Production of reactive oxygen by mitochondria from normoxic and hypoxic rat heart tissue.
Reactive oxygen species (ROS), which may be involved in ischemic or reperfusion heart injury, can be produced by mitochondria. Previous work indicated that coupled mitochondria from ischemic heart tissue incubated in calcium-free medium produced less ROS than normal. The effects of calcium, which may be elevated in hypoxic or ischemic tissue, were not examined. The relative production of ROS by mitochondria from normoxic or hypoxic rat heart tissue was estimated by measuring the oxidation of dichlorofluorescin to the fluorescent compound, dichlorofluorescein. ROS were detectable during succinate-stimulated State 4 respiration. In the absence of calcium, mitochondria from hypoxic (60 min) heart tissue produced less ROS than mitochondria from normoxic heart tissue. In the presence of 0.1, 1 or 10 microM calcium, ROS produced by hypoxic mitochondria were increased to normoxic levels. While function was depressed in mitochondria from hypoxic tissue, the presence of 0.1 and 1 microM calcium had no further effect. Respiration was uncoupled in the presence of 10 microM calcium in mitochondria from both normoxic and hypoxic heart tissue. ROS production was increased in mitochondria from hypoxic tissue with both increasing concentrations of calcium and increasing duration of exposure. ROS production in mitochondria from normoxic heart tissue was only stimulated after 200 or more seconds of exposure to 1 or 10 microM calcium. Production of ROS in mitochondria from hypoxic tissue in the presence of 1 microM calcium was inhibited by rotenone (80%), ruthenium red (69%), and a combination of these agents (96%). In contrast, ruthenium red had no effect on ROS production by mitochondria from normoxic heart tissue.(ABSTRACT TRUNCATED AT 250 WORDS)
Paraidathathu
T
T
Division of Pharmacology and Toxicology, College of Pharmacy, University of Texas, Austin 78712-1074.
de Groot
H
H
Kehrer
J P
JP
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D008297:Male; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012402:Rotenone / Q000494:pharmacology; D012430:Ruthenium Red / Q000494:pharmacology
D005452:Fluoresceins; D017382:Reactive Oxygen Species; D012402:Rotenone; D012430:Ruthenium Red; C037631:2',7'-dichlorofluorescein; D002118:Calcium