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Locomotion in Larval Zebrafish: Influence of Time of Day, Lighting and Ethanol
Citation:
MACPHAIL, R. C., J. Brooks, D. L. HUNTER, B. K. PADNOS, T. D. Irons, AND S. J. PADILLA. Locomotion in Larval Zebrafish: Influence of Time of Day, Lighting and Ethanol. NEUROTOXICOLOGY. Intox Press, Inc, Little Rock, AR, 30(1):52-58, (2009).
Impact/Purpose:
Remarkably few chemicals in commerce have ever been tested for their potential developmental neurotoxicity, owing in part to the resources, time and expense required for testing. As a result, alternative test methods are being promoted by regulatory, scientific, and stakeholder organizations that can be used to screen and prioritize chemicals for later in-depth evaluation of their neurotoxic potential. Alternative animal models include zebrafish larvae, which have been used increasingly in developmental neurobiology, toxicology and pharmacology. Understanding the behavior of the larvae is crucial for successful validation of an in vivo screening battery. This manuscript describes a series of studies that parametrically determined the effects of time of day and lighting conditions on locomotion in larval zebrafish. Activity was recorded simultaneously in individual larvae housed in a 96-well microtiter plate, allowing testing of large numbers of larvae and chemicals. Results showed that larval locomotion could be recorded efficiently in microtiter plates, and that the activity was highly sensitive to both time of day and lighting conditions. Ethanol was also administered as a positive neurotoxic control treatment, and produced a variety of behavioral effects that depended on lighting conditions. These results identified optimal testing parameters for larval locomotion that can be used in a broad-based chemical screening effort.
Description:
The increasing use of zebrafish (Danio rerio) in developmental research highlights the need for a detailed understanding of their behavior. Behavior represents the unique interface between intrinsic and extrinsic forces that determine an organism’s health and survival. We studied the locomotion of individual zebrafish larva (6 days post fertilization) in 96-well microtiter plates. Movement was recorded using a video-tracking system. Time-of-day results indicated locomotion in darkness (infrared) decreased gradually from early morning to a stable level between 13:00 and 15:30 hr. All further experiments were conducted during this time window. Locomotion in darkness increased initially to a maximum at 4 min, then decreased steadily to a low level by 20 min. Locomotion during light was initially low and then gradually increased to a stable level after 20 min. When 10-min periods of light and dark were alternated, activity was low in light and high in dark; curiously, activity during alternating dark periods was markedly higher than during either continuous dark or light. Further experiments explored the variables influencing this alternating pattern of activity. Varying the duration of the initial dark period (10-20 min) did not affect subsequent activity in either light or dark. The activity increase on return to dark was, however, greater following 15-min than 5-min of light. Acute ethanol increased activity at 1% and 2% and severely decreased activity at 4%. One-percent ethanol retarded the transition in activity from dark to light, and the habituation of activity in dark, while 2% ethanol increased activity regardless of lighting condition. Collectively, these results show that locomotion in larval zebrafish can be reliably measured in a 96-well microtiter plate format, and is sensitive to time-of-day, lighting conditions, and ethanol.
