Potentiation of intracellular Ca2+ mobilization by hypoxia-induced NO generation in rat brain striatal slices and human astrocytoma U-373 MG cells and its involvement in tissue damage.
The relationship between nitric oxide (NO) and intracellular Ca2+ in hypoxic-ischemic brain damage is not known in detail. Here we used rat striatal slices perfused under low-oxygen and Ca2+-free conditions and cultured human astrocytoma cells incubated under similar conditions as models to study the dynamics of intracellular NO and Ca2+ in hypoxia-induced tissue damage. Exposure of rat striatal slices for 70 min to low oxygen tension elicited a delayed and sustained increase in the release of 45Ca2+. This was potentiated by the NO donors sodium nitroprusside (SNP) and spermine-NO and inhibited by N-omega-nitro-L-arginine methyl ester (L-NAME) or by the NO scavenger 2-phenyl-4,4,5,5 tetramethylimidazoline-1-oxyl-3-oxide (PTIO). A membrane-permeant form of heparin in combination with either ruthenium red (RR) or ryanodine (RY) also inhibited 45Ca2+ release. In human astrocytoma U-373 MG cells, hypoxia increased intracellular Ca2+ concentration ([Ca2+]i) by 67.2 +/- 13.1% compared to normoxic controls and this effect was inhibited by L-NAME, PTIO or heparin plus RR. In striatal tissue, hypoxia increased NO production and LDH release and both effects were antagonized by L-NAME. Although heparin plus RR or RY antagonized hypoxia-induced increase in LDH release they failed to counteract increased NO production. These data therefore indicate that NO contributes to hypoxic damage through increased intracellular Ca2+ mobilization from endoplasmic reticulum and suggest that the NO-Ca2+ signalling might be a potential therapeutic target in hypoxia-induced neuronal degeneration.
Meini
Antonella
A
Istituto di Scienze Farmacologiche, Università di Siena, via A Moro 2, 53100 Siena, Italy.
Benocci
Alberto
A
Frosini
Maria
M
Sgaragli
Gian Pietro
GP
Garcia
Julian Blanco
JB
Pessina
Gian Paolo
GP
Aldinucci
Carlo
C
Palmi
Mitri
M
D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology; D001254:Astrocytoma; D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D003342:Corpus Striatum / Q000187:drug effects / Q000378:metabolism*; D003497:Cyclic N-Oxides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004338:Drug Combinations; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D016257:Fura-2 / Q000031:analogs & derivatives* / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D006836:Hydro-Lyases / Q000378:metabolism; D000860:Hypoxia / Q000378:metabolism* / Q000503:physiopathology; D007093:Imidazoles / Q000494:pharmacology; D066298:In Vitro Techniques; D042541:Intracellular Space / Q000378:metabolism; D008297:Male; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009599:Nitroprusside / Q000494:pharmacology; D010477:Perfusion / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D012428:Ruthenium / Q000494:pharmacology; D012433:Ryanodine / Q000494:pharmacology
D000925:Anticoagulants; D003497:Cyclic N-Oxides; D004338:Drug Combinations; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D007093:Imidazoles; D020030:Nitric Oxide Donors; C049925:fura-2-am; D012433:Ryanodine; D009599:Nitroprusside; C079391:2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide; D009569:Nitric Oxide; D012428:Ruthenium; D006493:Heparin; D006836:Hydro-Lyases; C045581:lactate dehydratase; D002118:Calcium; D016257:Fura-2; D019331:NG-Nitroarginine Methyl Ester