Y/T Rodent Maze

The traditional Y-Maze and T-Maze are commonly used to study spatial learning and memory in rodents.  They are also used for analyzing hippocampal place cell firing, which is used to provide a cognitive map of the subject’s environment and consequently can be used to understand spatial navigation strategies including goal-directed behavior (Grieves, Wood, & Dudchenko, 2016). The Y-T Rodent Maze, which is a combination of the Y and T-Mazes, effectively analyzes spatial learning and memory, and spatial navigation strategies. The maze utilizes a win-stay paradigm to evaluate the subject’s performance. During testing, the subject must correctly navigate past two-choice points to reach the rewarded goal box. The configuration of the maze makes four routes, which lead to the goal boxes. Therefore, the subject must correctly analyze and remember the correct route to take to find the reward. The reward can be placed in the same goal box for one block of trials and then moved to another goal box for the next block of trials to associate all four routes with the reward. 

Since Routes 2 and 3 are overlapping routes to reach the center goal box. A transparent barrier can be inserted on one of the entries of the center goal box to prevent the subject from reaching it from that route or side of the maze, which challenges their spatial navigation further. However, utilizing the maze without the transparent barrier can be used for win-stay free choice trials. Barriers can also be placed at the ends of the second choice point on either side of the maze to convert the maze to a standard Y-Maze. Similarly, the other parts of the maze can be blocked off to convert it to the traditional T-Maze.

The Y-T Rodent Maze is made of acrylic and has a start box, three choice points, three goal boxes, and connecting alleyways.  The start box leads the subject to the first choice point of the maze, which has a left and right choice arm connected to it at 45°, similar to the standard Y-Maze. Entry into either choice arm leads the subject to the second choice point, which also has a left and right choice arm connected perpendicularly to it, similar to a T-Maze.

The goal boxes are present at each end of the choice arms, making a left goal box, a right goal box, and a central goal box. The center goal box connects the two T-Mazes. Different objects can be placed in the goal boxes to allow the subject to distinguish them from one another. The goal boxes contain heavy ceramic reward dishes.

The maze has four routes, connecting the start box to the goal boxes. The leftmost route leads to the Left Goal Box (Route 1); the center-left path to Center Goal Box (Route 2); the center-right path to Center Goal Box (Route 3); and the rightmost path to Right Goal Box (Route 4).

The maze is available in a choice of colors and can be placed on a table/stool or stand to elevate it from the floor. A removable transparent barrier can be inserted at the end of the start box to confine the subject within it. Another removable transparent barrier can be placed at either entrance of the central goal box to prevent the subject from entering the goal box through that entrance.

Rodent location can be tracked using a video tracking software package such as Noldus EthoVision, ANY-Maze, or BehaviorCloud.

Clean the maze with 70% ethanol after every trial to remove odor cues from previous trials. Appropriately light the apparatus. The following is a sample protocol using a win-stay task and free-choice task to assess spatial navigation behaviors in rodents.

Y-T Maze Win-Stay Task

Place a food reward in one of the goal boxes. Place different objects in the goal boxes and attach different colored drawings, symbols, or letters to the walls of the goal boxes.

Place a barrier at the start box exit. Place the subject in the start box and confine it in it for a few seconds. Remove the barrier and allow the subject to explore the maze and enter one of the three goal boxes. Allow the subject to eat the reward for three seconds if it enters the correct box and score it as a correct choice. If the subject enters an unrewarded goal box, score it as an incorrect choice. Lift the subject from the maze and place it in the start box along with the barrier present for a minimum of three seconds. Conduct trials until the subject enters the correct goal box. Conduct eleven additional trials in which the same goal box is reinforced, which makes one trial block. Move the reward to another goal box for the next block of trials and repeat the procedure.

If the reward is placed in the center goal box, block one of the entrances of the goal box with a transparent barrier. Therefore, the subject can only reach the center goal box from one of the two paths from the start box. If the reward is placed in the left or right goal boxes, place the transparent barrier on the entry to the center goal box furthest from the rewarded box. Therefore, if the reward is placed in the left goal box, place the transparent barrier at the right entry of the center goal box and vice versa. This ensures that the final choice made is between two open goal boxes.

 Y-T Rodent Maze Win-Stay Free-Choice Task

Conduct trials in the same manner as the previous task; however, remove the transparent barrier to the middle goal box. Therefore, the subject can access the center goal box from any of the two central routes leading to the center goal box from the start box. Place the reward in one goal box for one trial block, and reinforce the reward in the center goal box for two blocks. Conduct twelve sessions.

