Krupp Joshua J, Levine Joel D
Department of Biology, University of Toronto.
J Vis Exp. 2010 Jul 18(41):2242. doi: 10.3791/2242.
In Drosophila melanogaster, as in other insects, a waxy layer on the outer surface of the cuticle, composed primarily of hydrocarbon compounds, provides protection against desiccation and other environmental challenges. Several of these cuticular hydrocarbon (CHC) compounds also function as semiochemical signals, and as such mediate pheromonal communications between members of the same species, or in some instances between different species, and influence behavior. Specialized cells referred to as oenocytes are regarded as the primary site for CHC synthesis. However, relatively little is known regarding the involvement of the oenocytes in the regulation of the biosynthetic, transport, and deposition pathways contributing to CHC output. Given the significant role that CHCs play in several aspects of insect biology, including chemical communication, desiccation resistance, and immunity, it is important to gain a greater understanding of the molecular and genetic regulation of CHC production within these specialized cells. The adult oenocytes of D. melanogaster are located within the abdominal integument, and are metamerically arrayed in ribbon-like clusters radiating along the inner cuticular surface of each abdominal segment. In this video article we demonstrate a dissection technique used for the preparation of oenocytes from adult D. melanogaster. Specifically, we provide a detailed step-by-step demonstration of (1) how to fillet prepare an adult Drosophila abdomen, (2) how to identify the oenocytes and discern them from other tissues, and (3) how to remove intact oenocyte clusters from the abdominal integument. A brief experimental illustration of how this preparation can be used to examine the expression of genes involved in hydrocarbon synthesis is included. The dissected preparation demonstrated herein will allow for the detailed molecular and genetic analysis of oenocyte function in the adult fruit fly.
在黑腹果蝇中，与其他昆虫一样，表皮外表面的一层蜡质层主要由碳氢化合物组成，可提供防止脱水和应对其他环境挑战的保护。这些表皮碳氢化合物（CHC）中的几种还作为信息化学信号起作用，因此介导同一物种成员之间或在某些情况下不同物种之间的信息素通讯，并影响行为。被称为oenocytes的特化细胞被认为是CHC合成的主要部位。然而，关于oenocytes在调节有助于CHC输出的生物合成、运输和沉积途径中的参与情况，人们了解得相对较少。鉴于CHCs在昆虫生物学多方面发挥的重要作用，包括化学通讯、抗脱水和免疫，更深入了解这些特化细胞内CHC产生的分子和遗传调控非常重要。黑腹果蝇的成年oenocytes位于腹部体壁内，呈节段性排列成带状簇，沿着每个腹部节段的内表皮表面辐射状分布。在这篇视频文章中，我们展示了一种用于从成年黑腹果蝇制备oenocytes的解剖技术。具体来说，我们详细分步演示了（1）如何去骨制备成年果蝇腹部，（2）如何识别oenocytes并将它们与其他组织区分开来，以及（3）如何从腹部体壁中取出完整的oenocyte簇。还包括一个关于如何使用这种制备方法来检测参与碳氢化合物合成的基因表达的简要实验示例。本文展示的解剖制备方法将有助于对成年果蝇中oenocyte功能进行详细的分子和遗传分析。
