The WGTA apparatus (described in Harlow, 1959) is used for a battery of tests including Black/White Discrimination, Black/White Reversal, Hamilton Search, Hamilton Search Set Breaking, Hamilton Search Forced Set Breaking, and Learning Set. The following is a general description of the procedures for each task. Read every part of this protocol carefully.

Procedures for All Phases

1. Always check to make sure that the apparatus is working properly—door closes completely, clock works properly, etc., before start of testing.
2. Check to see whether any of the subjects assigned to you are sick or otherwise not to be run.
3. Transfer all animals to and from the apparatus in transport cages.
4. Keep interaction with the animals to a minimum; do not interact at all while the animal is in the apparatus, except as specifically noted in the protocols.
5. Clean the transport cage after each use.
6. Clean the WGTA cage with a spray-on disinfectant between animals, and wash it at the end of the day.

General Information—WGTA Testing

1. The data sheets used for WGTA testing are forms 18-39 in the Appendix. The heading on the data sheets includes the same basic information for all tasks: Animal number, Problem, Tester, Date, and Time. Enter all information in pencil for easy editing.

   Animal code: Enter the animal's six-digit number.

   Problem codes: 01  Black/White Discrimination Adaptation
   02  Black/White Discrimination, White Rewarded
   03  Black/White Discrimination, Black Rewarded
   04  Black/White Reversal, White Previously Rewarded
   05  Black/White Reversal, Black Previously Rewarded
   06  Hamilton Search Adaptation
   07  Hamilton Search
   08  Hamilton Search Set Breaking
   09  Hamilton Search Forced Set Breaking
   11  Learning Set—A Rewarded
   12  Learning Set—B Rewarded
   

   For Learning Set, enter the stimulus pair number at the top of the box for each problem.

   Tester code: A 2-digit experimenter number will be assigned to anyone running WGTA experiments.

   Date code: Enter month, day and year as six digits.

   Time code:
   1  until 12 noon
   2  12 noon until 5 p.m.
   3  after 5 p.m.
   

   Error Codes
   -1  Subject makes choices but fails to choose correct box after 3 min; testing
   -2  Subject makes no choice within 3 min; after 5 consecutive balks, end testing
   -3  Subject balks due to external noise
   -4  Missing data due to timer failure
   -9  Trial not run

2. Record latencies in the space provided, rounded to the nearest second (i.e., 18.4 sec = 18, 18.7 sec = 19, 19.5 sec = 20).
3. Run each animal according to the condition in its home cage; e.g., give it a diaper if it has one in its home cage, but otherwise don't.
4. If an animal will not run for 3 min on five successive trials, terminate the run. In any case, an animal must receive five trials in a day, even if all are balks. If an animal does not make a correct choice in Hamilton Search in 3 min, enter a -1 in the latency column and go on to the next trial.
5. Do not talk to the animals.
6. Set up each run after the animal has been placed in the testing cage.
7. Always note any mistakes you make and report them to the supervisor.
8. Record any behavior that is out of the ordinary; e.g., an animal acts terrified of an object in the Learning Set and refuses to respond or responds only to the other object (be it a rewarded one or not).
9. If balking or other circumstances not covered by this protocol arise, see your supervisor.

Problem 01—Black/White Adaptation

Use Yellow Block for adaptation.

For each stage, run each subject to a criterion of five trials in a row, each having a latency of 60 sec or less. Each animal receives a maximum of 25 trials per day. Each trial has a maximum time limit of 180 sec (3 min). If an animal does not complete a given stage within 25 trials, stop testing for that day and start on the same stage the next day. If an animal balks (no response for 3 min in five successive trials) under these circumstances, move back one stage at a time until it responds. Then proceed with adaptation. During all stages of adaptation, lower the screen between trials and keep the one-way mirror screen down. Start with Stage 2, running Stage 1 only if the animal will not do Stage 2 for 5 consecutive test days.

Stage 1. Run this stage only if the animal balks at Stage 2 for a week. Run the animal to criterion, offering it food from your hand, with your hand centered between the food wells.

Stage 2. Using the Gellerman randomization order, place reward in front of the food well.

Stage 3. Using Gellerman, place reward in the food well.

Stage 4. Same as 3, but place the adaptation block in back of the food well.

Stage 5. Cover the food well half with the adaptation block, using the same procedure.

Stage 6. Cover the reward completely with the object, using the same procedure. Run 23/25 of stage 6 for adaptation. For this stage of adaptation only, trials do not have to be under 1 min.

