Monitoring Neuronal Activity and Inducible Gene Expression using MRI

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DESCRIPTION (provided by applicant): Over the last two decades, reporter genes have contributed tremendously to biomedical research, and have revolutionized the way many experiments are designed and carried out. The major applications include: (i) monitoring the level of gene expression; (ii) investigating dynamic molecular interactions between proteins; (iii) studying cellular interactions (such as neuronal networks); (iv) tracking cell fates in normal and abnormal development, immunological processes, and in (stem) cell transplantation therapy; and (v) monitoring gene therapy. Availability of reporter genes that can be detected non-invasively using magnetic resonance imaging (MRI) would be particularly attractive, since it would allow co-registration of the site of gene expression with (high resolution) anatomical structures as well as with functional information. In this project, we propose to test the hypothesis that neuronal activation-induced gene expression can be monitored using chemical exchange saturation transfer (CEST) MRI. This MRI contrast is based on the transfer of radiofrequency saturation from the reporter's exchangeable protons to water. Since different CEST MRI reporters can be designed such that each will be responsive to unique radiofrequencies, our new imaging approach should allow simultaneous imaging of the expression of multiple genes. CEST-based MRI reporter genes such as lysine rich protein (LRP) will be cloned and expressed under the regulation of the inducible promoter c-fos and the constitutive cytomegalovirus (CMV) promoter. While the CMV promoter drives constant levels of gene expression, the genes under the c-fos promoter will be expressed only in response to an extracellular stimulation. In the brain, c-fos is induced in neurons in response to activation by various stimuli such as stress and somatosensory stimulation. The reporter expression will be evaluated in vitro in cell cultures, and in vivo after transduction into regions of the rat brain using a lentiviral vector. Following neuronal activation using kainic acid injection and forepaw stimulation, the CEST MRI measurements will be correlated with changes in neuronal activity as measured with functional MRI (fMRI). If these studies are completed successfully, it will represent a unique way to study brain plasticity in vivo. Non-invasive CEST MRI during differential expression of multiple genes may potentially lead to visualizing and identifying pathways that underlie common neurological disorders which may further help in designing improved treatments. PUBLIC HEALTH RELEVANCE: In the healthy brain as well as in many brain disorders neuronal activity is associated with dynamic changes in gene expression. Following the expression of multiple genes using noninvasive technologies could tremendously promote the identification of pathways that underlie many neurological disorders and could also lead to improved treatments. The objective of this study is to develop a platform for the noninvasive imaging of gene expression in concert with neuronal function in real time. Gene expression will be monitored using artificial reporters genes designed specifically for Magnetic Resonance Imaging (MRI) and neuronal activity using functional MRI (fMRI).