Document Detail

Glutamate-induced cytotoxicity in PC12 pheochromocytoma cells: role of oxidation of phospholipids, glutathione and protein sulfhydryls revealed by bcl-2 transfection.
MedLine Citation:
PMID:  9757062     Owner:  NLM     Status:  MEDLINE    
Incubation of mock-transfected PC12 rat pheochromocytoma cells (PC12) for 2 h with increasing concentrations of glutamate caused progressive loss of viability (e.g., 67% with 15 mM glutamate). In contrast, the viability of bcl-2-transfected cells (PC12/bcl-2) was unaffected by glutamate. Neither PC12 nor PC12/bcl-2 cells showed a significant incidence of apoptosis in response to glutamate. Conventional phospholipid analysis by high-performance TLC and phosphorous determination showed no significant changes in the phospholipid composition of either cell line incubated with </=15 mM glutamate. Phospholipid peroxidation was quantified in the cells using our newly developed method based on fluorescence-HPLC analysis of metabolically incorporated oxidation-sensitive and fluorescent fatty acid, cis-parinaric acid. Unlike previous studies that measured total phospholipid oxidation, this novel technology permitted quantitation of oxidative stress in different classes of labeled phospholipids (the amount of labeled phospholipids in the cells did not exceed 1% of total phospholipids). Significant peroxidation of phosphatidylcholine and phosphatidylethanolamine occurred in PC12 cells treated with >5 mM glutamate. The peroxyl radical initiator 2,2'-azobis(2,4-dimethylvaleronitrile) caused a pronounced loss of all major phospholipid classes in PC12 cells, but no loss of cell viability. No phospholipid peroxidation was detected in PC12/bcl-2 cells incubated with </=15 mM glutamate or with 2, 2'-azobis(2,4-dimethylvaleronitrile). These results directly demonstrate that peroxidation of membrane phospholipids is not responsible for the cytotoxicity of glutamate in PC12 cells. Total cellular thiol, protein thiol and GSH reserves were quantified by a previously described electron paramagnetic resonance spectrometric method. Total thiols were ca. 1.5-fold greater in PC12/bcl-2 than in PC12 cells. Glutamate (</=5 mM) caused a progressive and equally significant decrease in total thiols and GSH in both PC12 and PC12/bcl-2 cells. High glutamate concentrations caused oxidation of protein sulfhydryls in PC12 cells, but not in PC12/bcl-2 cells. The results suggest that the changes in cellular milieu caused by bcl-2 gene transfection protect PC12 cells from the toxic effects of glutamate in a manner consistent with prevention of protein sulfhydryl oxidation.
V A Tyurin; Y Y Tyurina; P J Quinn; N F Schor; R Balachandran; B W Day; V E Kagan
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Brain research. Molecular brain research     Volume:  60     ISSN:  0169-328X     ISO Abbreviation:  Brain Res. Mol. Brain Res.     Publication Date:  1998 Oct 
Date Detail:
Created Date:  1998-12-07     Completed Date:  1998-12-07     Revised Date:  2009-09-29    
Medline Journal Info:
Nlm Unique ID:  8908640     Medline TA:  Brain Res Mol Brain Res     Country:  NETHERLANDS    
Other Details:
Languages:  eng     Pagination:  270-81     Citation Subset:  IM    
Copyright Information:
Copyright 1998 Elsevier Science B.V.
Department of Environmental and Occupational Health, University of Pittsburgh, Pittsburgh, PA 15238, USA.
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MeSH Terms
Adrenal Gland Neoplasms
Azo Compounds / pharmacology
Cell Survival / drug effects,  physiology*
Genes, bcl-2*
Glutamic Acid / toxicity*
Glutathione / metabolism*
Neoplasm Proteins / metabolism
Nitriles / pharmacology
PC12 Cells
Phospholipids / metabolism*
Proto-Oncogene Proteins c-bcl-2 / genetics,  metabolism*
Recombinant Proteins / biosynthesis,  metabolism
Sulfhydryl Compounds / metabolism
Grant Support
F05 NS 10669/NS/NINDS NIH HHS; R01 CA 57288/CA/NCI NIH HHS; //Wellcome Trust
Reg. No./Substance:
0/Azo Compounds; 0/Neoplasm Proteins; 0/Nitriles; 0/Phospholipids; 0/Proto-Oncogene Proteins c-bcl-2; 0/Recombinant Proteins; 0/Sulfhydryl Compounds; 4419-11-8/2,2'-azobis(2,4-dimethylvaleronitrile); 56-86-0/Glutamic Acid; 70-18-8/Glutathione

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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