The Y maze is similar to the T maze, except with three arms at 120 degrees to each other. The rodent or mouse starts at the end of one arm, then chooses between the other two. Spontaneous alternation is measured to demonstrate learning.
The Y-maze is often preferred to the T-maze because gradual turns decrease learning time as compared to the sharp turns of the T-maze. It is also a smaller maze* to allow less degrees of freedom of movement, focusing the animal on the task at hand. The Y Maze can also be baited with food for rewarded alternation. Food wells are standard 1cm deep.
Rat (8cm diameter): $400
The Y-Maze is a widely used behavioral task in neuroscience for studying spatial learning and memory. This test is based on the fact that rodents are motivated to explore their environment and locate food quickly and efficiently. This maze gives the animal only two options: the left arm or the right arm, each containing a food reward. Once a food reward is retrieved from one arm, the animals’ natural tendency is to alternate their choice and obtain the food reward from the opposite arm. This ability to remember spatial locations has been adapted into a simple behavioral task used to test cognitive function. It requires use of hippocampal-dependent spatial reference memory, and this ability to remember the location of the visited arm can be effected by the administration of certain drugs or disease models.
The Y-Maze is very similar to the T-Maze and can be used to obtain very similar data. The Y-Maze is often preferred over the T-Maze because it has more gradual turns (120 degrees in the Y-maze compared to 90 degrees in the T-Maze), which can decrease learning times in animals. It is also sometimes adapted as a smaller maze with shorter arms to keep the animal focused on the present task.
Alternation behavior was first studied in animals by Dember and Fowler in the late 1950s. They published a series of papers describing spontaneous vs. free alternation behaviors, as well as the influence of rewards and reinforcement (Dember and Fowler 1959, Fowler et al. 1959a, Fowler et al. 1959b). Since that time, the Y-Maze and T-Maze have developed as relatively simple and widely used tasks because they measure spatial memory and have many variations. Due to their ability to readily test the function of hippocampal learning, the mazes has been used extensively to study drugs and toxins that effect spatial memory and age-related cognitive decline (Sharma et al. 2010). It has also been shown that pre-exposure to chronic stress impairs spatial memory but does not affect novelty-seeking behavior (Conrad et al. 2003, Wright & Conrad 2005). There are also differences in how male and female animals respond to stress in the maze, as females tend to recover more quickly than males (Conrad et al. 2003).
The apparatus used for the Y-Maze consists of a capital Y-shape maze, with three arms each 120 degrees to each other. Each arm ranges from 30 to 50 cm in length to accommodate mice, rats, and small primates. The width of the pathways is approximately 10 cm. Each of the two goal arms may contain a well with a food reward; alternatively, the goal arms may contain an exit tube that allows the animal to escape the maze (Deacon 2013). Generally, there are guillotine doors at the entrance of each arm that can be used to confine the animal to a specific arm or prevent the animal from entering one of the goal arms. The Y-Maze is typically used as an enclosed maze with approx 30 cm high walls. The entire apparatus can be raised as well, sometimes to about 50 cm above the floor. The maze is usually painted a dark color to prevent the animals from feeling extra anxiety while performing the task. Intra-maze cues can also be used to help animals distinguish and remember goal arms.
Fully automated mazes are available which can detect the location of the animal within the maze, open and close doors within the maze, and detect the presence of the food reward in the arm chambers.
The maze should be lit from above to prevent shadows within the maze and lighting should be sufficient to allow the animals to see the food rewards.
A mounted video camera is used to record the experiments from above the maze. Tracking software can be used to follow the moments of the animals within the maze. Live scoring can also be performed.
The purpose of the Y-Maze is to assess spatial memory and learning in animals, in a control vs. disease model/intervention group, by observing their ability to remember which arm they have previously entered. This test can provide information regarding hippocampal-dependent learning, specifically spatial memory. Typically, animals can remember which arm they entered previously and will alternate to the opposite arm in the next trial. This aptitude to remember decreases, and the number of incorrect choices increases, in aged or stressed animals.
There are several versions of protocols to be used with the Y-Maze. The two most commonly used have been outlined below. The rewarded alternation version allows the experimenter to decide which arm is the “correct choice” for the animal. The spontaneous alternation version uses the animals’ natural drive to explore and allows the animal to make the choice of which arm to explore first and second. This version has been more successful in testing animals with hippocampal lesions, as these animals sometimes develop a side preference (Deacon and Rawlins 2006).
Pre-Training for the Y-Maze
To encourage food-scavenging behaviors, animals will need to be food deprived the night prior to testing. Animals should maintain 90-95% of their free-feeding body weight throughout food rationing and the testing period.
To begin the training and testing process, prepare the apparatus by ensuring it is clean and free of odors. Place a food reward in the chambers at the end of each arm. Bring animals into the room, and allow an acclimation period if necessary.
The animals need to be familiarized with the maze prior to testing. This can be done by placing all the animals of one home cage in the maze for approximately three minutes. During this time, the food wells will contain food rewards to accustom the animals to consuming the rewards. Fruity Pebbles (Post Foods) work especially well as food rewards for rodents. Repeat this procedure four times with at least ten minutes in between exposures.
Next, allow individual animals to run from the start position to the end of a goal arm while the opposite goal arm is blocked by a closed door. Repeat this for an equal number of times for both arms.
Evaluation of Spatial Memory using Rewarded Alternation in the Y-Maze
When ready to begin the test run, place rewards at the ends of both arms. Close the door to one arm; this will be the “correct choice” arm. Place the animal gently in the start area. Allow the animal to run to the open arm and consume the reward. Open the door to the “correct choice” arm and return the animal to the start area. Allow the animal to choose which arm to explore. If it chooses correctly, allow it to consume the reward. If it chooses incorrectly, allow it to see that the food well is empty before removing it from the maze. Each trial should take no more than two minutes. Each of approximately ten animals will complete ten trials on each day for up to twelve testing days.
