Oxygen Free Radical Generation During in-Utero Hypoxia in the Preterm and Term Fetal Guinea Pig Brain. † 966

Abstract

Previous studies have shown that hypoxia increases the generation of oxygen free radicals in the brain of the term guinea pig fetus. The present study tests the hypothesis that free radical production during hypoxia changes with each stage of brain development. Pregnant guinea pigs at 40 days (preterm, n=12) and 60 days (term n=14) gestation were assigned to normoxic or hypoxic groups and exposed to 21% or 7% oxygen, respectively, for 60 minutes. Fetal cerebral cortical tissue was homogenized in N2 bubbled ice-cold 100 mMα -phenyl-N-tert-butylnitrone (PBN) to trap oxygen free radicals. Trapped PBN spin adducts were extracted in toluene and free radical generation was measured by electron spin resonance spectroscopy (ESR) performed at-20°C. Peak height of the spin adduct signal was used as an index of the intensity of free radical formation and expressed as units/g dry tissue. Fetal brain hypoxia was documented biochemically by decreased levels of ATP and phosphocreatine. In the preterm group, cortical tissue from hypoxic fetuses showed an increase in spin adducts (normoxic:41.4±3.5 units/g tissue vs. hypoxic:55.1±6.4 units/g tissue, 33% increase, p<0.01). In the term group, cortical tissue from hypoxic fetuses also showed a significant increase in spin adducts (normoxic: 33.2±7.3 units/g tissue vs. hypoxic: 57.9±9.2 units/g tissue, 71% increase, p<0.01). Baseline free radical generation during normoxia was 23% higher in preterm than term. The higher baseline in the normoxic preterm brain may reflect decreased levels of antioxidant enzymes at this stage of gestation. However, the greater magnitude of change in free radicals following hypoxia in the term fetus suggests that susceptibility to hypoxia-induced free radical injury increases with brain maturation. We speculate that the lower level of free radical generation during hypoxia in the immature brain may be due to the lower concentrations of membrane highly unsaturated fatty acids available as a source of free radical production, and/or to the underdevelopment of free radical generating mechanisms.