Name: dissection of oenocytes from adult drosophila melanogaster
Uploaded: 2010-07-18T17:00:00
Description: Watch this Scientific Journal Video about Dissection of Oenocytes from Adult Drosophila melanogaster at JoVE.com

We would like to thank Amsale Belay for her assistance in filming of this video article. This work was funded by CIHR, NSERC and CRC grants to JDL.

Part 1. Fillet preparation of the adult Drosophila abdomen.
The procedure outlined immediate below prepares the abdomen for the subsequent removal of the oenocytes (see Part2 of protocol). The same procedure can also be used for the preparation of the other tissues attached to the inner surface of the cuticle, including the dorsal vessel (i.e. heart) and fat body.
Note that in the video article the oenocyte dissection technique is demonstrated on an adult w...

<ol>
	<li>Wielowiejski, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ueber+das+blutgewebe+der+insekten.">Ueber das blutgewebe der insekten.</a> Zeit. Wiss. Zool. 43, 512-536 .</li><li>Krupp, J. J., Kent, C., Billeter, J. C., Azanchi, R., So, A. K., Schonfeld, J. A., Smith, B. P., Lucas, C., Levine, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Social+experience+modifies+pheromone+expression+and+mating+behavior+in+male+Drosophila+melanogaster.">Social experience modifies pheromone expression and mating behavior in male Drosophila melanogaster.</a> Curr. Biol. 18, 1373-1383 (2008).</li><li>Locke, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+membranes,+Golgi+complexes,+and+vacuolar+systems.">Surface membranes, Golgi complexes, and vacuolar systems.</a> Annu. Rev. Entomol. 48, 1-27 (2003).</li><li>Billeter, J. C., Atallah, J., Krupp, J. J., Millar, J. G., Levine, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specialized+cells+tag+sexual+and+species+identity+in+Drosophila+melanogaster.">Specialized cells tag sexual and species identity in Drosophila melanogaster.</a> Nature. 461, 987-991 (2009).</li><li>Gould, A. P., Elstob, P. R., Brodu, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insect+oenocytes:+a+model+system+for+studying+cell-fate+specification+by+Hox+genes.">Insect oenocytes: a model system for studying cell-fate specification by Hox genes.</a> J. Anat. 199, 25-33 (2001).</li><li>Lawrence, P. A., Johnston, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+lineage+of+the+Drosophila+abdomen:+the+epidermis,+oenocytes+and+ventral+muscles.">Cell lineage of the Drosophila abdomen: the epidermis, oenocytes and ventral muscles.</a> J. Embryol. Exp. Morphol. 72, 197-208 (1982).</li><li>Johnson, M. B., Butterworth, F. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maturation+and+aging+of+adult+fat+body+and+oenocytes+in+Drosophila+as+revealed+by+light+microscopic+morphometry.">Maturation and aging of adult fat body and oenocytes in Drosophila as revealed by light microscopic morphometry.</a> J. Morphol. 184, 51-59 (1985).</li><li>Gutierrez, E., Wiggins, D., Fielding, B., Gould, A. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specialized+hepatocyte-like+cells+regulate+Drosophila+lipid+metabolism.">Specialized hepatocyte-like cells regulate Drosophila lipid metabolism.</a> Nature. 445, 275-280 (2007).</li><li>Marcillac, F., Bousquet, F., Alabouvette, J., Savarit, F., Ferveur, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mutation+with+major+effects+on+Drosophila+melanogaster+sex+pheromones.">A mutation with major effects on Drosophila melanogaster sex pheromones.</a> Genetics. 171, 1617-1628 (2005).</li><li>Chertemps, T., Duportets, L., Labeur, C., Ueyama, M., Wicker-Thomas, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+female-specific+desaturase+gene+responsible+for+diene+hydrocarbon+biosynthesis+and+courtship+behaviour+in+Drosophila+melanogaster.">A female-specific desaturase gene responsible for diene hydrocarbon biosynthesis and courtship behaviour in Drosophila melanogaster.</a> Insect Mol. Biol. 15, 465-473 (2006).</li><li>Legendre, A., Miao, X. X., Da Lage, J. L., Wicker-Thomas, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolution+of+a+desaturase+involved+in+female+pheromonal+cuticular+hydrocarbon+biosynthesis+and+courtship+behavior+in+Drosophila.">Evolution of a desaturase involved in female pheromonal cuticular hydrocarbon biosynthesis and courtship behavior in Drosophila.</a> Insect. Biochem. Mol. Biol. 38, 244-255 (2008).</li><li>Shirangi, T. R., Dufour, H. D., Williams, T. M., Carroll, S. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+evolution+of+sex+pheromone-producing+enzyme+expression+in+Drosophila.">Rapid evolution of sex pheromone-producing enzyme expression in Drosophila.</a> PLoS Biol. 7, e1000168-e1000168 (2009).</li></ol>

