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Neuroscience

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Published: April 20th, 2013

DOI:

10.3791/50237

1Department of Biology, Institute of Cell and Developmental Biology, University of Fribourg

Here we describe a light-dark preference test for Drosophila larva. This assay provides information about innate and circadian regulation of light sensing and processing photobehavior.

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer basic questions on how light information is processed and shared between rapid and circadian behaviors. Drosophila larvae display a stereotypical avoidance behavior when exposed to light. To investigate light dependent behaviors comparably simple light-dark preference tests can be applied. In vertebrates and arthropods the neural pathways involved in sensing and processing visual inputs partially overlap with those processing photic circadian information. The fascinating question of how the light sensing system and the circadian system interact to keep behavioral outputs coordinated remains largely unexplored. Drosophila is an impacting biological model to approach these questions, due to a small number of neurons in the brain and the availability of genetic tools for neuronal manipulation. The presented light-dark preference assay allows the investigation of a range of visual behaviors including circadian control of phototaxis.

Here we describe a behavioral assay based on the larval preference for dark (or light). Larvae react with a strong and stereotypic photonegative response during foraging stages (L1 to early L3)1. The assay is aimed to assess the photophobic behavior of the larva and compares the light or dark preference of a group of larvae moving freely in a Petri dish coated with agar. This behavioral assay not only provides information about the sensitivity, integration and temporal plasticity of the visual system, it further provides hints on how light sensitivity and its process is controlled by the circadian system.

The Drosophila....

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1. Larval Rearing

  1. Keep fly strains or genetic crosses in mass culture at 25 °C on corn meal medium under a 12-hr light-12-hr dark cycle in a fly incubator equipped with light and timer.
  2. Dilute backer yeast in water to form a fluid paste (10 g of backer yeast diluted with 3-4 ml distilled H2O). Add a small drop to the corn meal food and cover the vials. Let dry for at least one hour to avoid adult flies sticking to the yeast paste. Putting a small drop of yeast diluted in water on th.......

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Following the protocol described above, we tested light-dark preference in early third larval stadium of wild type Canton-S flies at two different circadian times CT0 and CT12. Adults were reared 12-hr light-12-hr dark and left to lay eggs for 12 hr. Larvae grow the first two days under the same light-dark regime. Since we wanted to test circadian modulation under constant conditions (free running of the circadian clock), larvae were then transferred to constant darkness for the next three days until test was performed .......

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The light preference test described takes advantage of the larval innate photobehavior. The assay is easy to establish, allows many repetitions at low cost and delivers valuable information about light sensing and processing. The experimental paradigm allows relatively quick quantification of how many individuals prefer light or dark. Such preference can be displayed as crude percentages or alternatively as Preference Index (PREF). The PREF is expressed as the difference of animals that preferred light and animals that p.......

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We thank our colleagues at the Department of Biology, University of Fribourg for fruitful discussions. We thank the Bloomington Stock Center for providing fly stocks. This work was financially supported by the Swiss National Science Foundation (PP00P3_123339) and the Velux Foundation to S.G.S.

....

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Name Company Catalog Number Comments
Name of Reagent/Material Company Catalog Number Comments
Agar Sigma-Aldrich A5093-500G 2.5%; Sigma-Aldrich, 9471 Buchs, Switzerland
Petri dishes Greiner Bio-One GmbH 633180 90-mm diameter; Greiner Bio-One GmbH, 4550 Kremsmeinster, Austria
LEDs Lamp OSARAM 80012 White LED lamp, 80012 White
Environment Meter PCE PCE EM882 Lux, Temp, RH%
Thermostatic cabinet Aqua Lytic (Liebherr) ET636-6
Light timer Timer T 6185.104 230V/50HZ (check specifications for your country)
Universal thermostat Conrad UT200
Humidifier Boneco
Balck tape Tesa 5 cm
Glue Uhu
lncubator lamp Phillips Softtone 5W
Timer clock Ziliss Ziliss, Switzerland
Excel Software Microsoft Excel
Origin Software 8.5 OriginLab
Backer Yeast Migros Switzerland
Iron support stand 17X28CM Fisher Scientific S47808
Acetic acid Sigma Aldrich A6283-100ML 20% acetic acid dilluted in H2O
Red light lamp Phillips PFE712E*8C
Spatula Fisher Scientific 14-373-25A
Power supply EA EA PS 2042-06B Optional
Aluminium foil Prix Coop
Heater GOON NSB200C
Microwave Oven Intertronic
Standard corn meal fly food
Destilled water

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