Labelling and targeted ablation of specific bipolar cell types in the zebrafish retina
Labelling and targeted ablation of specific bipolar cell types in the zebrafish retina
复制标题
斑马鱼视网膜中特定双极细胞类型的标记和靶向消融
作者：
Xiao;S. Ellingsen;A. Fjose
功能视网膜的发育取决于几种神经元的调节分化和不同类型神经元之间高度复杂网络的产生。此外，每种类型的视网膜神经元包括几种不同的形态类型。我们对产生这种视网膜神经元多样性的机制知之甚少，这些神经元也可能表现出特定的区域分布模式。结果在斑马鱼的筛选中，我们使用携带工程酵母Gal4转录激活因子的捕获载体和UAS:eGFP报告盒，在视网膜双极细胞的特定亚群中鉴定出了两种转基因斑马鱼系，它们共同表达eGFP和Gal4。egfp标记有助于分析成人视网膜内丛状层内的轴突终末，并表明荧光双极细胞符合先前定义的形态类型。在成年斑马鱼中观察到egfp阳性双极细胞对视神经周围视网膜中央部分的强烈区域限制。此外，我们利用Gal4-UAS控制下的细菌硝基还原酶基因联合前药甲硝唑，在5天龄的幼虫中实现了标记双极细胞的特异性消融。在药物前治疗后，表达硝基还原酶的双极细胞被有效地消融，而不影响周围的视网膜结构，并且由于新的双极细胞的产生增加，在几天内恢复。结论增强子捕获可用于标记斑马鱼视网膜双极细胞的不同形态类型。这些细胞的遗传标记产生了修饰的Gal4转录激活子和荧光标记物eGFP的共表达。我们的工作还证明了Gal4-UAS系统在诱导其他转基因方面的潜在用途，包括细菌硝基还原酶融合基因，它可以促进双极细胞分化的分析，以及视网膜如何从这些细胞的特异性消融中恢复。
BackgroundDevelopment of a functional retina depends on regulated differentiation of several types of neurons and generation of a highly complex network between the different types of neurons. In addition, each type of retinal neuron includes several distinct morphological types. Very little is known about the mechanisms responsible for generating this diversity of retinal neurons, which may also display specific patterns of regional distribution.ResultsIn a screen in zebrafish, using a trapping vector carrying an engineered yeast Gal4 transcription activator and a UAS:eGFP reporter cassette, we have identified two transgenic lines of zebrafish co-expressing eGFP and Gal4 in specific subsets of retinal bipolar cells. The eGFP-labelling facilitated analysis of axon terminals within the inner plexiform layer of the adult retina and showed that the fluorescent bipolar cells correspond to previously defined morphological types. Strong regional restriction of eGFP-positive bipolar cells to the central part of the retina surrounding the optic nerve was observed in adult zebrafish. Furthermore, we achieved specific ablation of the labelled bipolar cells in 5 days old larvae, using a bacterial nitroreductase gene under Gal4-UAS control in combination with the prodrug metronidazole. Following prodrug treatment, nitroreductase expressing bipolar cells were efficiently ablated without affecting surrounding retina architecture, and recovery occurred within a few days due to increased generation of new bipolar cells.ConclusionThis report shows that enhancer trapping can be applied to label distinct morphological types of bipolar cells in the zebrafish retina. The genetic labelling of these cells yielded co-expression of a modified Gal4 transcription activator and the fluorescent marker eGFP. Our work also demonstrates the potential utility of the Gal4-UAS system for induction of other transgenes, including a bacterial nitroreductase fusion gene, which can facilitate analysis of bipolar cell differentiation and how the retina recovers from specific ablation of these cells.
影响因子：
2.7
作者：
Davison, Jon M.;Akitake, Courtney M.;Parsons, Michael J.
通讯作者：
Parsons, Michael J.
影响因子：
2.7
作者：
Bernardos, RL;Lentz, SI;Raymond, PA
通讯作者：
Raymond, PA