Note: For Black/White Adaptation only, record balks as follows:

			Latency	Stage
1	R	1	180	6	-2
2	L	0	18	6	1
3	R	1	180	6	-2

In trials 1 and 3 of this example, the animal balked. In trial 2 it responded correctly after 18 sec.

Problem 02—Black/White Discrimination, White Rewarded
Problem 03—Black/White Discrimination, Black Rewarded

1. Always reward either black or white for each animal. If you start your first animal with black, then start your second with white, your third animal with black, etc. In this way all animals will not be having the same color reward for Black/White Discrimination.
2. Use the Gellerman series form for this test (Appendix, forms 19-23). Begin your first animal with Series I, your second animal with Series II, etc. The different Gellerman series represent different randomizations of right/left reward.
3. Rewarding one color only for each animal, cover the appropriate well following the Gellerman randomization schedule. Criterion is 23 correct choices in 25 trials.
4. A balk is scored in Black/White Discrimination and Reversal when the subject has not responded for 3 min on five consecutive trials. The animal must be tested for five trials, which can include the balk trials, on any given day.
5. Scoring: Always enter correct latencies in the left-hand column of the form and incorrect latencies in the right-hand column. In the far right-hand column, indicate whether the animal made the correct choice (code 1) or an incorrect choice (code 0). All this scoring procedure facilitates data entry. A balk (code -2) should be written across the latency columns, and no mark made in the far right-hand column. If five balks occur in a row and testing is terminated, fill remaining blank columns with -9, in the same fashion as a balk is filled in.

Problem 04—Black/White Reversal, White Previously Rewarded
Problem 05—Black/White Reversal, Black Previously Rewarded

1. After an animal has run to criterion on Black/White Discrimination, begin Black/White Reversal. On this task reward black if white was rewarded on the initial discrimination series, and white if black was rewarded on the discrimination series. Always begin a new data sheet when Reversal is begun.
2. Reward the appropriate well according to the Gellerman randomization schedule (Appendix, forms 19-23). Run Black/White Reversal until the animal reaches criterion (23 correct responses in 25 trials).
3. The scoring is the same as for Black/White Discrimination: use the left-hand column for correct latencies and the right-hand column for incorrect latencies. Again, use the far right column to indicate whether the response was correct (code 1) or not (code 0). Code -2 (balk) extends across latency columns.

Problem 06—Hamilton Search Adaptation

1. Adaptation for Hamilton Search is done in two days (unless the animal has not learned how to manipulate the box). Using the single green box, begin adaptation with Stage 1. Record latency and stage on the Hamilton Search Adaptation Sheet (Appendix, form 24). If an animal balks on any one stage five times in a row, return to the previous stage the next testing day.
2. Stage 1. Place a reward in the box and hold the lid open. When the animal has responded five times in a row with latencies of less than 60 sec, continue to Stage 2.
3. Stage 2. Place a reward in the box, holding the box half open with your hand. Criterion is the same as in stage 1.
4. Stage 3. Place a reward in the box and leave the box closed. Criterion is correct responses on 23/25 trials. This stage will take an entire testing session.

Problem 07—Hamilton Search

1. The four little yellow boxes are used for this task. On the back of each box is a letter to help you randomize the box order each day. It doesn't matter what order the boxes are in. Just make sure that the boxes are in a different order for each animal each day. Make a note of the box order on the Hamilton Search data sheet.
2. There are three Hamilton Search data sheets, each with a different randomization schedule for the baited box. Do not use the same series sheet two days in a row. (See Appendix, forms 25-27.)
3. The Hamilton Search boxes are numbered 1-4 as seen from the experimenter's left to right. The numbers 1-4 refer to position. For each trial, bait the box listed in the order or "bait" column in the data sheet. For example:

   A	C	B	D
   1	2	3	4

4. After the correct box is baited, be sure all four boxes are completely closed, raise the door, start the timer, and record the order of box openings in the spaces marked Box 1, Box 2, Box 3, and Box 4. For example:

   Box 1	Box 2	Box 3	Box 4
   4	2	5	1,3

   In this example, Box 1 was the fourth box opened; Box 2, the second; Box 3, the fifth; and Box 4, the first and third. The Box 3 answer is circled because it is the correct response. Run 25 trials. Record latency to correct choice for each trial in the space marked "latency."

   Note: The subject only needs to open the correct box to terminate the trial—be sure the animal receives the reward, but do not record the time it takes to remove the reward.

