Characterizing choice behavior in Drosophila melanogaster olfactory classical conditioning

Drosophila melanogaster is a powerful model organism to understand the neural and molecular mechanisms underlying learning and memory. However, memory is usually studied by assessing choice behavior at only one point in time in D. melanogaster, which limits our understanding of how flies respond to a learned cue. We developed a simple open-field-style assay that records the continuous behavioral dynamics of flies as they form preferences between two competing olfactory stimuli. We trained flies to associate odors with either a reward (sucrose) or a punishment (electric shock) and then placed them in a circular arena where they encountered a choice between a paired or unpaired odor. Within 30 s, flies approached the sugar-associated odor or avoided the shock-associated odor. Their collective behavior dynamics suggest the flies became more decisive over time; they traveled more on the preferred side, stayed closer to the source of the preferred cue, made more stops there, and consistently turned away from the alternative cue. We confirmed these results by showing that a learning-deficient mutant strain displayed no choice behavior. This assay provides enhanced resolution for understanding decision making as a continuous, dynamic process in D. melanogaster.