Morris Water Maze Protocol

Description

The Morris water maze (MWM) consists of a round tank (pool) of water or milk with a hidden platform for the animal to locate. It is one of the best assays for spatial learning and memory in laboratory rodents, and is at the center of contemporary neuroscience research. Water or milk removes olfactory cues and provides motivation for movement. The interior is smooth to minimize allothetic cues.

The hidden platforms are included in your order.

Pricing

4 Ft (Mouse)

$ 1390

  •  Polyethylene exterior
  • Non cloggable drain placed on side or under. Choice on order.
  • 1 Platform included

5 Ft (Intermediate)

$ 1490

  • Polyethylene exterior
  • Non cloggable drain placed on side or under. Choice on order.
  • 1 Platform included

6 Ft (Rat)

$ 1590

  • Polyethylene exterior
  • Non cloggable drain placed on side or under. Choice on order.
  • 1 Platform included

Accessories

Gantry

Multiple Styles, Heights, Camera Fits

Adjustable Platform

Manual, Multiple Styles

Adjustable Platform

Manual, Multiple Styles

Floating Platform

Spatial Navigation

Radial Arm Inserts

4, 6, 8, 10, 12 arm

Star Maze

5 Arms

Y Maze Inserts

Visual Discrimination

Snowcone Inserts

Corner Snowcone cue

Plus Insert

For Spatial Navigation

Heater

0ºF to 90ºF with 1ºF differential

Multiple Beacons

Spatial Memory

Radial Arm Tread

False Escape Holes

Swim Channel: Full, Half Length

MWM Training

Release Device

Documentation

Introduction

The Morris Water Maze is a widely used behavioral task in neuroscience for studying spatial learning and memory. This test is based on the fact that an animal will try to escape a stressful situation or stimulus, which in this case is a large pool of water. The pool contains a small platform, either visible above the water level, or just below the surface of the water. This small platform allows the animals to escape the water and allows them to stand without the stress of swimming, and is designed with a mesh or grooved material that allows for easy handling. Pre-training occurs by introducing the location of the escape platform and using a platform that is visible above the water surface. On the following days, the actual test is performed, in which the platform is hidden beneath the water surface. To escape swimming in the water, the animal must remember the location of the escape platform using visual cues in the testing area, which requires use of hippocampal-dependent spatial reference memory, and this ability to remember the location of the platform can be effected by the administration of certain drugs or disease models.
The MWM was first used by Richard Morris at the University of St. Andrews in Scotland in the early 1980s. Since then, it has become one of the most widely used tools in behavioral neuroscience because of its easy of use and training, its many variations, and its ability to test various areas of brain function. Morris published a series of papers describing the maze and its evaluation of hippocampal-dependent learning over several years (Morris 1981, Morris 1982, Morris 1984, Morris 1986). The maze also gained popularity when it was used by Ian Whishaw’s group in Canada (Kolb et al. 1982, Kolb et al. 1983). Since these initial papers, the maze has been used to study various disease models, including endocrine abnormalities, strokes, Alzheimer’s disease, other neurodegenerative diseases, and their effects on learning and memory (Brandeis et al. 1989).

Apparatus and Equipment

The apparatus used for the Morris Water Maze consists of a large cylindrical pool. The diameter can range from 120 to 180 cm and the height can range from 55 to 95 cm in height depending on whether mice or rats will be tested. The pool is filled with clean, room temperature water to a depth that prevents animals from touching the bottom of the pool with their paws or tails and from climbing out the sides of the pool. The color of the pool should be in contrast to the color of the mice or rats being used.

Within the pool, there is an escape platform that the animal must locate to escape swimming in the water. This platform is generally 8 cm in diameter, and will be visible to the animal during training trials and will later be hidden below the water surface during testing trials. The color of the platform should match the color of the pool, making it difficult for the animals to locate. During the testing trials, the water of the pool can be made opaque using milk or non-toxic paint.

Visual cues may be placed in the pool above the surface of the water to help orient the animals and help them remember the location of the escape platform. Visual cues outside of the pool may also be used.

A mounted video camera is used to record the experiments from above the pool. Tracking software can be used to follow the moments of the animals within the pool. Live scoring with a stopwatch can also be performed.

Protocol

The purpose of the MWM is to assess memory and learning in animals, in a control vs. disease model/intervention group, by observing their ability to locate an escape platform within a stressful environment. This test can provide information regarding hippocampal-dependent learning, specifically spatial and long-term spatial memory. For example, the effects on memory abilities in animal models of aging or neurodegenerative diseases can be tested easily and reliably using the MWM. Typically animals are capable of learning and remembering the location of the escape platform within the water pool using visual cues. This aptitude to remember decreases, and the time the animal takes to locate the escape platform increases, in animals with impaired neurocognitive abilities. Levels of cortisol are also highly correlated with the outcomes observed in the maze.

