Odor–Taste Learning Assays in Drosophila Larvae

The Drosophila larva is an emerging model for studies in behavioral neurogenetics because of its simplicity in terms of cell number. Despite this simplicity, basic features of neuronal organization and key behavior faculties are shared with adult flies and with mammals. Here, we describe a pavlovian-type learning assay in fruit fly larvae. A group of larvae is sequentially exposed to specific odors in the presence or the absence of sugar, and then tested to determine whether they prefer the odor previously experienced with the reward. The protocol uses a two-group, reciprocal training design: One group of Drosophila larvae is exposed to n-amyl acetate (AM) with a sugar reward (+), then subsequently exposed to 1-octanol (OCT) with no reward (denoted AM+/OCT). The other group receives the reciprocal training (AM/OCT+). The two groups of larvae are then tested for their choices between AM and OCT. Relatively higher preferences for AM after AM+/OCT training than after AM/OCT+ training reflect associative learning and are quantified by the learning index (LI). This method offers a robust, simple, cheap, and reasonably quick test for learning ability (an aversive version is available as well, using either high-concentration salt or quinine as punishment). With the concerted efforts of the Drosophila research community, we anticipate it will allow us to unravel the full circuitry underlying odor-taste learning on a single-cell level.