5. Summary. To facilitate data entry, you must summarize your own data. In the column marked first choice, enter the first choice the animal made. In the case of the previous example, enter a 4 because Box 4 was opened first. In the four spaces following "first choice," enter how many times each of the boxes was opened. If a box was not opened at all during the trial, enter a 0. Do not leave the space blank.
6. Run the Hamilton Search Problem for 5 days. Balks during the run are acceptable; however, if five balks in a row occur (resulting in termination of testing), that day cannot count as one of the 5 days.
7. Remember: An incident in which an animal makes choices but doesn't make the correct response has an error code of -1. This does not count as a balk.

Problem 08—Hamilton Search Set Breaking

1. After Hamilton Search has been run for 5 days, determine which box was the least preferred. To do this, enter in a table how many times each was opened first for each day. For example:

   	Box 1	Box 2	Box 3	Box 4
   Day 1	10	3	8	4
   Day 2	8	5	6	6
   Day 3	11	2	8	4
   Day 4	7	3	10	5
   Day 5	6	4	8	7
   Total	42	(17)	40	26

   Across each day, the number of first lid openings should add up to 25 since there are 25 trials per day. In this case, Box 2 was opened first the least number of times.

2. The least preferred box is always baited for Hamilton Search Set Breaking. On the data sheet (Appendix, form 28) mark in red each of the entries in the "bait" column with the number of the least preferred box.
3. Run Hamilton Search Set Breaking the same as Hamilton Search—only reward the same least-preferred box for each of the 25 trials. Enter the order the boxes are opened in the same manner and summarize data in the same way.
4. Hamilton Search Set Breaking is run for 5 days. (Again, if an animal balks five times in a row resulting in termination of testing, that day cannot count as one of the 5 days.)

Problem 09—Hamilton Search Forced Set Breaking

1. For this problem, bait the same least preferred box as in Set Breaking for all 25 trials.
2. The difference between Forced Set Breaking and Set Breaking is that in Forced Set Breaking, the animal is given the opportunity to open only one lid before the door is brought down. It is extremely important that the animal not be allowed to make more than one choice, no matter how fast it is. Be faster.
3. Scoring for Forced Set Breaking is the same as for Set Breaking and Hamilton Search, except that there is just one entry per trial in the spaces provided for the order of box openings (Appendix, form 29).
4. In Forced Set Breaking, the summary is somewhat simplified. It is still necessary to indicate the first choice (which is the only choice), but there is no need to fill in the spaces to indicate how many times each box was opened.
5. Hamilton Forced Set Breaking is run for five days. The same balk rule as in Problems 07 and 08 applies.

Problem 11—Learning Set, A Rewarded
Problem 12—Learning Set, B Rewarded

1. There is no adaptation for Learning Set.
2. Learning Set is run for 36 trials per day (six item pairs, six trials with each pair).
3. In the WGTA run, there are 240 pairs of toys bagged in sets of six pairs each. Each box of toys represents 5 days of testing or running. The toys in each pair are labeled A or B. For any given animal, always reward either the A toy or the B toy throughout the 240 sets of toys. In other words, for a particular animal, A is rewarded all the time or B is rewarded all the time.
4. Each bag of toys represents a day of running. Begin running with the bag labeled 1-6 for the first day, 7-12 for the second day, etc. Write down the number of the pair of toys consecutively on the data sheets—on top of the data block for that particular set of toys.
5. Present each pair of toys to the animal six times, representing six trials per pair. Present six pairs of toys per day.
6. Learning Set is run similar to Black/White Discrimination. Place the reward in the wells according to the randomization schedule on the six-trial discrimination forms (Appendix, forms 30-39).
7. Enter the latency for a correct choice in the left column and the latency for an incorrect choice in the right column. Then in the far right column, enter a 1 if the choice was correct and a 0 if the choice was incorrect.
8. Learning Set takes approximately 8 weeks to run.
9. Balking is defined as 180 sec of no response. If the subject balks for 5 consecutive trials (this can occur over two toy sets), stop testing. Do not present the same toys to the animal on the next day's run, e.g., if an animal stops running on pair #157, start with set #158 the next day, finishing that six-pair bag of toys. For example, if four pairs had been run successfully before the animal balked, run only five and six the next day. Start the following test session with the next six pairs.

WGTA Data Processing
After data are collected for individual animals on the entire WGTA process, they are edited for computer entry. This edit is to ensure that testers have put all relevant information on the animal's daily data sheets (i.e., animal number, problem number, date, time, tester number) and that the data are presented correctly, with all data information available.