Pre-Training for the Morris Water Maze

This test requires pre-training for the animals prior to the actual experimental testing and data collection. This training repeatedly asks the animals to locate the escape platform in order to finish the task and be removed from the pool.

To begin the training process, prepare the apparatus by placing the escape platform in the pool and filling the pool with clean, room temperature water to a depth where the platform in 1 cm above the water surface, approximately 40 cm. Set up any visual cues within the pool or testing area. Bring animals into the room, and allow an acclimation period if necessary of approximately 15 minutes.

Each animal will go through three consecutive trials in the pool. To show the animal that an escape platform exists in the pool, place the animal on the platform for twenty seconds.

Within the pool, there are four possible starting positions: north, south, west, or east. Gently lower the animal into the water at one of the positions facing the wall of the pool. Start a timer.

Allow the animal to swim and search for the platform for sixty seconds. It is common for animals to swim around the edge of the pool looking for an escape. However, the animal should know that there is a platform available and will eventually search for it. Stop the timer when the animal reaches the platform. If it has not found the platform in sixty seconds, gently guide the animal to the platform using your hand. This will teach the animal that it must find the platform to escape the pool. Record this trial as sixty seconds. Repeat this procedure for two more trials with the same animal, each time starting in a different directional position. When the three trials are complete, dry the animal carefully using towels and a heat lamp prior to returning to its home cage. Repeat the training procedure for the other animals.  

The animals are now trained and ready to perform the water maze test.

Evaluation of Spatial Learning and Memory Using the Morris Water Test

Fill the pool with clean, room temperature water so the escape platform is 2 cm below the surface. Mix in non-fat dry milk or non-toxic white paint to make the water opaque. All other conditions, such as testing room, lighting, and water temperature, should be the same as during the training session.

Each animal will undergo twelve trials, three from each starting direction. Do not use the same starting direction twice in a row. Each trial will again last sixty seconds. Begin the video recording before placing the animal in the water.

Gently lower the animal into the water at one of the positions facing the wall of the pool. Start a timer.

Allow the animal to swim and search for the platform for sixty seconds. If the animal finds the platform in less than sixty seconds, record the searching time. Allow the animal to sit on the platform for at least ten seconds. If the animal does not find the platform, guide it to the platform and allow it to sit there for at least ten seconds. Dry the animal and return it to its home cage. Repeat Trial 1 for all animals, and then move on to Trial 2 for all animals, and so on for all twelve trials.

After the completion of the testing trials, perform a probe trial for each animal in which the escape platform is removed. This test verifies the animals’ understanding of the location of platform. Place the animal in the water at one of the starting directions facing the wall of the pool. Record the number of times the animal crosses the platform location during thirty seconds. Remove the animal from the water after the thirty seconds, dry, and return to its home cage.

Modifications

Since Morris initially introduced the water maze, a variety of pool sizes have been used and proved successful for different applications, including for use with transgenic mice (D’Hooge and De Deyn 2001). Many others have described variations of this test, including alternative pre-training, training, and probing protocols (D’Hooge and De Deyn 2001).

Spooner et al. also developed the use of on-demand platforms that only rise to the surface of the water when the animal swims in proximity of it (1994). This prevents the animal from finding the platform simply by chance and results in a highly focused searching strategy.

Markowska et al. suggested performing repeated probe trials after each training trial, which allowed for the generation of extensive learning curves (1993).

In some protocols, the platform is moved to a new location on each testing day. This prevents the animal from knowing the location of the platform during the first trial each day, but once the platform is located, the animal can generally learn and remember the location in one trial (Steele and Morris 1999). The time between trials can be adjusted to study spatial memory.

Data

The data obtained from the Morris Water Maze is generally visualized by graphing the time it takes the animal to locate the escape platform, which is referred to as the latency time. This time is obtained by observing the animals in the maze live or by analyzing the recorded experiments with a stopwatch. As the animal learns the location of the platform with repeated trials, the latency time decreases. The latency time can be easily graphed and compared across the sham control and disease model or intervention groups, as shown in the example graph below.

Using graphs similar to this to compare the latency time between different disease or treatment groups allows for easy visualization of the effect on spatial memory and learning. Animals in the control groups should show significant decrease in latency time as they rapidly learn the location of the escape platform. Animals as disease models of neurodegenerative disorders, for example, should show a much slower learning curve with higher latency times, even after several trials. Generally, animal cohorts of 20-30 animals are sufficient to obtain p-values of <0.05 using ANOVA, t-tests, or Bonferroni’s post hoc tests (Harrison et al. 2008).

