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. 2010 Feb 3;26(1):8–16. doi: 10.1007/s12264-010-0813-7

Excitotoxicity effects of glutamate on human neuroblastoma SH-SY5Y cells via oxidative damage

谷氨酸致人神经母细胞瘤细胞兴奋性毒损伤的机制

Zhong-Wei Sun 1, Lan Zhang 1, Shu-Jia Zhu 1, Wen-Chun Chen 1, Bing Mei 1,✉
PMCID: PMC5560379  PMID: 20101268

Abstract

Objective

To investigate the mechanisms of excitotoxic effects of glutamate on human neuroblastoma SH-SY5Y cells.

Methods

SH-SY5Y cell viability was measured by MTT assay. Other damaged profile was detected by lactate dehydrogenase (LDH) release and by 4′, 6-diamidino-2-phenylindole (DAPI) staining. The cytosolic calcium concentration was tested by calcium influx assay. The glutamate-induced oxidative stress was analyzed by cytosolic glutathione assay, superoxide dismutase (SOD) assay and extracellular malondialdehyde (MDA) assay.

Results

Glutamate treatment caused damage in SHSY5Y cells, including the decrease of cell viability, the increase of LDH release and the alterations of morphological structures. Furthermore, the concentration of cytoplasmic calcium in SH-SY5Y cells was not changed within 20 min following glutamate treatment, while cytosolic calcium concentration significantly increased within 24 h after glutamate treatment, which could not be inhibited by MK801, an antagonist of NMDA receptors, or by LY341495, an antagonist of metabotropic glutamate receptors. On the other hand, oxidative damage was observed in SH-SY5Y cells treated with glutamate, including decreases in glutathione content and SOD activity, and elevation of MDA level, all of which could be alleviated by an antioxidant Tanshinone IIA (Tan IIA, a major active ingredient from a Chinese plant Salvia Miltiorrhiza Bge).

Conclusion

Glutamate exerts toxicity in human neuroblastoma SH-SY5Y cells possibly through oxidative damage, not through calcium homeostasis destruction mediated by NMDA receptors.

Keywords: glutamate, excitotoxicity, cytosolic calcium, oxidative damage

Footnotes

These authors contributed equally to this work.

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