Be sure that your data sheets are completely filled out, as any errors in the data are difficult and time consuming to rectify.

References
Boothe, R. and Sackett, G. Perception and learning in infant rhesus monkeys. In G. Bourne (ed.), The Rhesus Monkey, p. 343-363. New York: Academic Press, 1975.

Harlow, H. The development of learning in the rhesus monkey. American Scientist 45:459-479, 1959.

[Table of Contents]

Subjects
Nonhuman primates

Schedule
Monday through Friday, generally between 8 a.m. and 5 p.m.

Procedures
Standard procedures for all phases of the learning tests are as follows:

1. Before testing, check the WGTA apparatus to be sure it is working properly.
2. Wear a lab coat and examination gloves at all times when handling and testing animals. Change gloves between animals.
3. Transport animals to and from the testing room in infant carriers.
4. Clean infant carriers after each animal.
5. Clean the WGTA cage at the end of the day.
6. If an animal cannot be tested because it is sick or because of experimenter error, write the reason in the daily log book.

General Information

Header Information
Test day and problem are already printed on the data sheet (a few examples are shown in the Appendix, forms 40-42). Fill out the rest of the header information completely and accurately. Include:

Animal code: the animal's six-digit number.

Procedure code:
01  Adaptation 1 (white board)
02  Adaptation 2 (white board)
03  Nonmatching to sample with familiar stimuli (blue board)
04  Nonmatching to sample with familiar stimuli (yellow board)
05  Nonmatching to sample with novel and familiar stimuli (blue board)
06  Nonmatching to sample with novel and familiar stimuli (yellow board)
07  Matching to sample with novel and familiar stimuli (reversal-yellow board)
08  Matching to sample with novel and familiar stimuli (reversal-blue board)
09  Conditional learning set with familiar stimuli (blue-nonmatch, yellow-match)
10  Conditional learning set with familiar stimuli (yellow-nonmatch, blue match)
11  Conditional nonmatching and matching to sample with novel and familiar stimuli (blue-nonmatch, yellow-match)
12  Conditional nonmatching and matching to sample with novel and familiar stimuli (yellow-nonmatch, blue-match)
13  Nonmatching to sample warm up with novel and familiar stimuli (blue board)
14  Nonmatching to sample warm up with novel and familiar stimuli (yellow board)
15  Nonmatching to sample 15-sec delay with novel and familiar stimuli (blue board)
16  Nonmatching to sample 15-sec delay with novel and familiar stimuli (yellow board)
17  Nonmatching to sample 30-sec delay with novel and familiar stimuli (blue board)
18  Nonmatching to sample 30-sec delay with novel and familiar stimuli (yellow board)

Tester code: Your two-digit experimenter number.

Date code: Month, day and year as six digits.

Time code:
1  before 12 noon
2  12 noon to 5 p.m.
3  after 5 p.m.

Scoring Responses
Use the following codes when an animal completes a trial on its first presentation:

0 Response incorrect
1 Response correct

Problems and Errors That Can Occur During Testing
When you are testing an animal, certain problems can arise. The most frequent problems are that the animal balks (won't respond) or a test stimulus breaks mid-trial. A balk is defined as refusal to respond within 45 sec to the sample stimulus or refusal to respond to the test stimuli within 45 sec after displacing the sample stimulus. If either of these situations occurs, record an asterisk near the trial number on the data sheet. Do not record any latencies at this time. Should the animal continue balking for five consecutive trials, end testing for the day. On the next day the animal is tested, resume testing from the first of the five previous balk trials. In other words, if an animal is to be tested for 24 trials and balks on trials 5-10, start testing the next day on trial 5. Finish that data sheet and wait until the next day to start a new one.

If an animal balks intermittently during a test session but does not balk for five consecutive trials, continue testing until the data sheet is completed. Then go back and rerun the balk trials one more time. In summary, indicate all balk trials on the first presentation with an asterisk. Do not record latencies or responses at this time. When the balk trials are rerun (either the same day or the next), record the appropriate latencies as well as one of the following response codes:

2  Rerun balk, response incorrect
3  Rerun balk, response correct
4  Rerun balk, balk repeated

If a test object or the apparatus breaks mid-trial, code the trial as a 5. Again, write this code in the response column.

A less frequent problem will be the placement of the test object on the wrong side. Code right as 1 and left as 0. If you notice after a trial that you have placed the object on the wrong side, change the number on the data sheet (e.g., if you have placed it on the left instead of the right, change the 1 to a 0).

