The overall goal of the following experiment is to test hippocampal dependent learning and memory. On day one, the mouse is trained to find a raised flagged platform that is randomly placed in a pool of clear water to allow the mouse to learn that the task has an escapable endpoint. As a second step, the mouse is trained on subsequent days by being released into a pool of opaque water from different positions with the platform submerged and fixed in one location.
This allows the mouse to learn the location of the platform by using the surrounding cues. Next, the platform is removed on the final day and the swim pattern of the mouse is tracked for one minute in order to access task acquisition. And hence, learning and memory results are obtained.
That show impairments in spatial learning and memory based on the escape latencies and path length seen in the three different testing conditions. The main advantage of this technique over existing masses is that it's relatively simple to differentiate between special hidden platform and non-special visible platform conditions. Generally, people new to this method may struggle cause it's sensitive to experimental handling.
You have to ensure the mice aren't stress during the procedures, this could affect the results. Demonstrating this procedure will be Ms.Kelly Bromley breaths and Miss Sweden graduate students from my laboratories First secure a circular pool. If the experimental mice are black, use a white pool and if mice are white, use a black pool.Now.
Arrange the room with dividers such that the animals being tested will not see the experimenter during testing place. High contrast spatial cues about the room and on the interior of the pool at a location that will be above the water surface. Fill the pool with water and equilibrate to room temperature.
Then place a 10 centimeter diameter platform in the pool, one centimeter above the water surface, white for a white pool, or clear plexiglass for a black pool. Calibrate the pool in the computer software so the camera can create physical distance information. From pixel based information.
Divide the pool into four quadrants. Specify the platform zone as a variable zone, which can change with each trial. Then create five platform sub zones, one in each quadrant, and one in the center of the pool.
Save the calibration and use it for the remaining test days. Next, set the maximum trial time as 60 seconds. If the mouse finds the platform before this time, program the software to stop the trial when the platform is found.
Also specify the program to begin tracking automatically when the experimenter exits the testing area. Utilize any reflection minimization options the software package provides. Finally, track path, length, escape, latency, and time spent in each quadrant.
On the day of testing, transfer the mice from their housing facility to the behavior room In an area where they cannot see the pool or spatial cues. Let them adjust to the new environment for at least 30 minutes. Before testing, load the pool calibration into the tracking software.
Create five trials with an appropriate inter trial interval program, the platform location, and starting direction to differ with each trial. Place a flag on the platform to increase its visibility. To begin testing, lift the mouse from the home cage by the base of the tail, supporting the mouse during transfer to the testing area.
Then gently place the mouse into the water facing the edge of the pool. Quickly leave the testing area. If the mouse finds the platform before the 62nd cutoff, allow the mouse to stay on the platform for five seconds, then return it to its home cage.
If the mouse does not find the platform, place the mouse on the platform and allow it to stay there for 20 seconds before returning it to its home cage. Repeat for all mice in the trial. Begin each subsequent trial with a different platform location and starting direction according to the program software.
When testing is complete, return the mice to their housing facility in preparation for the following day. Remove the flag from the platform and add additional water to the pool to submerge the platform to one centimeter below the surface. Load the pool calibration into the tracking software and create five trials with an appropriate inter trial interval.
Then program the platform location to remain in the same position throughout all trials and days, but have the starting direction differ with each trial each day. For black mice, add non-toxic white powdered temperate paint to the pool and mix thoroughly. Use enough paint such that the submerged platform is not visible from the surface of the water.
Proceed to test individual mice in the water as described earlier for the final probe trial. Load the pool calibration into the tracking software. Create one trial with no platform zone and one starting direction.
Preferably the starting direction farthest from the platform quadrant used on days two to five. Also set the trail length to 60 seconds. Now remove the platform from the pool and test individual mice in the water as described earlier for each day and each mouse average the five trials to give a single path length and escape latency for each test subject.
Further, analyze and interpret data as detailed in the accompanying manuscript using the Morris water maze. Seven month a PP 23 transgenic mice carrying the human Swedish mutant A PP gene were tested after one month of daily valproic acid or vehicle solution injections during the first day of visible platform tests. The VPA treated and control A PP 23 mice exhibited a similar latency to escape onto the visible platform.
The VPA treated and control A PP 23 mice had similar swimming distances before escaping onto the visible platform. In the visible platform test in hidden platform tests on the third and fourth day, VPA treated a PP 23 mice showed a shorter latency to escape onto the hidden platform. On the third and fourth day, the VPA treated a PP 23 mice had a shorter swimming length before escaping onto the hidden platform.
In the probe trial on the sixth day, compared to control mice, the VPA treated a PP 23 mice traveled significantly more times into the third quadrant where the hidden platform was previously placed. While attempting this procedure is important to remember to monitor the environmental factors that might cause stress, such as noise, temperature, And light. Following this procedure, other methods such as queued and contextual fear conditioning can be performed to answer additional questions such as whether the learning deficits are specific to the hippocampus or whether it's part of a broader neurocognitive problem, which may include the amygdala.
