Afterimages drive a shared visual motion-reversal illusion in Drosophila

Illusions expose core computations in perception. In one visual apparent-motion illusion, perceptual direction is reversed when phase-shifted gratings are interleaved with uniform frames. Here, we demonstrate that Drosophila exhibits the same direction reversal reported in mammals. Combining behavior, targeted silencing, two-photon imaging, and modeling, we localize the origin of this illusion to elementary motion pathways. Silencing direction-selective T4/T5 neurons abolishes the reversal, and recordings reveal that downstream wide-field neurons invert their directional preference as interleave duration increases. Replacing periodic gratings with random binary patterns preserves the reversal, implicating afterimages rather than spatial periodicity. Imaging neurons upstream of T4/T5 shows signatures of an afterimage, whose emergence depends on interleave luminance. Critically, dark interleaves suppress afterimages and eliminate both the neural and behavioral reversal, whereas light interleaves preserve or enhance it. Thus, afterimages are central to this shared illusion and explain a deficiency of canonical motion-energy accounts. These results link a classic apparent-motion phenomenon to identified circuit elements and reveal a simple stimulus manipulation that switches an illusion on and off.