If other errors or problems occur and you feel the need to comment on them, please do so on the back of the data sheet. The data will eventually be entered in a computer and unnecessary writing on the data sheet is distracting to the computer operator.

To summarize, problems such as balks, test object breakage, and experimenter error can occur during a test session. The most frequently occurring problems have error codes that need to be entered in the response column of a particular trial. Familiarize yourself with codes 2-5 to record discontinued or balk trials accurately.

Toy Grabbing
It is not uncommon for monkeys to try to grab the stimuli and pull them into the testing cage. If the animal successfully pulls the sample toy into the test cage, stop timing and close the center barrier. Immediately retrieve the toy from the monkey. (These animals have been hand-raised and will not bite unless frightened or provoked. Just reach into the testing cage and remove the toy.) Place the sample stimulus behind the center well, raise the opaque screens, and proceed with the trial as usual. If the animal grabs one of the test stimuli, stop timing and retrieve the toy from the monkey. Record latencies and the response as usual. The response will be correct or incorrect regardless of whether the stimulus is calmly displaced or pulled into the cage. If a monkey is a chronic toy grabber, you can chain down the stimuli to avoid the constant retrieval of toys.

Test Stimuli
All stimuli used in the Advanced Learning Series are drawn from two pools of objects. Toy pairings 1-10 are considered familiar stimuli and pairings 11-130 are considered novel stimuli. Novel and familiar stimuli can be combined in four ways: 1) sample and test object novel; 2) sample and test object familiar; 3) sample novel, nonmatch familiar, match novel; 4) sample familiar, nonmatch novel, match familiar. These pairing types were designed to assess the effects of relatively novel and familiar stimuli on the development of learning strategies. do not alter the pairings that are shown for each trial!

Board Colors
Before an animal begins Advanced Learning, it will be assigned a specific color combination. Two combinations are possible: 1) blue-nonmatch and yellow-match, and 2) yellow-nonmatch and blue-match. If your animal is assigned blue-nonmatch and yellow-match, run all nonmatching to sample tests on a blue presentation board. When the animal begins the matching to sample task, present all trials on a yellow board (i.e., the board color that is opposite from the one that was used during the nonmatch tests). On the conditional tests, the animal is required to nonmatch when the board is blue and match when the board is yellow. Note that the subject was presented with the blue board on all previous nonmatch tests and the yellow board on the previous match test. If your animal is assigned the yellow-nonmatch and blue-match color combination, the test progression is identical except that the opposite board colors are used. This procedure is used to counterbalance the test animals across the color combinations. Examples of both color combinations follow:  

Phase 1—Adaptation (White Board)

Adaptation 1—Middle well baited, no test stimuli
Put a reward in the middle well and cover it with an object. Opaque Plexiglas screens on either side of the middle well (where the sample will be placed during the actual tests) will prevent the animal from touching and seeing the test objects until the sample has been displaced. During the first part of adaptation, there will be no stimuli behind these screens. Run a total of 25 trials per day.

There are three stages during Adaptation 1:

Stage 1—well baited, toy behind well

Stage 2—well baited, toy covering half of well

Stage 3—well baited, toy fully covers well

Begin with stage 1. When an animal responds on five consecutive trials in less than 45 sec per trial, go on to stage 2. When an animal responds on five consecutive trials in less than 45 sec per trial, go on to stage 3. Perform stage 3 until the animal gets 23 out of 25 correct in one test session. This is criterion, so proceed to Adaptation 2 the next day. If an animal balks on five consecutive trials at any stage, stop testing for the day. The next day, begin testing at the stage previous to the one the animal balked on. For example, if an animal balked on stage 3, begin with stage 2 on the next testing day. Present trials at stage 2 until the animal has responded for five consecutive trials and then go on to the next stage. Record stage number (1, 2, 3) on the data sheet for each trial (Appendix, form 40).

Adaptation 2—Side well baited, no test stimuli
Bait one of the test wells randomly. Place the adaptation object over the center well. Do not bait the center well. After the monkey displaces the object, raise the opaque screens so the animal can obtain the reward. Run 25 trials per day. Test the animal on this task until it displaces the center object and takes the reward 23/25 times in a single test session. Record results (Appendix, form 41).

