Glutamate uptake inhibition by oxygen free radicals in rat cortical astrocytes.
Formation of reactive oxygen species and disfunction of the excitatory amino acid (EAA) system are thought to be key events in the development of neuronal injury in several acute and long-term neurodegenerative diseases. Recent evidence suggests that the two phenomena may be interdependent. The present study is aimed at exploring possible molecular mechanisms underlying oxygen radical-EAA interaction. Exposure of cortical astrocytic cultures to either xanthine + xanthine oxidase (X/XO), a free radical-generating system, or hydrogen peroxide (H2O2) results in a marked decrease of high-affinity glutamate transport. Within 10 min of X/XO application, uptake falls to approximately 60% of its control value. In parallel no detectable release of lactate dehydrogenase occurs. X/XO effect is abolished in the presence of a mixture of scavenger enzymes (superoxide dismutase+catalase) or by the disulfide-reducing agents glutathione and dithiothreitol (DTT), but not by lipophilic antioxidants or ascorbate. The time course of inhibition shows an almost linear decline of glutamate transport during cell exposure to free radicals, while upon their inactivation the decline stops but established inhibition persists for at least 1 hr. In this situation, application of DTT significantly restores transport function. These data suggest that free radicals inhibit glutamate uptake primarily by long-lasting oxidation of protein sulfhydryl (SH) groups. Chemical modifiers of free SH groups, such as p-chloromercuribenzoate and N-ethylmaleimide, also induce uptake inhibition. Na+/K+ ATPase is a known target of oxygen radicals and may be involved in glutamate uptake inhibition. Indeed, ouabain, a blocker of the pump, reduces uptake in astrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
Volterra
A
A
Center of Neuropharmacology, Institute of Pharmacological Sciences, University of Milan, Italy.
Trotti
D
D
Tromba
C
C
Floridi
S
S
Racagni
G
G
D015763:2-Amino-5-phosphonovalerate / Q000494:pharmacology; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000494:pharmacology*; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002374:Catalase / Q000494:pharmacology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000378:metabolism*; D002729:Chloromercuribenzoates / Q000494:pharmacology; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D016166:Free Radical Scavengers; D005609:Free Radicals / Q000494:pharmacology; D005971:Glutamates / Q000378:metabolism*; D018698:Glutamic Acid; D005978:Glutathione / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D013482:Superoxide Dismutase / Q000494:pharmacology; D013997:Time Factors; D019820:Xanthine; D014969:Xanthine Oxidase / Q000494:pharmacology; D014970:Xanthines / Q000494:pharmacology; D020245:p-Chloromercuribenzoic Acid
D000975:Antioxidants; D002729:Chloromercuribenzoates; D016166:Free Radical Scavengers; D005609:Free Radicals; D005971:Glutamates; D011810:Quinoxalines; D014970:Xanthines; D019820:Xanthine; D018698:Glutamic Acid; D020245:p-Chloromercuribenzoic Acid; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D015763:2-Amino-5-phosphonovalerate; D004228:Dithionitrobenzoic Acid; D006861:Hydrogen Peroxide; D007770:L-Lactate Dehydrogenase; D002374:Catalase; D013482:Superoxide Dismutase; D014969:Xanthine Oxidase; D005978:Glutathione; D005033:Ethylmaleimide; D004229:Dithiothreitol