Consistency of morphological endpoints used to assess developmental timing in zebrafish (Danio rerio) across a temperature gradient

Zebrafish (Danio rerio) are model organisms for testing developmental toxicity at the morphological level. In this study, influence of temperature (24.5–28.5° C) and silver nanoparticles on developmental staging, ear–eye distance, and ratio of ear–eye distance to inner ear diameter was investigated. As temperature decreased, all endpoints showed developmental delay, with differences between endpoints in amount and type of delay measured. Differences in developmental delay patterns were observed, with rate delays increasing over time when staging endpoints were utilized and rates remaining constant when using ear–eye measurements. Integrated predictive equations were created to normalize each endpoint for temperature. Influence of image rotation on ear–eye distance accuracy showed that more than 75% eye overlap during analysis is necessary to minimize error. Exposure to silver nanoparticles demonstrated a lack of consistency between developmental endpoints and highlighted the usefulness of a multi-endpoint approach when measuring changes to developmental timing.

► Increasing temperature significantly increased in the rate of zebrafish development.
► The amount and type of delay observed varied depending the endpoint being measured.
► Predictive equations were created to estimate developmental delays.
► Physical position of embryos during analysis greatly influences image data.
► Little consistency existed between endpoints used to measure developmental delay.