Evaluation of Spatial Learning using Spontaneous Alternation in the Y-Maze
For this version of the task, the animals are not familiarized with the maze prior to testing, as the novelty of the maze promotes exploration and alternation. Prepare the maze and place food rewards in both goal arms. Open all doors. Place the animal in the start area and allow it to choose a goal arm. Close the door to chosen arm and confine the animal there for thirty seconds. Remove the animal and open both doors. Replace the animal at the start area and allow it to again choose a goal arm. If it chooses correctly, allow it to consume the reward. If it chooses incorrectly, allow it to see that the food well is empty before removing it from the maze. Each trial should take no more than two minutes. Each of approximately ten animals will complete ten trials on each day for up to twelve testing days.
Alternatively, other spontaneous alternation Y maze protocools allow for natural alternation between the previously chosen arm and the new arm. Spontaneous alternation looks at % of chosen new arms in a timed trial without any intervention during the trial.
Similar to the T-Maze, there are several variations of the Y-Maze that have developed as cognitive neuroscience research has advanced. These modifications allow for the study of particular aspects of spatial learning and alternation behavior.
The goal arms of the maze may contain some cues that the animals must recognize and to which they must respond. For example, a large enamel-painted rock could be placed in each of the goal arms. These rocks could be distinguished by texture or paint color/design. These cues can reduce the difficulty of the task and help the animals remember which of the goal arms they have visited. Animals can be trained to always choose the arm with a specific rock or cue. (Wright & Conrad 2005)
An aquatic version of the maze has also been adapted. A Y-shaped tank is filled with water, and the animals are forced to make an arm choice while swimming. They can be trained to select a specific arm by learning which contains an escape platform, ladder, or tube. (Deacon 2013)
The data obtained from the Y-Maze is generally very straightforward and consists of the number of correct (the animal enters the opposite arm on the second run) vs. incorrect (the animal enters the same arm previously entered on the previous run) arm entries in each trial. The time it takes the animal to retrieve the food reward can also be recorded. As the animal learns that entering a new arm results in a food reward, the number of incorrect arm entries should decrease. The percentages of correct arm choices can be graphed and compared across a sham control group and a disease model/intervention group, as shown in the following graph:
Using graphs similar to this to compare arm entries between different disease or treatment groups allows for easy visualization of the effect on spatial memory and learning. Animals in the controls groups should show significant improvements in their correct arm choices. Animals as disease models of neurodegenerative disorders, for example, should show a much slower learning curve with more incorrect choices, even after several trials. Generally, animal cohorts of 10-30 animals are sufficient to obtain p-values of <0.05 using ANOVA (Wright & Conrad 2005, Conrad et al. 2003).
A major strength of the Y-Maze is its relative simplicity compared to other tasks that test spatial learning and memory. It requires minimal time and training to perform and its simplicity allows for easily reproducible results. The Y-Maze also places minimal stress on the animals. Other mazes that also test spatial learning and memory, such as the Morris Water Maze, subject the animal to significantly more stress as it must be submerged in water, swim in order to survive, and search for an escape platform (Hodges 1996). The absence of significant stressors and familiarization with the maze prior to testing allow for better observations of working memory in the animals as they perform in the maze.
A limitation of the Y-Maze is that it only has a single point choice with only two alternatives. This increases the possibility of success because by default the probability of the animal choosing the correct arm is naturally 50%. It also allows for the possibility that the animal may use a strategy other than spatial learning to solve the maze.
As with all mazes that measure aspects of learning and memory, it is important to remember that many different processes play into behavior in the maze. In many cases, the Y-Maze is used in conjunction with other mazes to study disease models or transgenic animals and gain a fuller understanding of spatial learning and memory.
- The Y-Maze is an extensively used maze to test spatial learning and memory
- This task asks animals to retrieve food rewards at the end of both arms by alternating their arm choice
- Different groups have adapted this maze in order to collect data regarding different aspects of spatial learning
- Animals in control groups show rapid learning as they remember the location of the arm from which they have retrieved the food reward, while in comparison, animals as disease models show a much slower learning curve
- The maze has been used to study different aspects of cognitive function, such as hippocampal lesions, age-related cognitive decline, and chronic stress
Conrad, C.D., Grote, K.A., Hobbs, R.J., Ferayorni, A. Sex differences in spatial and non-spatial Y-maze performance after chronic stress. Neruobio. Learning and Memory 79, 32-40 (2003).
Deacon, R.M.J. Shallow water (paddling) variants of water maze tests in mice. J. Vis. Exp. 76, e2608 (2013).
Dember, W.N., Fowler, H. Spontaneous alternation after free and forced trials. Can. J. Psychol. 13, 151-154 (1959).
Fowler, H., Blond, J., Dember, W.N. Alternation behavior and learning: the influence of reinforcement magnitude, number, and contingency. J. Comp. Physiol. Psychol. 52, 609-614 (1959).
Fowler, H. Fowler, D.E., Dember, W.N. The influence of reward of alternation behavior. J. Comp. Physiol. Psychol. 52, 220-224 (1959).
Hodges, H. Maze procedures: the radial-arm and water maze compared. Cog. Brain Research 3, 167-181 (1996).
Sharma, S., Rakoczy, S., Brown-Borg, H. Assessment of spatial memory in mice. Life Sciences 87, 521-536 (2010).
Mouse Y Maze (CM)
Mouse Y Maze (CM)
- Single Arm Width: 5
- Single Arm Length: 35
- Single Arm Height: 20
Rat Y Maze (CM)
Rat Y Maze (CM)
- Single Arm Width: 10
- Single Arm Length: 50
- Single Arm Height: 30