In response to the Research Objective 14 (Functional Senescence) Elevated levels of reactive oxygen species contribute to the age-related functional decline of many organs and are associated with stroke and major neurodegenerative disorders such as Parkinson's and Alzheimer's diseases. Therefore, techniques for constant evaluation of free radical generation in vivo are required. However, presently there are no such methods. As a result it is impossible to monitor the changes in the levels of free radicals occurring in aging brain. In the current project, we will enlarge the technology base available to study free radical generation in vivo using the new fullerene-based nanosensors. Fullerenes were discovered by a Nobel laureate Richard Smalley in1985 and are physiologically inert. The long-term objective of this proposal is to develop a new approach to in vivo imaging of free radical generation, and apply it for the analysis of age-related changes in free radical generation in brain.
Specific aims of this project are: 1) To develop a new approach to image intracellular free radical reactions utilizing fullerenes. Use magnetic resonance imaging (MRI) as a reporter method. 2) Apply the new approach for detection and monitoring the levels of free radicals in brains of young and old rats. Compare the free radical levels generated in response to transient brain ischemia in young and old rats. The technique utilizes a double-cage principle by encaging the MRI tracer into fullerenes with subsequent covering of the loaded fuilerenes by a free-radical sensitive polymer. The outer coating allows for transmembrane delivery and prevents release of the caged tracer. Degradation of the cover results in the tracer release, signaling the generation of free radicals. We collaborate with Prof. Richard Smalley on the development of fuilerene nanosensors and after completion of this task we will study the time course and regional localization of free radical generation induced in rat brain by experimental stroke (permanent occlusion of carotid artery). The experiments will be carded out in young (6 months) and old (24 months) rats (ad lib fed and group housed inbred F344tNnia). Initially the levels of free radicals will be analyzed using fluorescently labeled tracers and animals will be sacrificed. At a later stage free radicals will be monitored using magnetic resonance tracer and MRI detection.
