Neuronal development and synaptic plasticity in Cdc42 cenditional knockout mice

为了研究Rho家族GT3基因Cdc42在突触形成和维持、突触可塑性和学习中的作用，我们产生了Cdc42条件性敲除小鼠。我们通过基因打靶的方法将loxP序列导入cdc42等位基因中，并在胚胎干细胞中进行表达。然后我们使用突变的ES细胞产生了flox-Cdc 42小鼠。通过将Cdc42小鼠与两种不同的神经元特异性Cre系杂交，我们产生了两种不同的神经元特异性Cdc42敲除系。(1)前脑特异性Cdc42基因敲除（FB-Cdc42 KO）小鼠为了避免胚胎死亡，我们用Emx1-Cre小鼠通过Cre-loxP重组破坏cdc42基因。Emx 1启动子/增强子诱导Cre重组酶仅在胚胎第10.5天（E）的端脑背侧表达，从而从大脑皮质形成开始消除皮质投射神经元中的Cdc 42表达。对从FB-Cdc 42 KO大脑皮质中制备的蛋白质进行蛋白质印迹分析表明，Cdc 42在KO大脑皮质中被敲低。我们发现FB-Cdc42基因敲除小鼠大脑皮层扩张，皮层形成异常。此外，FB-Cdc42 KO小鼠海马的形态学严重扭曲。这些结果表明，Cdc42在皮层发育过程中控制神经元的细胞增殖和分化。(2)浦肯野细胞特异性Cdc42基因敲除（PC-Cdc42 KO）小鼠为了研究Cdc42在小脑浦肯野细胞中的作用，我们用L7-Cre小鼠破坏了Cdc42基因。PC-Cdc42 KO小鼠未显示共济失调步态。PC-Cdc42 KO小脑的组织学分析显示浦肯野细胞树突的形态没有明显异常。

In order to investigate the role of Rho family GTPase, Cdc42, in synapse formation and maintenance, synaptic plasticity, and leaning, we generated Cdc42 conditional knockout mice. We introduced loxP sequences into cdc42 allele by gene targeting in the embryonic stem cells. Then we generated flox-Cdc42 mice by using the mutant ES cells. By crossing flox-Cdc42 mice with two different neuron-specific Cre lines, we generated two distinct neuron-specific Cdc42 knockout lines.(1) Forebrain-specific Cdc42 knockout (FB-Cdc42 KO) miceTo avoid embryonic lethality, we have disrupted cdc42 gene via Cre-loxP recombination using Emx1-Cre mice. Emx1 promoter/enhancer induces Cre recombinase expression exclusively in the dorsal telencephalon as early as embryonic day (E) 10.5, tcogehereby eliminating Cdc42 expression in cortical projection neurons from the beginning of cerebral cortinesis. Western blot analysis of the protein prepared from FB-Cdc42 KO cerebral cortex showed that Cdc42 was knocked down in the KO cerebral cortex. We have found expanded cerebral cortex with abnormal layer formation in the FB-Cdc42 KO mice. Furthermore, the morphology of hippocampus was severely distorted in the FB-Cdc42 KO mice. These results suggested that Cdc42 controls the cell proliferation and differentiation of the neuron during cortical development.(2) Purkinje cell specific Cdc42 knockout (PC-Cdc42 KO) miceTo investigate the role of Cdc42 in cerebellar Purkinje cells, we have disrupted cdc42 gene using L7-Cre mice. PC-Cdc42 KO mice did not show ataxic gait. The histological analysis of the PC-Cdc42 KO cerebellum showed no apparent abnormality in morphology of the Purkinje cell dendrites.