Displacement experiments provide evidence for path integration in Drosophila.

Like many other animals, insects are capable of returning to previously visited locations using path integration, a memory of travelled direction and distance. Recent studies suggest that also Drosophila can use path integration to return to a food reward. However, the existing experimental evidence for path integration in Drosophila has a potential confound: pheromones deposited at the site of reward might enable flies to find previously rewarding locations even without memory. Here we show that pheromones indeed can cause naïve flies to accumulate where previous flies had been rewarded in a navigation task. Therefore, we designed an experiment to determine if flies can use path integration memory despite potential pheromonal cues by displacing the flies shortly after an optogenetic reward. We found that rewarded flies returned to the location predicted by a memory-based model. Several analyses are consistent with path integration as the mechanism by which flies returned to the reward. We conclude that while pheromones may often be important in fly navigation and must be carefully controlled in future experiments, Drosophila may indeed be capable of performing path integration.