Research reportFree radical-induced elevation of ornithine decarboxylase activity in developing rat brain slices
Introduction
Fetal and neonatal hypoxia and cerebral ischemia are widely held to contribute to developmental abnormalities, central nervous system dysfunction, neurological handicaps as well as increased neonatal morbidity and mortality 5, 19, 24, 35. Nonetheless, despite significant advances in understanding the basic mechanisms of hypoxic-ischemic brain damage in immature animals [43], a complete understanding of the pathophysiology of these changes has yet to be established [25].
Several reports have provided evidence of a strong link between oxidative stress and brain damage 8, 22, 23. The brain is believed to be particularly vulnerable to oxidative damage, in part, because it contains high concentrations of easily peroxidizable lipids (i.e. it is rich in polyunsaturated fatty acids), it has regions that are particularly enriched in iron and it has comparatively low-to-moderate levels of endogenous antioxidant molecules [33].
Ornithine decarboxylase (ODC) and polyamine concentrations are relatively high in developing neural tissues 26, 40and are required for cell growth and proliferation 11, 14. ODC is the rate-limiting enzyme in the biosynthesis of the positively charged polyamines which play key roles in DNA, RNA and protein synthesis. Recently, we have shown, both in vivo 26, 32and in vitro [31], that hypoxia increases ODC activity and polyamine concentrations in fetal and neonatal rat brain. It is not known, however, to what extent this hypoxic-associated increase in ODC activity is mediated by oxygen free radicals. The aim of this study was to test the hypothesis that free radicals are associated with the hypoxic-induced increase in ODC activity in the developing rat brain.
Section snippets
General
We used brain tissue from over 300 male and female 3–4-day-old Sprague-Dawley rat pups. We chose this age group because of our previous studies on ODC activity and hypoxia 26, 31, 32. Pregnant and nursing dams (Charles River, Portage, MI), on a regular 12/12-h (light/dark) cycle, were housed one per cage (40×20×18 cm) and given free access to food and water as we have previously described 16, 26. Animals were maintained in fully accredited facilities and in accord with all United States
Effect of oxygen free radicals on ODC activity
To examine the slice ODC activity as a function of time, we measured ODC activity for 2 h after the initial 1-h equilibration. As shown in Fig. 1, under control conditions, ODC activity was stable during the 2-h post-recovery period. In response to 5×10−7 M xanthine+10 mU XO, ODC activity increased 230% at 1.5 h post-recovery. In contrast, in response to 1.25×10−4 M xanthine+10 mU XO, ODC activity decreased gradually, being reduced almost 60% by the end of the 2-h post-recovery period (Fig. 1).
General
Brain slice preparations from adult rat, guinea-pig, rabbit or mouse are now widely utilized for a variety of neurobiological investigations 1, 2, 6, 27, 37, 38. We have previously used neonatal rat brain slices to demonstrate acute hypoxic-induced increase in ODC activity [31]. In the present study, we used the neonatal brain slice preparation to examine the role of free radicals in the induction of ODC activity.
Effect of oxygen free radicals on ODC activity
To examine free radical-induced perturbation of ODC activity by reactions which
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