The following parameters can be observed using the Y/T Maze

• The following parameters can be observed using the Y-T Rodent Maze:
• The number of correct trials
• The number of error trials before discovering the rewarded goal box at the beginning of each trial block
• The number of error trials after discovering the rewarded goal box during each block
• The route taken by the subject for each trial
• Trial duration

Investigation on whether place cells encode route or destinations

Grieves, Wood, and Dudchenko (2016) investigated whether hippocampal place cells encode routes or destinations during spatial navigation in rodents using the Y-T Maze. Twelve male Lister Hooded rats were used in the study.

The rats first performed a Win-Stay Task in which they were required to explore the maze and find the rewarded goal box. A transparent barrier was placed at one entry of the center goal box to block one part of the maze so that the subject could only get to the reward using one route. The goal boxes contained different objects: a small white opaque bottle with a cork stopper, a small gray elephant stature, and a small black and red box with a slanted lid. The boxes also had a different reflective colored Latin alphabet character affixed to the walls. The objects and alphabets were used as cues for the subject to distinguish the boxes across trials. Trials were conducted until the subject found the correct goal box and then eleven additional trials were conducted to reinforce the rewarded location. The reward remained in the same goal box for each block of trials and then was moved to another goal box for the next trial block.

The number of error trials before and after discovering the correct goal box, the route chosen by the subject to reach the goal box, and the trial duration were recorded to analyze its spatial navigation performance. The results indicated that the subjects reduced the time it took to find the reward and the number of errors after finding the correct goal box in each block of trials. Therefore, the subjects learned to apply the win-stay or lose-shift strategy to find the reward. However, it was observed that when routes 1 or 4 to the Left and Right Goal Boxes were rewarded, the animals made fewer errors during training than when Routes 2 or 3 to the Center Goal Box were rewarded.

The subjects also performed a free-choice task without the presence of a transparent barrier. Therefore, the subject could access the center goal box using either route of the maze. Unfortunately, this variation of the task was ineffective for determining differences in place cell activity between routes and goals since all of the rats quickly formed a preference for only one route to the Center Goal Box.

Place cell activity analysis was performed on eight of the subjects that underwent surgery for the insertion of electrodes to record place cell firing. These subjects performed the task pre-surgery as well as after recovery from surgery. To analyze whether place cells encode route versus destination, the reward was placed in the center goal box since two routes led to the same destination. It was observed that only two cells (4.2%) of the cells that fired on these two routes displayed goal-dependent firing (firing similarly on both routes), as opposed to 95.8% of the cells that displayed route-dependent firing (firing on only one path). Additionally, route-dependent place cells and place cells, in general, overrepresented the start location and the less discriminable paths. These findings suggest that the animal’s route, not its goals, is represented in place cell firing on overlapping routes and that this firing may facilitate spatial discrimination.

Strengths

The Y-T Maze is used to examine spatial learning and memory in rodents. Unlike the Y- and T-Maze, which only has a single choice point that leads to a goal, the Y-T has three choice points in which the subject has to correctly navigate past two-choice points to reach the rewarded goal box. Different objects can be placed in the goal boxes and different colored symbols or drawings can be affixed to their walls to be used as visual cues for the subjects to distinguish them. Barriers can easily be placed in the start box or goal box to confine the subject. Moreover, barriers can also easily convert the maze into the standard Y- and T-Maze by using them to block one part of the maze from the other. Therefore, experimenters can also utilize the Y-T maze for two-choice experiments.  

 Limitations

The maze should be blocked off from the experimental room using a curtain or black-out sheets to prevent the subject from utilizing extra maze cues for task performance. Preference for one route of the maze over the others can affect results.  Odor cues emanating from the rewarded goal box can influence behavioral performance; therefore, the other food bowls in the unrewarded goal boxes should be sprinkled with food dust.  

The Y-T Rodent Maze is used to study spatial learning and memory in rodents.

• It contains a start box, three goal boxes, and connecting alleyways. The maze is essentially a combination of the standard Y-Maze and two T-Mazes.
• The configuration of the maze makes four routes, which lead to the goal boxes. The left-most route leads to the left goal box (Route 1); the center-left path to the center goal box (Route 2); the center-right path to the center goal box (Route 3); and the right-most path to the right goal box (Route 4).
• The subject must correctly analyze and remember the correct route to
• Different objects can be placed in the goal boxes and different colored drawings, symbols, or letters can be affixed to the walls of the goal boxes to serve as cues.
• A transparent barrier can be inserted on one of the entries of the center goal box to prevent the subject from reaching it from that route or side of the maze.
• Barriers can also easily convert the maze into the standard Y- and T-Maze by using them to block one part of the maze from the other.

Grieves, R. M., Wood, E. R., & Dudchenko, P. A. (2016). Place cells on a maze encode routes rather than destinations. eLife, 5, e15986. https://doi.org/10.7554/eLife.15986