Elucidation of the synapse formation mechanism with glutamatergic signaling

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AMPA-type glutamate receptors were composed of four subunits, GluR1, GluR2, GluR3 and GluR4 and the AMPA receptors lacking GluR2 shows high Ca^<2+>-permeability. The roles of GluR2, of which the genetical symbol is GRIA2, in process formations in neurons and glial cells were examined using GRIA2-knock out mice, obtained form the Jackson Laboratory (Maine, USA). Although the knockout mice of GluR2 were introduced into Gunma University in 2004 and the propagation was made from the beginning of this project, the mice exhibited poor breeding rates. In heterozygote mice, GRIA2(+/-), One of two 11th exons of GRIA2 was substituted to a neomycin resistance gene by homologous recombination. Embryos of wild type mice, GRIA2(+/+), and knock-out mice, GRIA2(-/-) were obtained as follows. Females and males GRIA2(+/-) mice were mated overnight and separated the next morning and checked for the presence of gastration plug. This will be counted as gastration day 0.5(E0.5). Dissociated hippocampal culture was prepared from pups E16.5 or E17.5, killing the mother these embryos on the pregnancy 16.5 _17.5 days. For genotyping, PCR were performed according to the protocol of Jackson Laboratory Neurites of cultured neurons were thinner in GRIA2(-/-). The numbers of dendrites were fewer. In addition the spines of dendrites in glutamatergic neurons in GRIA2(-/-) mice were smaller than that in wild type. Since the limited periods of this grant, immunohistochemical analysis and electrophysiological experiments were not performed in the present study, remaining many subjects in the further study.