Oxidation of Bilirubin by Brain Mitochondrial Membranes—Dependence on Cell Type and Postnatal Age

Abstract

Bilirubin is oxidized by brain mitochondrial membranes at a rate which may contribute significantly to clearance of bilirubin from the brain. Neurons appear to be more sensitive to bilirubin toxicity than glial cells. Clinical experience has suggested that sensitivity to bilirubin neurotoxicity may be greater in the neonate than later in life. We hypothesized that the ability to oxidize bilirubin would be lower in mitochondrial membranes from a pure neuronal compared to a mixed glial/neuronal source, and lower in immature than more mature brains. Mitochondria of synaptosomal and nonsynaptosomal origin were obtained from young rat brains by differential centrifugation in sucrose gradients. Synaptosomes were lysed by hypoosmotic treatment, and mitochondria were ruptured by sonication. The change in optical density of a bilirubin solution at 440 nm was measured over time following addition of the membrane suspension. The rate of bilirubin oxidation was significantly higher in nonsynaptic than in synaptic mitochondrial membranes [99.1 ± 42.3 (mean ± SD) vs 69.9 ± 30.9 pmol/min/mg protein,t= 4.835,P= 0.0003]. “Crude” mitochondrial membranes were obtained by differential centrifugation in sucrose from the forebrains of rats of 7, 14, and 21 days postnatal age as well as adults, and from rabbits of 1 and 7 days postnatal age as well as adults. In both species the rates of bilirubin oxidation increased significantly with postnatal age (rats:F= 55.3,P< 0.0001; rabbits:F= 101,P< 0.0001). Mitochondrial membranes from a pure neuronal source oxidize bilirubin at a significantly lower rate than membranes from a mixed glial/neuronal source. This suggests that neurons may be less able to detoxify bilirubin locally and thus might contribute to the apparent higher sensitivity to bilirubin toxicity in these cells vs glia. Similarly, the lower bilirubin-oxidizing ability of mitochondrial membranes from immature brains seems compatible with the clinical impression of increased vulnerability to bilirubin neurotoxicity in the newborn.