Department of Pharmacology, School of Medical Sciences, University of New South Wales, and Vincent's Centre for Applied Medical Research, St. Vincent's Hospital, Sydney, Australia.
PLoS One. 2010 Nov 30;5(11):e14123. doi: 10.1371/journal.pone.0014123.
Astrocytes have long been perceived only as structural and supporting cells within the central nervous system (CNS). However, the discovery that these glial cells may potentially express receptors capable of responding to endogenous neurotransmitters has resulted in the need to reassess astrocytic physiology. The aim of the current study was to characterise the expression of NMDA receptors (NMDARs) in primary human astrocytes, and investigate their response to physiological and excitotoxic concentrations of the known endogenous NMDAR agonists, glutamate and quinolinic acid (QUIN). Primary cultures of human astrocytes were used to examine expression of these receptors at the mRNA level using RT-PCR and qPCR, and at the protein level using immunocytochemistry. The functionality role of the receptors was assessed using intracellular calcium influx experiments and measuring extracellular lactate dehydrogenase (LDH) activity in primary cultures of human astrocytes treated with glutamate and QUIN. We found that all seven currently known NMDAR subunits (NR1, NR2A, NR2B, NR2C, NR2D, NR3A and NR3B) are expressed in astrocytes, but at different levels. Calcium influx studies revealed that both glutamate and QUIN could activate astrocytic NMDARs, which stimulates Ca2+ influx into the cell and can result in dysfunction and death of astrocytes. Our data also show that the NMDAR ion channel blockers, MK801, and memantine can attenuate glutamate and QUIN mediated cell excitotoxicity. This suggests that the mechanism of glutamate and QUIN gliotoxicity is at least partially mediated by excessive stimulation of NMDARs. The present study is the first to provide definitive evidence for the existence of functional NMDAR expression in human primary astrocytes. This discovery has significant implications for redefining the cellular interaction between glia and neurons in both physiological processes and pathological conditions.
星形胶质细胞长期以来仅被认为是中枢神经系统 (CNS) 中的结构和支持细胞。然而,发现这些神经胶质细胞可能潜在地表达能够响应内源性神经递质的受体,这导致需要重新评估星形胶质细胞的生理学。本研究的目的是表征原代人星形胶质细胞中 NMDA 受体 (NMDAR) 的表达,并研究它们对生理和兴奋性毒性浓度的已知内源性 NMDAR 激动剂谷氨酸和喹啉酸 (QUIN) 的反应。使用 RT-PCR 和 qPCR 在 mRNA 水平上,使用免疫细胞化学在蛋白质水平上检查这些受体在原代人星形胶质细胞中的表达。使用细胞内钙内流实验和测量谷氨酸和 QUIN 处理的原代人星形胶质细胞中外源性乳酸脱氢酶 (LDH) 活性来评估受体的功能作用。我们发现,所有七种目前已知的 NMDAR 亚基 (NR1、NR2A、NR2B、NR2C、NR2D、NR3A 和 NR3B) 在星形胶质细胞中表达,但水平不同。钙内流研究表明,谷氨酸和 QUIN 均可激活星形胶质细胞 NMDAR,这刺激 Ca2+ 流入细胞,并可能导致星形胶质细胞功能障碍和死亡。我们的数据还表明,NMDAR 离子通道阻滞剂 MK801 和美金刚胺可以减弱谷氨酸和 QUIN 介导的细胞兴奋性毒性。这表明谷氨酸和 QUIN 神经毒性的机制至少部分是通过过度刺激 NMDAR 介导的。本研究首次提供了明确的证据,证明原代人星形胶质细胞中存在功能性 NMDAR 表达。这一发现对于重新定义生理和病理条件下胶质细胞和神经元之间的细胞相互作用具有重要意义。