Generation of Brain Subregion-Restricted Conditional

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The enormous complexity of the brain is derived from hundreds of neuronal cell types and extensive synaptic connections between them. Studies of the localized function of the brain-subregions have traditionally been facilitated by the various brain lesion techniques such as aspiration, electrical or ibotenic acid lesions. However, these procedures often caused neuronal degeneration of the projecting axons, potentially resulting in the impaired function of the projection area as well. Pharmacological intervention, such as infusion of TTX, sodium channel blocker, and muscimol, GABA agonist, into brain subregions, has also been utilized to elucidate their localized functions. This approach is more molecule selective and temporally controllable in a reversible manner. However, the drug administration into the brain subregion does not distinguish specific cell types and often lead to cellular damage and toxicity. To overcome these limitations, a new conditional transgenic technology has been revolutionized by the development of genetic engineering that ideally switches gene expression on and off in a particular cell-type of certain brain subregion in vivo. For example, Cre recombinase of the P1 bacteriophage has proven invaluable for conditional transgenic manipulation in post-mitotic neuronal cells of the adult brain. Tetracycline responsive system has also been shown to be useful in the brain in a reversible manner. Toward understanding of the significance of brain subregions in higher cognitive functions, such as learning and memory, emotional state including anxiety and fear, attention, and awareness, we initiated a project to create a variety of brain-subregion or cell-type restricted conditional transgenic mice. Since any particular brain functions are affected by many brain subareas, we are focusing onto several brain subregions, such as hippocampal CA1/CA3, amygdala, entorhinal cortex, prefrontal cortex, nucleus accumbens and the ventral tegmental area. The key issue of this project is the choice of genetic promoter which determines the cell type or brain subarea specificity of transgene expression. Among them, a BAC (bacterial artificial chromosome) clone carrying a promoter for hippocampal CA3 restricted expression is in our hand from our previous study. We have further identified most of BAC clones that carries promoter for the gene expression predominantly in the above brain subregion, respectively. Currently, my colleagues and I are making DNA constructs for generation of transgenic mice in which Cre recombinase will be predominantly expressed in the above brain subregions. Concurrent with this effort, by combining Cre/loxP system, we are trying to establish a genetic knockdown system of protein synthesis in a cell-type specific manner. In a few years, we expect to establish a variety of brain subregion restricted genetic manipulation system that hopefully provides incredibly valuable animal tools for understanding of the function of each brain subregion at a system level. We also hope that this study leads to make fundamental discoveries critical to our understanding of the most serious neuropsychiatric disorders, such as bipolar disorders and schizophrenia.