Effects of Bilirubin on Mitochondrial Reactions

Abstract

Abstract We have studied the effects of bilirubin on respiration, respiratory control, oxidative phosphorylation, and swelling of mitochondria from rat liver, bovine heart, and brain of chick, rabbit, and monkey. Low (l∼20 µm) concentrations of bilirubin increase and high concentrations (g50 µm) decrease the respiration of liver or heart mitochondria. This biphasic effect is not observed in brain mitochondria, in which respiration is always inhibited by bilirubin. Bilirubin in micromolar concentrations abolishes respiratory control, uncouples oxidative phosphorylation, and induces swelling of mitochondria. The bilirubin-induced swelling is irreversible and its amplitude is large. The swelling process requires a monovalent cation, any permeable anion, Mg++, and energy. The energy may be supplied from the oxidation of a substrate by the respiratory chain or from external ATP. Swelling measured by the decrease of absorbance at 590 mµ is parallel to the increase of water content and packed volume of mitochondria. In the absence of Mg++, a damped oscillation occurs which lowers the amplitude. The swelling is also accompanied by proton ejection to the medium. When ATP is used as an energy source, liberation of inorganic phosphate follows the pattern of swelling with a short initial lag. The Km value (50% of the maximum) of bilirubin for the over-all swelling is about 2.5 µm in liver mitochondria and is approximately 2 µm (about 0.1 mg of bilirubin per 100 ml) in brain mitochondria. Bovine serum albumin prevents uncoupling and swelling; estimation from the titration curves for either process gives a molar ratio of 1 for bilirubin to albumin. A probable role of bilirubin in affecting mitochondrial reactions is discussed.