| Research Abstract |
The methods of gene targeting have greatly contributed to understanding of the brain function. However, it has also become clear that the methods have some limitations. 1)Its innate molecular loss hinders normal development and causes death, making analysis impossible. 2)Abnormal formation of tissues and organs caused by knockout (for instance, abnormal nerve network etc.) does not correctly reflect the molecular function in adults. 3)Functional compensation could be made by other molecules prohibiting correct evaluation. Moreover, knockout mice by other molecules prohibiting correct evaluation. Moreover, knockout mice made by prevalently used 129 strain-ES cells have a congenital hypoplasia in the corpus callosum, for which they are unsuitable for behavioral analysis. In order to solve these problems, we have developed a new system. As the first step, we established a method of producing a gene-manipulated mouse using the C57BL/6 origin embryonic stem cell and applied its results for a patent. To elucidate the physiological function of GluRε2 in adulthood, a conditional knockout mouse was generated using the Cre/loxP recombination system. We foxed the M4-containing exon of GluRε2f and GRγ1Cre mouse strain, which expressed Cre in the hippocampal CA3 region. GRε2f and GRγ1Cre mice were crossed to obtain the GluRε2 conditional targeting mice. Hippocampal CA3 pyramidal cells mainly receive three distinct excitatory inputs, so we analyzed their synapses electrophysiologically in the mutant mouse. NMDA receptor responses were hardly detected at any of the synapses, though GluRε1 existed in these regions. These results suggest that GluRε2 is crucial to form functional NMDA receptor in the hippocampal CA3 specific deficient mice. Their growth and mating were normal, but learning ability of the contextual fear conditioning was increased.
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