Phase 2—Nonmatching to Sample (Yellow or Blue Board)

Nonmatching to Sample with Familiar Stimuli
This procedure is designed to introduce the animal to the concept of nonmatching to sample. Set the test objects on a three-well presentation board before raising the center screen of the WGTA apparatus to begin the trial. Always place the sample stimulus over the middle food well and do not bait it. Place the test stimuli behind the opaque screens and over the left and right food wells. Bait the nonmatching test stimulus. After you raise the center divider and the animal displaces the sample object, raise the opaque screens to give the animal an opportunity to displace one of the test objects. As soon as the animal makes a choice, lower the opaque screens of the WGTA and record the response on the data sheet (a sample is shown in the Appendix, form 42). Give the animal 24 trials per session for 5 testing days on this procedure.

Nonmatching to Sample with Novel and Familiar Stimuli
The basic procedure is identical to that described in Part 1 except that the novel-familiar stimulus pairings are now introduced. Present six trials of each of the stimulus variations per session for a total of 24 trials per day. Run the animal either 15 days or 20 days on this procedure. When it completes 15 days of this task, calculate its mean performance level over test days 13, 14, and 15. If, across these 3 days, the animal's mean performance is 80% correct or greater, proceed to the next test in the series (matching to sample). If the animal has not reached this standard of performance, continue running the same procedure for an additional five sessions. After the 5 additional days, go on to Phase 3 regardless of the subject's final level of performance.

Phase 3—Matching to Sample (Yellow or Blue Board)

Matching to Sample with Novel and Familiar Stimuli
This phase consists of one test in which the animal is required to match rather than nonmatch to sample. This provides a measure of a subject's ability to reverse a previously learned strategy. The procedure is the same as that outlined for Phase 2 except that the test stimulus that matches the sample is now rewarded. The nonmatching test object has become the incorrect choice and should not be baited. Present six trials of each novel-familiar pairing type per session for a total of 24 trials per day. Test animals either 25 or 30 days on this procedure. When the subject completes 25 days of matching to sample, calculate its mean level of performance over test days 23, 24, and 25. If, across these 3 days, the mean percentage correct is 80% or greater, advance to Phase 4. If the animal is not performing at this level, continue testing with the same procedure for five more sessions. When 30 days of this task have been completed, proceed to the next test regardless of the animal's final level of performance.

Phase 4—Conditional Nonmatching and Matching to Sample
(Yellow and Blue Board)

This phase consists of two parts: conditional learning set and conditional nonmatching and matching to sample. The correct response on this task is contingent on the color of the presentation board. Half of the animals will be required to match to sample when the board is yellow and nonmatch when the board is blue. The other half will be required to match to sample when the board is blue and nonmatch to sample when the board is yellow.

Conditional Learning Set
This task is designed to introduce the idea of using the color of the presentation board as a cue. The basic procedure is the same as that described for the nonmatch and match tests, but each trial is presented twice. Present five nonmatch problems and five match problems each day. For a two-trial learning set procedure, this will total 20 trials per day. Test the animal on this procedure for 16 days.

Conditional Nonmatching and Matching to Sample
This procedure differs from conditional learning set in that each trial is presented only once. Present six trials of each novel-familiar variation each session for a total of 24 trials per day. Test the animal either 35 or 40 days on this procedure. When it completes 35 days of this task, calculate its mean level of performance over test days 33, 34, and 35. If the score is 80% or greater, the animal should be advanced to the next test in the series. If it is not performing at this level, continue testing with the same procedure for an additional five sessions. When the animal completes 40 days of this task, proceed to the next test regardless of its final level of performance.

Phase 5—Delayed Nonmatching to Sample (Yellow or Blue Board)

Nonmatching to Sample Warm Up
This task is designed to reacquaint the subjects with a testing paradigm that requires only the nonmatch response. The basic procedure is identical to that described for the earlier nonmatch tests. Give the animal 24 trials each session and test it for either 5 or 10 days on this procedure. When the subject completes 5 days of nonmatching to sample warm up, calculate its mean level of performance across test days 3, 4 and 5. If the mean percentage is 80% or greater, advance the animal to the 15-sec delay task. If it is not performing at this level, continue testing on nonmatch warm-up for an additional 5 days. When the animal has completed 10 days of this task, proceed to the 15-sec delay task regardless of its final level of performance.

Nonmatching to Sample—15-sec delay
This procedure requires a delay between the displacement of the sample and the presentation of the test stimuli. As in earlier procedures, set the stimuli on the presentation board with the center divider of the WGTA down. As soon as the animal displaces the sample, remove it from the board and begin a 15-sec delay. After 15 sec have elapsed, raise the opaque screens surrounding the test stimuli and allow the animal to make a choice. Give the animal 24 trials per day for 5 days on this procedure.

