Section of Human Anatomy and Histology, Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.
Section of Physiological Sciences, Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.
J Neuroendocrinol. 2020 Mar;32(3):e12837. doi: 10.1111/jne.12837. Epub 2020 Feb 19.
It has been well established, particularly in animal models, that oestrogens exert neuroprotective effects in brain areas linked to cognitive processes. A key protective role could reside in the capacity of oestrogen to modulate the inflammatory response. However, the direct neuroprotective actions of oestrogens on neurones are complex and remain to be fully clarified. In the present study, we took advantage of a previously characterised primary culture of human cholinergic neurones (hfNBM) from the foetal nucleus basalis of Meynert, which is known to regulate hippocampal and neocortical learning and memory circuits, aiming to investigate the direct effects of oestrogens under inflammatory conditions. Exposure of cells to tumour necrosis factor (TNF)α (10 ng mL ) determined the activation of an inflammatory response, as demonstrated by nuclear factor-kappa B p65 nuclear translocation and cyclooxygenase-2 mRNA expression. These effects were inhibited by treatment with either 17β-oestradiol (E ) (10 nmol L ) or G1 (100 nmol L ), the selective agonist of the G protein-coupled oestrogen receptor (GPER1). Interestingly, the GPER1 antagonist G15 abolished the effects of E in TNFα-treated cells, whereas the ERα/ERβ inhibitor tamoxifen did not. Electrophysiological measurements in hfNBMs revealed a depolarising effect caused by E that was specifically blocked by tamoxifen and not by G15. Conversely, G1 specifically hyperpolarised the cell membrane and also increased both inward and outward currents elicited by a depolarising stimulus, suggesting a modulatory action on hfNBM excitability by GPER1 activation. Interestingly, pretreating cells with TNFα completely blocked the effects of G1 on membrane properties and also significantly reduced GPER1 mRNA expression. In addition, we found a peculiar subcellular localisation of GPER1 to focal adhesion sites that implicates new possible mechanisms of action of GPER1 in the neuronal perception of mechanical stimuli. The results obtained in the present study indicate a modulatory functional role of GPER1 with respect to mediating the oestrogen neuroprotective effect against inflammation in brain cholinergic neurones and, accordingly, may help to identify protective strategies for preventing cognitive impairments.
已经有充分的证据表明，雌激素在与认知过程相关的大脑区域中发挥神经保护作用，这在动物模型中尤为明显。雌激素的一个关键保护作用可能在于其调节炎症反应的能力。然而，雌激素对神经元的直接神经保护作用是复杂的，仍有待充分阐明。在本研究中，我们利用了先前从梅内尔特基底核中分离的人胆碱能神经元（hfNBM）的特征性原代培养物，该培养物已知可调节海马体和新皮层的学习和记忆回路，旨在研究在炎症条件下雌激素的直接作用。细胞暴露于肿瘤坏死因子（TNF）α（10ngmL-1）会导致炎症反应的激活，这可通过核因子-kappa B p65 核易位和环氧化酶-2 mRNA 表达来证明。这些作用可通过用 17β-雌二醇（E2）（10nmolL-1）或 G1（100nmolL-1）处理来抑制，G1 是 G 蛋白偶联雌激素受体（GPER1）的选择性激动剂。有趣的是，GPER1 拮抗剂 G15 消除了 E2 在 TNFα 处理的细胞中的作用，而 ERα/ERβ 抑制剂他莫昔芬则没有。在 hfNBM 中的电生理测量显示 E2 引起的去极化作用，该作用被他莫昔芬特异性阻断，而不是被 G15 阻断。相反，G1 特异性地使细胞膜超极化，并增加由去极化刺激引起的内向和外向电流，这表明通过激活 GPER1 对 hfNBM 兴奋性产生调节作用。有趣的是，用 TNFα 预处理细胞可完全阻断 G1 对膜特性的影响，并显著降低 GPER1 mRNA 的表达。此外，我们发现 GPER1 定位于局灶性黏附位点的特殊亚细胞定位，这暗示了 GPER1 在神经元对机械刺激的感知中具有新的可能作用机制。本研究的结果表明，GPER1 具有调节功能作用，可介导雌激素对大脑胆碱能神经元炎症的神经保护作用，因此有助于确定预防认知障碍的保护策略。
