The molecular changes that cause neuronal death following various insults to the brain such as stroke and trauma have not been established. However, oxidative stress and excessive generation of free radicals appear to accompany many brain insults. Free radicals are highly reactive and can potentially cause cell injury by inducing protein oxidation, lipid peroxidation, and DNA damage. The excitotoxic injury that may accompany ischemia can also generate DNA damage, but little is known concerning how neuronal cells respond to DNA damage and the importance of such a response to neuronal viability. The characteristics of certain genetic diseases involving DNA repair deficits, such as xeroderma pigmentosum (XP), suggests that the neuronal response to DNA damage is important. Patients with XP have severe neurologic abnormalities and neuronal degeneration, but the cause of selective vulnerability of neurons to defects in DNA repair is unclear. A better understanding of the role of DNA damage in neuronal injury is needed to aid in defining methods to ameliorate such injury. In the present study, we will use radiation to induce free radicals and DNA damage in neurons and astrocytes growing in vitro. The proposed studies will quantify and compare the response of neurons and astrocytes to radiation-induced DNA damage in terms of the ability of these cells to repair damage and whether the cells undergo apoptosis or necrosis. Repair of DNA damage will be quantified using pulsed-field gel electrophoresis. The studies will also compare the response of neurons to DNA double-strand breaks versus lipid peroxidation in order to determine whether DNA damage specifically can lead to neuronal death. Finally, the studies will use agents that alter the concentration of free radicals to determine if modulation of oxidative stress can modify the response of neurons to radiation. Such studies will provide a basis for a better understanding of the pathophysiology of neuronal injury and will supply the background needed in future studies to develop methods to ameliorate that damage.
