The way an animal responds to novel stimuli in the environment depends on its internal state. Animals that have recently had a stressful experience, for example, are more likely to be wary of novel stimuli. This assay measures the response of zebrafish larvae to a sudden transition to darkness.
Larval zebrafish (Danio rerio) are highly sensitive to light (Burgess and Granato, 2007). They display regular rates of motion while swimming under even illumination, and can react to a sudden change in light with stereotyped locomotion alterations. This makes them ideal for studies of internal states (De Marco et al., 2016) and behavioural screens. The Zantiks MWP unit allows for measuring the activity of multiple larvae and control light of varying power and wavelength.
Experimental set up
Larval zebrafish are placed individually in each well of a 6, 12, 24, 48 or 96 well plate.
Experimental procedure
The multi-well plate with larvae is placed onto the light stimulation plate which is then placed into the chamber of the Zantiks MWP unit. The script can be written to control single or multiple light transitions. The length and power and wavelength of the stimuli can be controlled and automated in script as well as the system's temperature. The locomotor activity of each larva (measured as distance travelled) is measured and written to a data file which is exported as a .csv file of processed data at the end of the experiment.
In this example protocol, fish are habituated to the well plate for 10 minutes in the light before the experiment begins. Following habituation, 10 minutes of data in the light is recorded. After 10mins, fish are subjected to a sudden transition to the dark and distance travelled is recorded for a further 10 minutes. In this example, the fish are exposed to a further 3 transitions to light or dark at 10minute intervals. The number of transitions and length of time between transitions can be easily edited in the script.
Results
Zebrafish larvae in a 24-well plate during a light/dark transition test. (1 frame per sec.)
Protocols scripts downloads
There is one example script for the light/dark transitions test for zebrafish larvae in the MWP system. This script can be adapted to change the number and duration between light & dark transitions. The script can also be adapted for use with adult fish in the LT and AD systems by changing the asset, the data headings, and the tracking settings.
Assets
You will need to upload the correct well plate asset into the Asset directory on your Zantiks Control Console and ensure the correct asset name is in the LOAD(ARENAS,"name_of_asset") command in the script.
See the Calibrating your Zantiks unit page and Asset building in the MWP unit page for details on how to create assets customised to your system.
