Visual learning in tethered bees modifies flight orientation and is impaired by epinastine

Visual-orientation learning of a tethered flying bee was investigated using a flight simulator and a novel protocol in which orientation preference toward trained visual targets was assessed in tests performed before and after appetitive conditioning. Either a blue or a green rectangle (conditioned stimulus, CS) was associated with 30% sucrose solution (unconditioned stimulus, US), whereas the other rectangle was not paired with US. Bees were tested in a closed-looped flight simulator 5 min after ten pairings of the US and CS. Conditioned bees were preferentially oriented to the CS after such training. This increase in preference for CS was maintained for 24 h, indicating the presence of long-term memory. Because the total orienting time was not altered by conditioning, conditioning did not enhance orientation activity itself but increased the relative time for orientation to CS. When 0.4 or 4 mM epinastine (an antagonist of octopamine receptors) was injected into the bee's head 30 min prior to the experiment, both short- and long-term memory formation were significantly impaired, suggesting that octopamine, which is crucial for appetitive olfactory learning in insects, is also involved in visual orientation learning.