title: Effect of glutamate on lysosomal membrane permeabilization in primary cultured cortical neurons.
abstract: Glutamate is the principal neurotransmitter in the central nervous system. Glutamate-mediated excitotoxicity is the predominant cause of cerebral damage. Recent studies have shown that lysosomal membrane permeabilization (LMP) is involved in ischemia‑associated neuronal death in non‑human primates. This study was designed to investigate the effect of glutamate on lysosomal stability in primary cultured cortical neurons. Glutamate treatment for 30 min induced the permeabilization of lysosomal membranes as assessed by acridine orange redistribution and immunofluorescence of cathepsin B in the cytoplasm. Inhibition of glutamate excitotoxicity by the NMDA receptor antagonist MK‑801 and the calcium chelator ethylene glycol‑bis (2‑aminoethylether)‑N, N, N', N'‑tetraacetic acid, rescued lysosomes from permeabilization. The role of calpain and reactive oxygen species (ROS) in inducing LMP was also investigated. Ca2+ overload following glutamate treatment induced the activation of calpain and the production of ROS, which are two major contributors to neuronal death. It has been reported that lysosomal‑associated membrane protein 2 (LAMP2) and heat shock protein (HSP)70 are two calpain substrates that promote LMP in cancer cells; however, it was found that calpains were activated by glutamate, but only LAMP2 was subsequently degraded. Furthermore, LMP was not alleviated by treatment with the calpain inhibitors calpeptin and SJA6017, which blocked the cleavage of the calpain substrate α‑fodrin. It was demonstrated that LMP was significantly alleviated by treatment with the antioxidant N‑Acetyl‑L‑cysteine, indicating that LMP involvement in early glutamate excitotoxicity may be mediated partly by ROS rather than calpain activation. Overall, these data shed light on the role of ROS-mediated LMP in early glutamate excitotoxicity.
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Min
authors.initials: M
authors.lastname: Yan
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Wenbo
authors.initials: W
authors.lastname: Zhu
authors.affiliation: Department of Pathology, The First Affiliated Hospital, Sun Yat‑sen University, Guangzhou, Guangdong 510089, P.R. China.
authors.forename: Xiaoke
authors.initials: X
authors.lastname: Zheng
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Lipeng
authors.initials: L
authors.lastname: Tang
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Bingzheng
authors.initials: B
authors.lastname: Lu
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Wenli
authors.initials: W
authors.lastname: Chen
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Pengxin
authors.initials: P
authors.lastname: Qiu
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Tiandong
authors.initials: T
authors.lastname: Leng
authors.affiliation: Guangzhou Cellprotek Pharmaceutical Co., Ltd., Guangzhou, Guangdong 510663, P.R. China.
authors.forename: Suizhen
authors.initials: S
authors.lastname: Lin
authors.affiliation: Department of Pharmacology, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Guangmei
authors.initials: G
authors.lastname: Yan
authors.affiliation: Department of Biochemistry, Zhongshan School of Medicine, Sun Yat‑sen University, Guangzhou, Guangdong 510080, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Yin
mesh_terms: D000818:Animals; D002154:Calpain / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D018698:Glutamic Acid / Q000494:pharmacology*; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D061251:Primary Cell Culture; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D017382:Reactive Oxygen Species; D018698:Glutamic Acid; D002154:Calpain
references.citation: J Exp Biol. 2004 Aug;207(Pt 18):3221-31
references.pmid: 15299043
references.citation: Oncogene. 2004 Apr 12;23(16):2881-90
references.pmid: 15077151
references.citation: Photochem Photobiol. 1990 Jan;51(1):67-76
references.pmid: 2304980
references.citation: Neuroreport. 1998 Aug 3;9(11):2437-42
references.pmid: 9721910
references.citation: Eur J Neurosci. 1998 May;10(5):1723-33
references.pmid: 9751144
references.citation: Curr Biol. 2005 Jul 12;15(13):1249-54
references.pmid: 16005300
references.citation: Pediatr Res. 2006 May;59(5):684-9
references.pmid: 16627882
references.citation: Biochim Biophys Acta. 2006 Apr;1765(2):101-25
references.pmid: 16412578
references.citation: J Neurochem. 2006 Sep;98(5):1597-609
references.pmid: 16923169
references.citation: J Neurochem. 2008 Jul;106(1):56-69
references.pmid: 18363826
references.citation: Ann N Y Acad Sci. 2008 Nov;1144:97-112
references.pmid: 19076369
references.citation: J Biol Chem. 2010 Jan 1;285(1):667-74
references.pmid: 19850931
references.citation: Biochem Biophys Res Commun. 2010 Mar 19;393(4):806-11
references.pmid: 20171158
references.citation: Apoptosis. 2010 May;15(5):527-40
references.pmid: 20077016
references.citation: Neurobiol Dis. 2010 Oct;40(1):293-302
references.pmid: 20600913
references.citation: Prog Neurobiol. 2010 Oct;92(2):184-211
references.pmid: 20685377
references.citation: Mech Ageing Dev. 2011 Aug;132(8-9):405-11
references.pmid: 21729715
references.citation: Eur J Pharmacol. 2013 Jan 5;698(1-3):6-18
references.pmid: 23123057
references.citation: Biochim Biophys Acta. 2013 Oct;1833(10):2244-53
references.pmid: 23747342
references.citation: Annu Rev Neurosci. 1990;13:171-82
references.pmid: 1970230
references.citation: FASEB J. 1992 Dec;6(15):3338-44
references.pmid: 1464368
references.citation: J Biol Chem. 1995 Mar 24;270(12):6425-8
references.pmid: 7534762
references.citation: Eur J Neurosci. 1996 Sep;8(9):1932-44
references.pmid: 8921284
references.citation: Proc Natl Acad Sci U S A. 1998 Jun 23;95(13):7748-53
references.pmid: 9636222
references.citation: Nat Rev Mol Cell Biol. 2002 Feb;3(2):94-103
references.pmid: 11836511
references.citation: Biol Chem. 2002 May;383(5):785-91
references.pmid: 12108543
references.citation: Brain Res. 2003 May 2;971(1):1-8
references.pmid: 12691831
references.citation: Hippocampus. 2003;13(7):791-800
references.pmid: 14620874
references.citation: J Neurosci. 2004 Jan 14;24(2):565-75
references.pmid: 14724257
references.citation: J Exp Med. 2004 Aug 16;200(4):425-35
references.pmid: 15314073