Methods to Assay Drosophila Behavior

Summary

Drosophila melanogaster is a genetically and behaviorally tractable model system that has been used to understand the molecular and cellular basis of many important biological processes for over a century ¹. Drosophila has been well exploited to gain insights into the genetic basis of fly behavior.

Abstract

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases¹. We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials^2-4. The rapid iterative negative geotaxis (RING) assay⁵ has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Protocol

1. Larval Crawling Assay

1. Larvae Collection

1. Set up an 8 ounce bottle of flies (10-15 males + 10-15 females).
2. Let flies lay eggs for 24 hours, then clear bottle of flies. (Transfer the adults into a new bottle and repeat as necessary).
3. Incubate bottle for 3-4 days, or until third instar larvae are visible.
4. Add 50 - 100 ml of 20% sucrose to the bottle with larvae and let sit for 20 minutes. Larvae will float to the top.
5. Collect larvae using a 25 ml serological pipette with the tip cut off, and place into a mesh basket.
6. Wash larvae in the mesh basket two times with deionized H_2<....

Discussion

Drosophila behavior is tightly regulated by genetic and environmental factors. We, and others, have previously used the assays described here to gather data to examine genes related to fly behaviors and to human neurodegenerative diseases modeled in Drosophila^5-19. For the crawling assay, careful selection of 3^rd instar larvae is a critical step. If treating with a drug, it will take 10-15 minutes (or more depending on the type and nature of the drug) to achieve maximal effect if i.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Sucrose	Fisher Scientific	S5-500
Agarose	Invitrogen	16500-500
6 oz Drosophila bottle	Genesee Scientific	32-130
Paint Brush (#1)	Ted Pella, Inc.	11859
Cornmeal	Fisher Scientific	NC9109741
Agar	Genesee Scientific	66-104
Molasses	Fisher Scientific	NC9349176
Propionic acid	Acros Organics	14930-0010
Tegosept	Apex	20-258
Ethanol	Fisher Scientific	BP2818-4
Yeast	Genesee Scientific	62-107