than those in pull-downs from non-AD controls, indicating a reduced degree of α7 nAChR/NMDAR complex formation in the diseased tissue. A similar difference in GluN1/α7 ratios was observed between pull-downs from cortical homogenates from adult 3xTg-AD and age-matched wild type (WT) mice, whereas the GluN1/α7 ratios determined in pull-downs from young 3xTg-AD and age-matched WT mice did not differ significantly. The observation that pretreatment with oligomeric amyloid-β1–42 reduced GluN1/α7 ratios in BGT pull-downs from human cortical homogenate in a concentration-dependent manner provided a plausible molecular mechanism for this observed reduction. In conclusion, while it will be important to further challenge the existence of the putative α7 nAChR/NMDAR complex in future studies applying other methodologies than biochemical assays and to investigate the functional implications of this complex for cholinergic and glutamatergic neurotransmission, this work supports the formation of the complex and presents new insights into its regulation in healthy and diseased brain tissue.α7烟碱型乙酰胆碱受体（α7 nicotinic acetylcholine receptors, α7 nAChRs）与N-甲基-D-天冬氨酸受体（N-methyl-D-aspartate receptors, NMDARs）分别是中枢胆碱能与谷氨酸能神经传递的关键介导因子。除二者间已被大量研究证实的诸多功能性相互作用外，已有研究提出这两类受体可形成多聚体复合物；本研究针对人体及鼠类脑组织中的这一推定α7 nAChR/NMDAR组装体展开了探究。通过α-银环蛇毒素（α-bungarotoxin, BGT）亲和纯化实验，我们从人体及鼠类皮层与海马体匀浆中共同纯化得到了α7亚基与NMDAR亚基，证实了两类受体在人脑及啮齿动物脑中均属于同一多聚体复合物的组成部分这一观点。值得注意的是，阿尔茨海默病（Alzheimer’s disease, AD）患者皮层匀浆经α-银环蛇毒素亲和下拉所得样本中，GluN1与α7的水平比值显著低于非AD对照样本的对应比值，提示患病脑组织中α7 nAChR/NMDAR复合物的形成程度有所降低。成年3xTg-AD小鼠与同龄野生型（wild type, WT）小鼠皮层匀浆的亲和下拉实验中，也观察到了类似的GluN1/α7比值差异；而幼年3xTg-AD小鼠与同龄野生型小鼠的亲和下拉样本中，该比值并无显著差异。用寡聚体β淀粉样蛋白1–42预处理可呈浓度依赖性地降低人体皮层匀浆经α-银环蛇毒素亲和下拉所得样本的GluN1/α7比值，这一发现为上述比值降低现象提供了合理的分子机制解释。综上，尽管未来研究需采用生化实验以外的其他方法进一步验证推定的α7 nAChR/NMDAR复合物的存在，并探究该复合物对胆碱能与谷氨酸能神经传递的功能性影响，但本研究支持该复合物的形成，并为其在健康与患病脑组织中的调控机制提供了新的见解。