Abstract
Purpose :
Modern genomics has enabled the identification of an unprecedented number of genetic variants, many of which are extremely rare, associated with blinding disorders. These advances underscore the need for tractable models to rapidly validate candidates as disease causing and to characterize their functional roles. Zebrafish eye structure is similar to humans and the rapid development and small size of zebrafish facilitates genetic screens. In addition, zebrafish display robust quantifiable behavioral responses to visual stimuli. The goal is to evaluate a spectrum of visual impairments to facilitate high-throughput in vivo screens.
Methods :
Our lab uses two specific assays in five day old zebrafish: Visual Interrogation of Zebrafish maNipulations (VIZN) and OptoMotor Response (OMR). VIZN uses interruptions in constant light to induce a startle response. Fish that are visually compromised will not respond to the interruption of light. We simultaneously capture the motion of 96 larval zebrafish in 1 second intervals as we subject them to “startle” stimuli with ViewPoint hardware. We developed software (VIZN) in the R programming language to automatically analyze the response of individual zebrafish at each stimulus, group by genotype, plot the data, and perform statistics. In OMR, fish move in the direction of apparent motion created by an animated gradient of sinusoidal waves. Larvae respond by orienting their body and swim the direction of the motion, while the visually impaired move in random directions. To quantify behavior, we developed a scoring rubric.
Results :
We have successfully used VIZN to validate candidate vision loss genes. Blind fish also do not respond to OMR. In preliminary analysis, activity of selected cataract and glaucoma genes were manipulated in zebrafish and we find that they are responsive to VIZN, but not to OMR. We are currently adapting the OMR into a high throughput test for partial vision loss. To understand the functional roles, we evaluate retinal structure and expression domains in the developing zebrafish.
Conclusions :
These assays allow for rapid screening of many candidate genes related to vision loss and hold the potential to follow disease progression and serve as a platform for drug testing.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.