Müller cell protection of rat retinal ganglion cells from glutamate and nitric oxide neurotoxicity.
Müller cell protection of rat retinal ganglion cells from glutamate and nitric oxide neurotoxicity.
复制标题
DOI：
--
复制
发表时间：
2000-10
影响因子：
4.4
通讯作者：
A. Kawasaki;Y. Otori;C. Barnstable
中科院分区：
文献类型：
--
作者：
A. Kawasaki;Y. Otori;C. Barnstable
目的低浓度的谷氨酸和一氧化氮等兴奋性毒性物质会降低纯化的视网膜神经节细胞(RGC)的存活率。在视网膜中，视网膜节细胞被视网膜Müller胶质细胞突起包裹。本研究的目的是确定RMG细胞是否能保护RGC免受这些兴奋性毒性损伤。方法从7日龄或8日龄Long Evans大鼠分离纯化视网膜神经节细胞，在无血清条件下接种于包被多聚赖氨酸/层粘连蛋白的玻片上培养2d。然后将盖片移到含有融合的RMG单层的培养皿中，或在无谷氨酸的培养液中不含胶质细胞。一些融合的RMG细胞培养皿中加入谷氨酸摄取阻断剂D，L-苏氨酸-β-羟基天冬氨酸(THA)。加入不同浓度的谷氨酸或NO供体2，2‘-(羟基亚硝基-肼)双乙胺3d后，用钙黄绿素-乙氧基甲酯染色检测RGCs的存活率。用氨基酸分析法测定培养液中的谷氨酸浓度。结果在无RMG细胞的情况下，增加浓度(5~500微米)的谷氨酸3d后可引起RGC死亡的剂量依赖性增加。谷氨酸的神经毒性作用在RMG细胞共培养中被阻断，即使在细胞类型之间没有直接接触的情况下也是如此。THA对RMG细胞的保护作用减弱。NO对RGC也有毒性作用。RMG细胞只有在与RGC直接接触时才能阻止这种毒性。结论RMG细胞对谷氨酸对RGCs有保护作用，无神经毒性。我们认为RMG的谷氨酸摄取功能紊乱可能是青光眼的病因之一。
PURPOSE Low concentrations of excitotoxic agents such as glutamate and nitric oxide decrease survival rates of purified retinal ganglion cells (RGCs). In the retina, RGCs are ensheathed by retinal Müller glial (RMG) cell processes. The purpose of this study was to determine whether RMG cells could protect RGCs from these excitotoxic injuries. METHODS RGCs were purified from 7- or 8-day-old Long Evans rats and cultured on polylysine/laminin-coated coverslips in serum-free medium for 2 days. The coverslips were then moved to dishes containing either confluent RMG monolayers or no glial cells in glutamate-free medium. Some dishes with confluent RMG cells were exposed to D,L-threo-beta-hydroxyaspartate (THA), a blocker of glutamate uptake. Three days after exposure to various concentrations of glutamate or the NO donor, 2, 2'-(hydroxynitroso-hydrazino)bisethanamine, survival rates of RGCs were measured by calcein-acetoxymethyl ester staining. Glutamate concentrations in the medium were measured using amino acid analysis. RESULTS Without RMG cells, the application of increasing concentrations (5-500 microM) of glutamate caused a dose-dependent increase in RGC death after 3 days. The neurotoxic effects of glutamate were blocked in the RMG cell cocultures, even when there was no direct contact between the cell types. The protective effect of RMG cells was weakened by THA treatment. NO also had toxic effects on RGC. RMG cells prevented this toxicity but only when in direct contact with the RGCs. CONCLUSIONS RMG cells can protect RGCs from glutamate and NO neurotoxicity. We suggest that functional disorders of glutamate uptake in RMGs might be one of the etiologies of glaucoma.