Regulation by Glycogen Synthase Kinase-3β of the Arborization Field and Maturation of Retinotectal Projection in Zebrafish
Regulation by Glycogen Synthase Kinase-3β of the Arborization Field and Maturation of Retinotectal Projection in Zebrafish
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DOI：
10.1523/jneurosci.22-23-10324.2002
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发表时间：
2002-12
期刊：
影响因子：
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通讯作者：
H. Tokuoka;Tomoyuki Yoshida;Naoto Matsuda;M. Mishina
中科院分区：
文献类型：
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作者：
H. Tokuoka;Tomoyuki Yoshida;Naoto Matsuda;M. Mishina
视网膜顶投影是研究中枢神经系统中突触形成的分子基础的最佳系统之一，因为它具有良好的地形连接特征和活动依赖的细化。本研究利用烟碱乙酰胆碱受体β3 （nAChRβ3）基因启动子构建了斑马鱼视网膜顶突突触前神经元特异性基因操作系统。在携带achr β3基因启动子导向的EGFP表达载体的活转基因斑马鱼系中，绿色荧光蛋白（EGFP）表达信号增强，可视化了整个视网膜神经节细胞（RGC）轴突向顶盖投射的发育。微量注射achr β3基因启动子驱动的双盒载体，指导显性负糖原合成酶激酶-3β (dnGSK-3β)和EGFP的表达，使我们能够跟踪单个rgc的发育，并研究该分子对活斑马鱼同一神经元轴突树突和成熟的影响。我们发现，斑马鱼显性阴性形式GSK-3β的表达抑制了顶盖RGC轴突末端的乔木化场，这可以通过乔木分支长度和乔木面积的减少来估计。此外，GSK-3β活性的抑制增加了顶盖神经支配早期RGC轴突末端囊泡相关膜蛋白2-EGFP点的大小。这些结果表明GSK-3β在体内调节RGC轴突末端的树突范围和成熟。
The retinotectal projection is one of the best systems to study the molecular basis of synapse formation in the CNS because of the well characterized topographic connections and activity-dependent refinement. Here, we developed a presynaptic neuron-specific gene manipulation system in the zebrafish retinotectal projection in vivo using the nicotinic acetylcholine receptor β3 (nAChRβ3) gene promoter. Enhanced green fluorescent protein (EGFP) expression signals in living transgenic zebrafish lines carrying thenAChRβ3 gene promoter-directed EGFP expression vector visualized the development of entire retinal ganglion cell (RGC) axon projection to the tectum. Microinjection of thenAChRβ3 gene promoter-driven double-cassette vectors directing the expression of both dominant-negative glycogen synthase kinase-3β (dnGSK-3β) and EGFP enabled us to follow the development of individual RGCs and to examine the effect of the molecule on the axonal arborization and maturation of the same neurons in living zebrafish. We found that the expression of the dominant-negative form of zebrafish GSK-3β suppressed the arborization field of RGC axon terminals in the tectum as estimated by the reduction of arbor branch length and arbor areas. Furthermore, the suppression of GSK-3β activity increased the size of vesicle-associated membrane protein 2–EGFP puncta in RGC axon terminals at the early stage of innervation to the tectum. These results suggest that GSK-3β regulates the arborization field and maturation of RGC axon terminals in vivo.