Nonmatching to Sample—30-sec delay
This is essentially the same task as the 15-sec delay except that the delay period is lengthened to 30 sec. Give the animal 24 trials each day for 5 days.  

Reference
Fredrickson, W.T., Gould, P.L., Gunderson, V.M., and Grant-Webster, K.S. Complex learning in low birthweight and normal birthweight juvenile pigtailed macaques (Macaca nemestrina). Develop. Psychol. 23:483-489, 1987.

[Table of Contents]

Purpose

• To prepare animals for housing in large peer groups of 10-20 monkeys.
• To document the pattern of adaptation during the first 2 weeks of permanent group housing using indices of behavior and pituitary-adrenal axis responses. The housing change from individual cage to group housing in a novel room may constitute a particularly stressful challenge for some individuals. Use the following procedures to document each individual's unique pattern or style of adapting when it is first introduced to living with peers in a group cage.

Subjects
Nonhuman primate infants, 9 months or older, that meet the following criteria:

• have been members in the group for at least 4 weeks
• have participated daily in a standard socialization schedule as infants
• have been observed for at least the week prior to the housing change and preferably for 2 weeks
• have completed the WGTA protocol series if tested on the battery
• are not on medical treatment.

Procedures
Introduction to the New Group Cage
Place each member of the social group in a transfer cage and carry it to the group cage ready to be released. When all members are in transfer cages and ready, release them as quickly as possible between 2:55 and 3:00 p.m. Everyone except the behavioral observer should leave the area after the release.

Blood Sampling
All blood draws are at 3:30 p.m. Two basal samples are taken 1 to 2 weeks prior to the housing change, matched for day of week as day 1 and day 3. After the housing change, blood is drawn 30 min post-introduction, then on days 3, 7 and 14.

All samples must be obtained within 3 min of initial approach toward the animal. Draw blood with a 22-gauge needle (or smaller) and a 3-cc syringe. The needle will be heparinized with sodium heparin. Draw 1 cc blood from the femoral vein and occlude the area immediately after the draw. Unscrew the needle and inject the blood sample into a green-top vacutainer. Centrifuge immediately, then draw and freeze the plasma. The plasma is later assayed for cortisol and DHEA-S.

Behavioral Observations
All behavioral observations are collected with a digital data acquisition system with a key to measure duration of locomotion. The coding system follows the plan for Playroom Testing (see p. 45), without the 4th digit.

Base-line observations. Collect base-line observations on each member of the group during playroom sessions at least 1 week before the housing change.

Post-housing change observations. Morning observations are optional, done whenever time permits. Begin morning observations between 9:00 and 9:30. Afternoon observations are mandatory; begin at 3:00 and finish by 3:30.

On day 0 begin observations immediately after the animals are introduced to group housing. Observe each animal two times for 3 min each time in a counterbalanced order (for a total of 6 min of observation time per subject). On days 1 through 14, observe each animal once for 5 min in a counter-balanced order.

[Table of Contents]

The use of a standard protocol as the monkeys progress through changes in their feeding and housing is extremely important to both research goals and animal health care. The following charts detail when changes in feeding and housing occur. The first chart outlines the schedule for feeding changes. The second chart outlines the schedule of housing changes from incubator to group housing. Prepare feeding and housing schedules (Appendix, forms 43 and 44) for each animal and mark all changes as they occur. Record daily intake on the intake data sheet (Appendix, form 45). Record all housing movements on the movement sheet (Appendix, form 46).

These procedures should not be adhered to at the expense of an animal's health. The veterinary staff will examine the progress of each animal daily, thus ensuring that no health hazards exist. When a monkey is taken off this schedule for health-related purposes, the nursery personnel and veterinary staff will establish an individual feeding regimen for that monkey. When the monkey has returned to health, the normal feeding schedule will be reinstated as appropriate. If you have any questions regarding these procedures, talk to the nursery manager, a research technologist, or the lab supervisor at any time.

Reference
Ruppenthal, G.C. Survey of protocols for nursery-rearing infant macaques. In G.C. Ruppenthal and D.J. Reese (eds.), Nursery Care of Nonhuman Primates, p. 165-186. New York: Plenum Press, 1979.

