Elucidation of the synapse formation mechanism with glutamatergic signaling

AMPA型谷氨酸受体由GluR1、GluR2、GluR3和GluR4四个亚基组成，缺乏GluR2的AMPA受体表现出高的钙离子通透性。利用来自美国缅因州杰克逊实验室的GRIA2基因敲除小鼠，研究了GluR2(其遗传符号为GRIA2)在神经元和神经胶质细胞过程形成中的作用。尽管GluR2基因敲除小鼠于2004年被引入群马大学，并从该项目开始就进行了繁殖，但这些小鼠的繁殖率很低。在杂合子小鼠中，GRIA2的两个第11外显子之一的GRIA2(+/-)通过同源重组被替换为新霉素抗性基因。获得了野生型小鼠GRIA2(+/+)和基因敲除小鼠GRIA2(-/-)的胚胎。雌性和雄性GRIA2(+/-)小鼠交配过夜，第二天早上分开，检查是否存在断断塞。这将被算作绞刑第0.5天(E0.5)。取E16.5或E17.5仔鼠制备分离的海马区培养物，在妊娠16.5~17.5天处死母鼠。根据Jackson实验室的方法进行基因分型，培养神经元的突起在GRIA2(-/-)中较薄。枝晶数量较少。此外，GRIA2(-/-)小鼠谷氨酸能神经元的树突较野生型小。由于这笔赠款的期限有限，本研究没有进行免疫组织化学分析和电生理实验，许多对象留在了进一步的研究中。

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.