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The MazeEngineers Zebrafish Larvae Y Maze utilizes a distinctive backlighting system to enable precise behavioral assessments in zebrafish larvae. Similar to the Zebrafish Larvae T apparatus and Drosophila mazes, this device features a starting lane, bidirectional swimming pools, and specialized backlighting for straightforward video tracking. The user-friendly cover seals the pools and watertight chambers, ensuring durability and long-term use. The apparatus includes:
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Spontaneous alternation behavior (SAB) is effectively assessed in adult zebrafish using the Y-maze to observe their innate tendency to make alternating choices.
The Zebrafish Larval Y-Maze apparatus primarily comprises two starting arms and two goal arms, each paired with a separate swimming pool. It features a unique backlighting system to facilitate video tracking of subtle behaviors and locomotor skills in larvae.
Spontaneous alternation behavior tests combine the examination of consecutive choices (Lalonde, 2002), forced turn trials (Dember and Richman, 2012), stimulus satiation, and action decrement (Dember and Fowler, 1958; Hughes, 2004).
To conduct a spontaneous alternation behavior test, larvae are placed at the starting arm of the larval Y-maze to observe their natural path choices toward the goal arm. This process is repeated to monitor changes in consecutive choices. Learning is reinforced by generating external sensory stimuli within the apparatus. The larval Y-Maze’s flexible design allows for the opening and closing of different arms, making SAB tests versatile for various memory studies by encouraging and enforcing decision-making in zebrafish larvae.
The larval Y-Maze setup consists of four arms and two pools. The start and main arms each measure 50 mm in length, while the goal arms are each 25 mm long. These dimensions enable the Zebrafish larvae to effectively exhibit their exploration and learning behaviors.
Each corridor in the apparatus is 5 mm wide and 10 mm deep. The intersections where the corridors meet have an area of 25 mm². Each goal arm leads to a distinct pool with an area of 1950 mm².
The Y-Maze is fitted with a fluorescent backlight, and it can be heated from below. These features enhance the observation of detailed behavioral patterns in Zebrafish larvae. Furthermore, the light and heat can be used for reinforcement stimuli experiments.
The entire chamber is waterproof and includes a user-friendly cover that can seal the pools securely.
This protocol mirrors the Zebrafish larvae T-maze procedure. Prior to the experiment, the larval Y-maze is filled with E3 medium, which is then adjusted to a favorable temperature of 28°C for the larvae. The test subject is placed in the starting arm of the apparatus 10 minutes before the experiment begins, with a translucent tube blocking the intersection between the starting and main arms. An ideal sample size is 20 larvae per trial, although the apparatus can be customized to accommodate larger samples if needed.
After the 10-minute adaptation period, the tube is removed to open the corridor between the starting and main arms, allowing the larvae to explore the path towards the goal arm. The primary objective of the test is to measure the time it takes for the larvae to enter the goal arms and the number of successful entries within the first 10 minutes of the trial.
A stopwatch with two buttons is used to record the time and the number of larvae entering the goal arm. One button tracks the number of entries, and the other records the time taken by each larva to reach the goal. If a larva returns to the main arm or enters the second goal arm after an initial successful entry, only the first entry is counted.
For automated tracking, Noldus EthoVision XT can be utilized to record the larvae’s movements and activities. This advanced video tracking system efficiently measures the larvae’s movement and can accurately track and record the number of larvae entering the goal arm. This data is valuable for revalidation and reconfirmation of results in future tests.
The entire test takes 20 minutes, including the 10-minute adaptation period. Once the test is complete, the larvae and E3 medium should be removed from the Y-maze. The apparatus must be disinfected before starting another experiment.
Larval Y-Maze apparatus can be used to evaluate the spontaneous alternation behavior in Zebrafish larvae. The number of consecutive turn directions contributes to the SAB score.
The larval Y-Maze apparatus is also effective in conducting the forced turn trials on Zebrafish larvae. The arms can be conveniently closed to force the subjects to move towards the desired corridor.
SAB experiments using larval Y-Maze apparatus are essential in observing the early cognitive functions like memory and learning in Zebrafish larvae. This learning is imperative to develop the navigational skills that are essential for survival.
The Zebrafish Larval Y-Maze apparatus offers an exceptional opportunity to study the memory capabilities of Zebrafish during their early developmental stages. It is also employed to observe and analyze the search patterns Zebrafish larvae use to find food and shelter.
One significant advantage of the backlighting feature in the larval Y-maze apparatus is its ability to facilitate the tracking of detailed behaviors in Zebrafish larvae, such as Spontaneous Alternation Behavior (SAB), and to closely examine their locomotive skills.
This innovative apparatus provides valuable utility for a variety of experiments aimed at investigating memory and behavioral studies across different developmental stages of Zebrafish larvae.
Manual time recording is susceptible to errors. This issue can be resolved by employing an automatic video tracking system like Noldus EthoVision XT, which offers a precise method for tracking and recording the animals’ movements.
Best, J. D., Berghmans, S., Hunt, J. J., Clarke, S. C., Fleming, A., Goldsmith, P. and Roach, A. G. (2008). Non-associative learning in larval zebrafish. Neuropsychopharmacology 33, 1206-1215.
Stefan Yu Bögli, Melody Ying-Yu Huang. (2016). Spontaneous alternation behavior in larval zebrafish. Journal of Experimental Biology; 220: 171-173. Doi: 10.1242/jeb.149336
Dember, W. N. and Richman, C. L. (2012). Spontaneous alternation behavior: Springer Science & Business Media.
Dember, W. N. and Fowler, H. (1958). Spontaneous alternation behavior: Psychological Bulletin 55, 412.
Lalonde, R. (2002). The neurobiological basis of spontaneous alternation: Neuroscience & Biobehavioral Reviews 26, 91-104.
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