A 3D-Printed and Freely Available Device to Measure the Zebrafish Optokinetic Response Before and After Injury

Abstract

Zebrafish eyes are anatomically similar to humans and have a higher percentage of cone photoreceptors more akin to humans than most rodent models, making them a beneficial model organism for studying vision. However, zebrafish are different in that they can regenerate their optic nerve after injury, which most other animals cannot. Vision in zebrafish and many other vertebrate animals, including humans, can be accessed using the optokinetic response (OKR), which is an innate eye movement that occurs when tracking an object. Because fish cannot use an eye chart, we utilize the OKR that is present in virtually all vertebrates to determine if a zebrafish has vision. To this end, we have developed an inexpensive OKR setup that uses 3D-printed and off-the-shelf parts. This setup has been designed and used by undergraduate researchers and is also scalable to a classroom laboratory setup. We demonstrate that this setup is fully functional for assessing the OKR, and we use it to illustrate the return of the OKR following optic nerve injury in adult zebrafish.

Keywords: OKR; neuroscience; optic nerve; optokinetic response; regeneration; vision.

FIG. 1.

OKR device setup. (A) An example photograph of a current complete OKR setup. (B) Schematic indicating how to restrain the zebrafish while performing the OKR assay. OKR, optokinetic response.

FIG. 2.

Manual computational analysis of the OKR. (A) Yellow line indicating the reference midline of the fish. To measure the angle of the eye surface (marked by the red line) use the angle tool in ImageJ to click in the three arrow delineated points in the indicated order. (B) A second frame from the same video as (A) shows expected change in eye angle. This change in eye angle can be seen in the difference in angles between the red lines in A and the green lines in B.

FIG. 3.

Analysis of eye angles during the OKR induced at varying rotational speeds. In this example, a zebrafish was stimulated with nine different rotational speeds for 1.5 min each. The graph is color-coded to illustrate that the OKR is seen at different rotational speeds. rpm, revolutions per minute.

FIG. 4.

Analysis of the OKR following recovery from optic nerve injury in adult zebrafish. (A) Tracking of OKR-related eye movements of five zebrafish shown in different colors from days 5 to 20 after optic nerve injury using the following scale: 0 showing no eye movement, 1 showing eye movements that do not coordinate with the expected OKR, 2 showing limited movements that correlate with the expected OKR, and 3 demonstrating normal OKR movements. (B) Average response of the same five fish across this period of time.