Albert Y. Sun - US grants

There is increasing experimental evidence to support the concept of free radical damage to cellular constituents. In general, an increase in free radical generation would lead to alterations of biochemical and biophysical properties of cells. The process of free radical generation may also be important in the pathological manifestations of aging and alcoholism. The overall objective of this project is to investigate the involvement of free radicals in brain membrane deterioration, and to examine whether alcohol- enhanced free radical formation will affect the aging process. Specifically, experiments will be designed to (1) test the hypothesis that free radicals play and important role in the aging process, and that ethanol accelerates aging process through potentiation or enhancement of the lipid peroxidation process; (2) evaluate specific membrane-dependent processes with respect to aging and chronic ethanol administration; (3) study the body protective system in brain against the toxic effects of free radical attack and (4) evaluate the importance of dietary antioxidants, such as vitamins E and C, in providing protection against the peroxidative damage resulted from alcohol ingestion. For experiments described in this project, we will continue to use C57BL/6NNIA mice as the animal model. The spin trapping technique will be employed to detect free radical formation in both in vitro and in vivo systems. In addition, the activity of enzymes involved in metabolism of the peroxidation process in brain, such as supreoxide dismutase, catalase, and glutathione peroxidase will also be investigated. Since many neuronal membrane processes are known to depend on the microenvironment of membrane lipids for proper functioning, we will specifically examine the effects of age and alcohol and (Na+, K+)-ATPase activity, enzyme its kinetics, phosphorylated intermediates and ouabain binding properties in mouse synaptic plasma membranes. In addition, other membrane-bound enzymes which are important in modulating membrane lipids will be examined and tested for their sensitivity to the radical formation. It is anticipated that through this systematic investigation, new information may be obtained regarding the mechanism of membrane deterioration due to age and/or alcohol intake, and the effectiveness of dietary supplement of vitamin E and vitamin C on alleviating some of the changes due to aging and/or alcohol abuse.

There is a general consensus that free radical generation leads to alterations of biochemical and biophysical properties of cell membranes and in turn, these changes may lead to pathological manifestations in aging and in degenerative diseases. Recent data from our laboratory has indicated that pro-oxidant agents may be involved in the pathogenesis of neurodegenerative diseases and that ethanol may potentiate these processes through participating in the free radical reaction. The overall objective of this project is to evaluate the hypothesis that the accelerated aging process and loss of neuronal function associated with alcohol drinking is related to the enhancement of free radical reaction by ethanol. Since activation of the glutamate receptor has been shown to cause global dementia and severe memory impairment similar to that in aging, we will use glutamate and other pro-oxidants (such as chelated iron and H2O2) as experimental tools to examine the effects of ethanol on the neurodegenerative processes. The specific aims of this proposal are: (1) to test the hypothesis that ethanol potentiates the free radical mechanism of neurodegenerative process; (2) to investigate the involvement of oxidative insults on the aging process, and to examine how alcohol affects aging through participation in the free radical reactions and (3) to investigate possible dietary and drug interventions against the alcohol-related damage as caused by glutamate and other pro-oxidant stressors. The effects of ethanol on pro-oxidant-induced cell death and tissue damage will be examined in PC-12 cells and in C57BI/6NNIA mice, respectively. Oxidative damage will be assessed by determining cell viability, formation of thiobarbituric acid reactive substances (TBARS), lactic acid dehydrogenase (LDH) release, membrane lipid composition and activity of membrane enzymes and uptake processes (e.g. Na-pump, catecholamine uptake, etc). In addition, the ability of antioxidants (such as vitamins E and C), and drugs (Ca2+-blockers, salicylate) to protect against oxidative damage elicited by ethanol will also be investigated. It is anticipated that through this investigation, important information on the mechanism of membrane deterioration in the brain due to aging and/or alcohol intake may be obtained and preventive measures for protecting against these changes may be proposed.