Strengths and Limitations

A major strength of the Morris Water Maze is its relative simplicity compared to other tests of cognitive function, learning, and memory. Despite the fact that the maze requires the animals to be trained prior to test, the training protocols are less laborious and time-consuming than other mazes. Another advantage is the ability to test both and differentiate between spatial and non-spatial memory by using visible or hidden escape platforms. Since the task can employ various modifications, it is capable of testing the brain function of many brain areas, not only the hippocampus, and this allows the test to evaluate more general cognitive function in addition to make specific learning and memory functions.

 

Despite the presence of an escape platform, this test places a significant amount of stress on the animals. Initially, when the animals are placed into the pool, they are forced into a stressful situation with no obvious escape route. The act of being immersed in water and forced to swim can be induced distress that may alter the outcomes of each repeated trial. To minimize stress experienced by the animals, it is extremely important to ensure that the water temperature is appropriate. Mazes can be purchased with temperature control to help reduce stress caused by the water being too cold or too hot. The Barnes Maze is an additional maze used to test spatial learning in animals, and does not require immersion in water or forced swimming.

Summary and Key Points

-This test can provide information regarding hippocampal-dependent learning, specifically spatial and long-term spatial memory

– The color of the platform should match the color of the pool, making it difficult for the animals to locate.

The water of the pool can be made opaque using milk or non-toxic paint.

-To minimize stress experienced by the animals, it is extremely important to ensure that the water temperature is appropriate

-Animals control groups show significant decrease in latency time as they rapidly learn the location of the escape platform. For comparison, animals as disease models of neurodegenerative disorders should show a much slower learning curve with higher latency times

References

Brandeis, R., Brandys, Y., & Yehuda, S. The use of the Morris Water Maze in the study of memory and learning. Int. J. Neurosci. 48, 29-69 (1989).

Bromley-Brits, K., Deng, Y., & Song, W. Morris Water Maze test for learning and memory deficits in Alzheimer’s disease model mice. JoVE 53, 2920 (2011).

D’Hooge, R., & De Deyn, P.P. Applications of the Morris Water Maze in the study of learning and memory. Brain. Res. Reviews 36, 60-90 (2001).

Harrison, F.E., Hosseini, A.H., & McDonald, M.P. Endogenous anxiety and stress response in water maze and Barnes maze spatial memory tasks. Behav. Brain Res. 198, 247-251 (2009).

Kolb, B., Sutherland, R.J., & Whishaw, I.Q. A comparsion of the contributions of the frontal and parietal association cortex to spatial localization in rats. Behav. Neurosci. 97, 13-27 (1983).

Kolb, B., Pittman, K., Sutherland, R.J., & Whishaw, I.Q. Dissociation of the contributions of the prefrontal cortex and dorsomedial thalamic nucleus to spatially guided behavior in the rat. Behav. Brain Res. 6, 365-378 (1982).

Markowska, A.L., Long, J.M., Johnson, C.T., & Olton, D.S. Variable-interval probe test as a tool for repeated measurements of spatial memory in the water maze. Behav. Neurosci. 107, 627-632 (1993).

Morris, R.G.M. Spatial localization does not require the presence of local cues. Learn. Motiv. 12, 239-260 (1981).

Morris, R.G.M. Development of a water-maze procedure for studying spatial learning in the rat. J. Neurosci. Methods. 11, 47-60 (1984).

Morris, R.G.M., Anderson, E., Lynch, G.S., & Baudry, M. Selective impairment of learning and blockade of long-term potentiation by an N-methyl-D-aspartate receptor antagonist. Nature 319, 774-776 (1986).

Morris, R.G.M., Garrud, P., Rawlins, J.N.P., & O’Keefe, J. Place navigation impaired in rats with hippocampal lesions. Nature 297, 681-683 (1982).

Nunez, J. Morris Water Maze experiment. JoVE 19 (2008)

Spooner, R.I.W., Thomson, A., Hall, J., Morris, R.G.M., & Salter, S.H. The Atlantis platform: a new design and further developments of Buresova’s on-demand platform for the water maze. Learning and Memory 1, 203-211 (1994).

Steele, R.J., & Morris, R.G.M. Delay-dependent impairment of a matching-to-place task with chronic and intrahippocampal infusion of the NMDA-antagonist D-AP5. Hippocampus 9, 118-136 (1999).

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Sizing Chart

Mouse

Mouse Morris Water Maze Size (CM)

  • Diameter: 120
  • Height: 81

Intermediate

Intermediate Morris Water Maze Size (CM)

  • Diameter: 155
  • Height: 89

Rat

Rat Morris Water Maze Size (CM)

  • Diameter: 180
  • Height: 76