L: Learning and Memory: Recognition

Objectives:

To examine rat learning using a test of object recognition.

Introduction:

Object recognition is a model of working memory. It is a useful model for understanding the neural basis of working memory (Mumby, 2001), as it is sensitive to pharmacological, genetic and surgical interventions. Additionally, object memory is useful in understanding memory deficits associated with various dementias such as Alzheimer’s disease. This laboratory exercise, describes two object recognition tasks, the Delayed non-matching to sample task and Novel object preference task. If there is a lot of time for this lab exercise (a couple of weeks) and the proper maze available, the Delayed non-matching to sample task can be used. The novel object preference task is more simple can run over a shorter time frame as training and hunger-motivation are not required.

Procedure:

Delayed nonmatching-to-sample Preparation: You will need to assemble a large number of objects. These objects should be large enough to cover the food cups, but light enough to be displaced by the rat. The objects should not have any sharp edges that could injure the rat. Also, objects with prominent odors should be avoided. About 50 objects will be needed if you run 25 trials. Rats will need to be trained on the apparatus ahead of time. This can be facilitated if they are a little hungry, so they may need to be maintained at about 95-100% of their free-feeding weight. This procedure required daily weighing and feeding of rats; negligence can be fatal! On the first day weigh the rats. Weigh out an excess of food (~50g) for the rats. At the same time the next day, weigh the food and the rat. The average of the rat weights between the two days will become its free-feeding weight. Whatever the rat ate will become the daily ration of food, to be given after the training/testing is complete. Adjust the amount of food up or down a gram each day to keep the rat in the 95-100% of free-feeding weight range. The following is based on the protocol of Mumby et al. (1990), and assumes you have access to a maze such as those presented by Mumby (2005).

First, habituate the rat to the choice maze by placing reward pellets in the food cups (VWR, catalog #s CA89067-518; 89067-534; or 89067-518).
Next, use an object discrimination task where they always are presented with the same 2 objects, and need to displace the same one to get a reward hidden beneath it. Do this over 2 days.
After they readily perform the object discrimination task, they must be trained to run back and forth so that you can get them to leave the reward area. Do this by alternating the end of the maze where reinforcement is received. Do this over 2 days.
Finally, if your maze has doors, train the rat in how they operate. Place the rat in the maze, and allow it to move to one end. When it does, close the distal door, and put a pellet in the cup on the other side of the door. When the rat approaches the door, raise it, and as it eats the reward, close the other door and bait a cup behind that one. Do this over 2 days.
Test:
To train the rat on the non-matching-to-sample task, use the following protocol based on that outlined by Mumby et al., (1990):
Place the rat in the central portion of the maze with both doors closed.
Place a food pellet in a single cup and cover this food cup with the sample object.
Place the novel object over an empty cup at the OPPOSITE end of the maze.
1 minute later, raise the door between the rat and the sample object food cup.
Once the rat moves the sample object, move this object to the opposite end of the maze and place it over the other empty food cup while the rat enjoys its treat.
Raise the distal door, and allow the rat to move one of the objects. Only reward the rat if it moves the novel object. On the first trial ONLY, allow the rat to correct its choice and be reward if it chooses the sample over the novel object initially.
When the rat eats its reward, remove the objects and close the distal door.
When the rat finishes eating returns to the central area, close the door behind it.
Drop a pellet into the central area if the rat does not return there after finishing its reward
Start the next trial as described above with 2 new objects, one designate sample and one designated novel.
As the rat acquired the task (21/25 correct on 2 consecutive days), increase the delay between when you remove the sample object and when you raise the door to the choice end to 15s, then 30s, then 60s, then 120s. These may need to be run on separate days.
For these longer retention intervals, count how many of the 25 trials the rat gets correct.
Provide the data to your TA who will collate the data and provide summary data to you.

Novel-Object Preference Preparation: You will need to assemble a large number of objects. The objects should not have any sharp edges that could injure the rat. Also, objects with prominent odors should be avoided. About 20 objects will be needed if you run 10 trials, and you will need at least 3 copies of each. Screen objects ahead of time for baseline amount of exploration evoked. Discard objects that evoke too little or too much baseline exploration (you want all the objects to be mildly interesting, but not too exciting or too boring) Habituate to test arena, won’t explore in unfamiliar locations (10-15 minuets over a couple days). Use a pair of identical objects during the habituation stage and do not use these same objects during the actual recognition experiments. You will need to develop an operational definition of object exploration. The following is an example: “A rat was considered to be engaged in object exploration when its head was oriented within 45° of an object and within 4 cm of it. Rearing with the head oriented upward was also included if at least one forepaw was on the object. Climbing over or sitting on the objects was not included” (Mumby et al., 2002). Alternatively, Any-Maze can be configured to score object exploration objectively. Test:

Place a pair of identical sample objects in the arena.
Place the rat in the arena for 5 minutes
Remove the rat to its homecage for the predetermined latency (e.g., 1 minute).
Remove the objects and place a new copy of the sample object along with a novel object.
Place the rat back in the arena for 3 minutes.
For each minute, score how long the rat investigates both the sample and the novel objects.
Run a total of 5 such trials
Provide the data to your TA who will collate the data and provide summary data to you.