IMAGING GENE EXPRESSION IN LIVING NEURAL TISSUE

Bowling Green State Univ Bowling Green, Bowling Green OH

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Abstract

Description (applicant's abstract): Gene expression can be followed continuously in living neural tissue by imaging bioluminescence produced from reporter genes that are based on the firefly enzyme luciferase and its substrate luciferin. This powerful, non-invasive technique can be modified to track gene activation in response to neural activity by combining the promoter for the immediate early gene c-fos with the luciferase gene. To help establish luminescence imaging in brain slice cultures as a standard neuroscience technique, an optical and tissue culture system is presented that is specifically designed for luciferase-based imaging. The penetration of luciferin into the brain slice and its influence on luminescence signals will be examined for the first time in transgenic mice that contain luciferase reporter genes. Software for providing confocal fluorescence images will be applied to bioluminescence image data. Finally, the imaging system will be validated using the c-fos/luciferase transgenic mouse and hypothalamic brain slices containing the circadian pacemaker that controls sleep and daily rhythms in behavior. Experiments will focus on imaging the interaction of the circadian pacemaker with adjacent areas. This approach has the potential of providing unparalleled views of interactions between brain regions which will complement established medical imaging techniques performed on the brain of intact subjects. Ultimately, the organization of the brain and temporal changes in brain function associated with several diseases states and behavioral disorders could be examined in animal models through in vitro bioluminescence imaging.