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.

Warming affects routine swimming activity and novel odour response in larval zebrafish.

Temperature is a primary factor affecting the physiology of ectothermic animals and global warming of water bodies may therefore impact aquatic life. Understanding the effects of near-future predicted temperature changes on the behaviour and underlying molecular mechanisms of aquatic animals is of particular importance, since behaviour mediates survival. In this study, we investigate the effects of developmental temperature on locomotory behaviour and olfactory learning in the zebrafish, Danio rerio. We exposed zebrafish from embryonic stage to either control (28 °C) or elevated temperature (30 °C) for seven days. Overall, warming reduced routine swimming activity and caused upregulation of metabolism and neuron development genes. When exposed to olfactory cues, namely catfish cue, a non-alarming but novel odour, and conspecifics alarming cue, warming differently affected the larvae response to the two cues. An increase in locomotory activity and a large transcriptional reprogramming was observed at elevated temperature in response to novel odour, with upregulation of cell signalling, neuron development and neuron functioning genes. As this response was coupled with the downregulation of genes involved in protein translation and ATP metabolism, novel odour recognition in future-predicted thermal conditions would require energetic trade-offs between expensive baseline processes and responsive functions. To evaluate their learning abilities at both temperatures, larvae were conditioned with a mixture of conspecifics alarm cue and catfish cue. Regardless of temperature, no behavioural nor gene expression changes were detected, reinforcing our findings that warming mainly affects zebrafish molecular response to novel odours. Overall, our results show that future thermal conditions will likely impact developing stages, causing trade-offs following novel olfactory detection in the environment.

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.

Zebrafish Retinal Ganglion Cells Asymmetrically Encode Spectral and Temporal Information across Visual Space.

In vertebrate vision, the tetrachromatic larval zebrafish permits non-invasive monitoring and manipulating of neural activity across the nervous system in vivo during ongoing behavior. However, despite a perhaps unparalleled understanding of links between zebrafish brain circuits and visual behaviors, comparatively little is known about what their eyes send to the brain via retinal ganglion cells (RGCs). Major gaps in knowledge include any information on spectral coding and information on potentially critical variations in RGC properties across the retinal surface corresponding with asymmetries in the statistics of natural visual space and behavioral demands. Here, we use in vivo two-photon imaging during hyperspectral visual stimulation as well as photolabeling of RGCs to provide a functional and anatomical census of RGCs in larval zebrafish. We find that RGCs' functional and structural properties differ across the eye and include a notable population of UV-responsive On-sustained RGCs that are only found in the acute zone, likely to support visual prey capture of UV-bright zooplankton. Next, approximately half of RGCs display diverse forms of color opponency, including many that are driven by a pervasive and slow blue-Off system-far in excess of what would be required to satisfy traditional models of color vision. In addition, most information on spectral contrast was intermixed with temporal information. Taken together, our results suggest that zebrafish RGCs send a diverse and highly regionalized time-color code to the brain.