Protocol for using UV stimuli to evoke prey capture strikes in head-fixed zebrafish larvae.

Hunting in larval zebrafish begins with eye convergence and orienting turns, proceeds to approach swims, and ends with the strike, where larvae consume the prey. Here, we describe a protocol to present UV stimuli to zebrafish, which greatly increases the occurrence of hunting initiation and strikes. We also describe how we record and analyze strike behavior in head-fixed larvae. Our goals are to increase the robustness of prey capture and to allow other labs to implement the strike behavioral assay. For complete details on the use and execution of this protocol, please refer to Khan et al. (2023).1.

Zebrafish larvae use stimulus intensity and contrast to estimate distance to prey.

The ability to determine the distance to objects is an important feature of most visual systems, but little is known about the neuronal mechanisms for distance estimation. Larval zebrafish execute different visual behaviors depending on distance; at medium distances, they converge their eyes and approach, but when the prey is close enough, they execute a strike and suck the prey into their mouths. To study distance estimation, we developed a head-fixed strike assay. We found that we could evoke strike behavior in head-fixed larvae and quantify head movements to classify the behavior as a strike. Strikes were dependent on distance to prey, allowing us to use them to study distance estimation. Light intensity is rapidly attenuated as it travels through water, so we hypothesized that larvae could use intensity as a distance cue. We found that increasing stimulus intensity could cause larvae to strike at prey that would normally be out of range, and decreasing the intensity could lower the strike rate even for very proximal stimuli. In addition, stimulus contrast is a key parameter, and this could allow larvae to estimate distance over the range of natural illumination. Finally, we presented prey in the binocular vs. monocular visual field and found that monocular prey did evoke strikes, although the binocular input produced more. These results suggest that strike behavior is optimally evoked by bright UV dots in the binocular zone with minimal UV background light and provide a foundation to study the neuronal mechanisms of distance estimation.