In this video article we present a detailed dissection protocol for the preparation of the oenocytes from adult D. melanogaster in a manner suitable for molecular analysis. An abbreviated text-based account of the dissection method has been described elsewhere2, and the resulting oenocyte preparation has been demonstrated to be appropriate for the extraction of both RNA and protein2. Using this preparation it may also be possible to develop methods for the culturing of explanted oenocytes....

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Sylgard</td>
<td>&nbsp;</td>
<td>Dow Corning</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Tungsten Wire</td>
<td>&nbsp;</td>
<td>Ted Pella, Inc.</td>
<td>27-11</td>
<td>0.005&#x201D;/20&#x2019;</td>
</tr>
<tr>
<td>Forceps</td>
<td>&nbsp;</td>
<td>Dumont</td>
<td>11252-30</td>
<td>#5</td>
</tr>
<tr>
<td>Shields and Sang M3 Insect Medium</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>S3652</td>
<td>&nbsp;</td>
<tr>
<td>RNeasy Micro Kit</td>
<td>&nbsp;</td>
<td>Qiagen</td>
<td>74004</td>
<td>&nbsp;</td>
<tr>
<td>qScript cDNA Synthesis Kit</td>
<td>&nbsp;</td>
<td>Quanta Biosciences</td>
<td>95047-100</td>
<td>&nbsp;</td>
<tr>
<td>Monoject Hypodermic Needle</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>722289</td>
<td>20G1 with aluminum hub</td>
</tr>		</tbody>
		</table>
		</div>

dissection of oenocytes from adult drosophila melanogaster

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.

Watch this Scientific Journal Video about Dissection of Oenocytes from Adult Drosophila melanogaster at JoVE.com

Dissection of Oenocytes from Adult Drosophila melanogaster

In insects, the oenocytes produce cuticular hydrocarbon compounds. These compounds protect against desiccation and facilitate chemical communication. Here we demonstrate a dissection technique used to isolate the oenocytes from adult Drosophila melanogaster, and illustrate how this preparation can be utilized to study genes involved in hydrocarbon synthesis.

Dissection of Oenocytes from Adult Drosophila melanogaster

In Drosophila melanogaster, as in other insects, a waxy layer on the outer surface of the cuticle, composed primarily of hydrocarbon compounds, provides ...

Research

JoVE Journal

Biology

Dissection of Drosophila Ovaries

Dissection of Larval CNS in Drosophila Melanogaster

The central nervous system (CNS) of Drosophila larvae is complex and poorly understood.  One way to investigate the CNS is to use immunohistochemistry to ...

Dissection of Imaginal Discs from 3rd Instar Drosophila Larvae

Whole Cell Recordings from Brain of Adult Drosophila

In this video, we demonstrate the procedure for isolating whole brains from adult Drosophila in preparation for recording from single neurons. We begin by ...

High-Resolution Video Tracking of Locomotion in Adult Drosophila Melanogaster

Flies provide an important model for studying complex behavior due to the plethora of genetic tools available to researchers in this field.  Studying ...

Dissection of Hippocampal Dentate Gyrus from Adult Mouse

The hippocampus is one of the most widely studied areas in the brain because of its important functional role in memory processing and learning, its ...

Dissection of Organs from the Adult Zebrafish

Over the last 20 years, the zebrafish has become a powerful model organism for understanding vertebrate development and disease.  Although experimental ...

Isolation of Drosophila melanogaster Testes

Isolation of Drosophila melanogaster Testes

The testes of Drosophila melanogaster provide an important model for the study of stem cell maintenance and differentiation, meiosis, and soma-germline ...

Insulin Injection and Hemolymph Extraction to Measure Insulin Sensitivity in Adult Drosophila melanogaster

Insulin Injection and Hemolymph Extraction to Measure Insulin Sensitivity in Adult Drosophila melanogaster

Conserved nutrient sensing mechanisms exist between mammal and fruit fly where peptides resembling mammalian insulin and glucagon, respectively function ...

Dissection and Immunostaining of Imaginal Discs from Drosophila melanogaster

Dissection and Immunostaining of Imaginal Discs from Drosophila melanogaster

A significant portion of post-embryonic development in the fruit fly, Drosophila melanogaster, takes place within a set of sac-like structures called ...