Glutathione metabolism in primary astrocyte cultures: flow cytometric evidence of heterogeneous distribution of GSH content.
The time-course of intracellular glutathione (GSH) values after incubation with L-buthionine-(S,R)-sulfoximine (BSO), a selective inhibitor of gamma-glutamylcysteine synthetase, showed that glutathione turns over with a half-life of 5 h. Intracellular GSH was assayed by flow cytometry using three different methods. Astrocytes showed a narrow range of cellular size but a wide range of intracellular GSH. This heterogeneity was resolved into three distinct subpopulations which represent 20%, 35% and 45% of the total astrocyte number. The less abundant subpopulation had the lower GSH content, while the most abundant was the subpopulation with the higher content. Over 95% of astrocytes were in the G0/G1 phase of the cell cycle, the distribution of cytosolic pH was homogeneous and the number of viable cells at the time of the assay was 90%. These results show that several pools exist when astrocyte GSH is considered and these findings may be relevant to the understanding of brain GSH metabolism.
Departamento de Bioquímica y Biología Molecular, Universidad de Valencia, Spain.
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A
Devesa
J E
JE
O'Connor
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C
Garciá
I R
IR
Puertes
J R
JR
Viña
D000818:Animals; D000831:Animals, Newborn; D000963:Antimetabolites; D001253:Astrocytes / Q000378:metabolism* / Q000648:ultrastructure; D001923:Brain Chemistry; D019328:Buthionine Sulfoximine; D002453:Cell Cycle; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D005434:Flow Cytometry; D005978:Glutathione / Q000378:metabolism*; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D008717:Methionine Sulfoximine / Q000031:analogs & derivatives; D009419:Nerve Tissue Proteins / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D000963:Antimetabolites; D009419:Nerve Tissue Proteins; D008717:Methionine Sulfoximine; D019328:Buthionine Sulfoximine; D004247:DNA; D005978:Glutathione