Testing Drosophila Olfaction with a Y-maze Assay

Summary

We present a choice test to reveal the influence of odorants on Drosophila behavior using a Y-maze assay.

Abstract

Detecting signals from the environment is essential for animals to ensure their survival. To this aim, they use environmental cues such as vision, mechanoreception, hearing, and chemoperception through taste, via direct contact or through olfaction, which represents the response to a volatile molecule acting at longer range. Volatile chemical molecules are very important signals for most animals in the detection of danger, a source of food, or to communicate between individuals. Drosophila melanogaster is one of the most common biological models for scientists to explore the cellular and molecular basis of olfaction. In order to highlight olfactory abilities of this small insect, we describe a modified choice protocol based on the Y-maze test classically used with mice. Data obtained with Y-mazes give valuable information to better understand how animals deal with their perpetually changing environment. We introduce a step-by-step protocol to study the impact of odorants on fly exploratory response using this Y-maze assay.

Introduction

Chemoreception through taste or olfaction is a key sensory modality for animal survival. It gives vital cues necessary for the detection of a danger or food sources, as well as for social interactions. It also helps animals to find a sex partner necessary for their reproduction. For more than 20 years, intensive research, including Nobel prize winning work by Richard Axel and Linda Buck in 2004 "for their discoveries of odorant receptors and the organization of the olfactory system", has been carried out to reveal the molecular and cellular bases of olfaction^1,2.

One of the favorite animal models for scientists to dissect....

Protocol

1. Before Starting

1. Use an isogenized reference stock bearing stable and robust behavioral phenotypes. There is no general rule for choosing this stock, since all potential controls may carry heterogeneous background alleles.
2. Use this control strain to backcross every other stock necessary for later steps. This backcrossing step is typically represented by at least 5 successive crosses of a single virgin female (to allow possible crossing-over between homologous chromosomes) to 2–3 isogenic reference males⁵. This step is important to homogenize the genetic background between the different fly stocks.
3. Maintain Drosoph....

Results

Figure 1D shows two representative responses using this Y-maze assay. Canton-S males strongly avoid 10% acetic acid diluted in distilled water, whereas they do not significantly avoid 10% phenylacetic acid. These assays are based on 10 males per replicate placed together in the loading vials. This protocol can sometimes lead to large standard error of the mean. If needed, it is possible to reduce this drawback by using 20 males per replicates instead of only 10. Mathematically, the choice of one individu.......

Discussion

Our Y-maze protocol is based on a previously described protocol⁹. However, we introduce two major differences. First, we use narrow pipette tips to prevent the flies from returning once they decide to enter in the vial containing the solvent or the solvent plus the odorant. These narrow tips are also useful to limit the odorant diffusion in the Y-maze. Second, we use a smaller loading vial to force the flies to enter in the Y-maze. It is important to have a high participation of these flies (80% to 100% after .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Drosophila Polystyrene tube	VWR europe	734-2255	30 x 25 mm  Y-maze
Drosophila Borosilicate tube	Dijon verre		95 X 25 mm Y-maze
Foam stopper	Dutscher	999038	Y-maze
Y-shaped connector	Europrix	11020605	Y-maze
100-1,000 µl pipette tips	Corning	4868	Join the following pipette tips to the Y-shaped connector. Cut 2 pipette tips at 65 mm from the wide end, and connect the narrow end (with a ∼2 mm opening) to 2 test vials. These openings will limit the U-turns once the flies enter the tubes containing the odors. Cut 1 pipette tip at 35 mm from the wide end, and connect it to the loading vial. Y-maze
Far-Red LED Bulb	Rubin-Lacaque	0RB180238	625-630 nm
Acetic Acid	Sigma-Aldrich	45725
Phenylacetic Acid	Sigma-Aldrich	P16621
Yeast	Sensient Flavors Strasbourg	1018880464
Cornmeal	eurogerm	Farine de maïs
Agar	Kalys	HP-697-25
Methyl hydroxy 4 benzoate	VWR international	25605293