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


CONCLUSIONS
Glutamate exerts toxicity in human neuroblastoma SH-SY5Y cells possibly through oxidative damage, not through calcium homeostasis destruction mediated by NMDA receptors.
Sun
Zhong-Wei
ZW
Shanghai Institute of Brain Functional Genomics, East China Normal University, Shanghai 200062, China.
Zhang
Lan
L
Zhu
Shu-Jia
SJ
Chen
Wen-Chun
WC
Mei
Bing
B
D045784:Abietanes; D000596:Amino Acids / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D007211:Indoles; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008315:Malondialdehyde / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D010616:Phenanthrenes / Q000494:pharmacology; D013482:Superoxide Dismutase / Q000378:metabolism; D013997:Time Factors; D014966:Xanthenes / Q000494:pharmacology
D045784:Abietanes; D000596:Amino Acids; D000975:Antioxidants; D018691:Excitatory Amino Acid Antagonists; D007211:Indoles; C114624:LY 341495; D010616:Phenanthrenes; D014966:Xanthenes; C021751:tanshinone; D018698:Glutamic Acid; C007293:DAPI; D008315:Malondialdehyde; D016291:Dizocilpine Maleate; D007770:L-Lactate Dehydrogenase; D013482:Superoxide Dismutase; D005978:Glutathione; D002118:Calcium