Days of Age	How Fed	What Fed	Delivery Type	Frequency	Explanation
0-1	bottle	Polycose	nipple	2 hr	For first 5 feedings after infant is removed from mother, feed Polycose every 2 hr.
1-4	bottle	50% SMA	nipple	2 hr	From sixth feeding until 6 a.m. on 4th day, feed 50% SMA every 2 hr. The 8 a.m. feeding on day 4 is the last 50% SMA 2-hr feeding. At noon on 4th day begin feeding 100% SMA.
4-9	bottle/feeder w/surrogate	100% SMA	nipple	4 hr	8 a.m.: Bottle feed. 12 noon: Feeder box; 150 cc SMA. Hold infant up to feeder at noon. Measure intake and record for noon feeding with code = held up. Replace feeder. 4 p.m.: Measure feeder and record amount taken as code = self-feeding (make sure to subtract amount taken when infant was held up at noon to determine amount taken by self). If amount taken from feeder was 0, bottle-feed at 4 p.m. and record intake with code = bottle-fed. Do not replace feeder. Rest of feedings: Bottle feed.
9-12	feeder w/surrogate	100% SMA	nipple	continuous 4-hr changes	The 8 a.m. feeding on the 9th day is the last bottle feeding. Beginning at noon on day 9, do the following: Feeder box; 150 cc SMA. Hold infant up and record amount with code = held up. Replace feeder. For rest of feedings until day 12, do the following: measure amount taken from feeder (make sure to subtract amount taken from previous feeding) and record with code = self-feed. Clean feeder, replace with 150 cc SMA. If intake was 0 for self-fed portion, hold infant up and measure amount taken. Record with code = held up. If intake is sufficient without holding up, stop trying.
12-14	feeder w/o surrogate	100% SMA	nipple	continuous 4-hr changes	Beginning at 8 a.m. on day 12, change, measure, and refill feeders with 150 cc SMA every 4 hr. Record intake with self-feeding code. Do not hold infant up at this time.
14-21	feeder	100% SMA	nipple	continuous 4-hr changes	At 8 a.m. on 14th day, remove infant from nursery and set up feeder in cage in quad room. Change, measure, and refill every 4 hr. Begin to give chow at this time.
21-56	feeder	100% SMA	nipple	4 hr on/ 4 hr/off	At noon on 21st day, remove feeders; measure and record intake. Put feeders back on at 4 p.m. Continue 4-hr on/4-hr off schedule until day 56.
56-91	feeder	100% SMA	rubber tube	4 hr on/ 4 hr off	At noon on 56th day, remove feeders with nipples; measure and record intake. Put feeders with rubber tubes back on at 4 p.m. Do this routine until day 91.
91-98	feeder	60% SMA	rubber tube	4 hr on/ 4 hr off	At noon on 91st day, remove feeders with 100% SMA, measure and record intake. Return feeders with 60% SMA at 4 p.m. Do this routine until day 98.
98-112	feeder	40% SMA	rubber tube	4 hr on/ 4 hr off	At noon on 98th day, remove feeders with 60% SMA, measure and record intake. Return feeders with 40% SMA at 4 p.m. Do this routine until day 112.
112-114	feeder	40% apple juice	rubber tube	continuous 8-hr changes	At noon on the 112th day, remove feeders with 40% SMA; measure and record intake. Return feeders with 40% apple juice immediately. Have feeders on continuously. Change every 8 hr. Do this routine until day 114.
114-119	feeder	20% apple juice	rubber tube	continuous 8-hr changes	At noon of the 114th day, remove feeders with 40% apple juice; measure and record intake. Return feeders with 20% apple juice immediately. Do this routine until day 119.
119-122	feeder	100% water	rubber tube	continuous 8-hr changes	At noon on the 119th day, remove feeders with 20% apple juice; measure and record intake. Return feeders with 100% water immediately. Do this routine until day 122.
122-126	feeder	100% water	steel tube	continuous 8-hr changes	At noon on the 122nd day, remove feeders with rubber tubes; measure and record intake. Return feeders with steel tubes immediately. Do this routine until day 126.
126 and after	feeder	100% water	steel tube	continuous changes but not measured	At noon on the 126th day, remove feeders; measure and record intake. Replace feeders. Care will be provided by animal technician from this time on. Recording intake stops at this point. Weights are taken weekly instead of daily.

Days of Age	Housing Type	Temperature Support	Comments
birth	incubator (diaper)	none	At delivery, place infant in incubator at a temperature of 93°-94°. Maintain this temperature until 8 a.m. the next day.
1-2	incubator (diaper)	none	At 8 a.m. on the day after delivery, turn incubator temperature down to 90°-91°. Check infant's temperature at noon and remaining diurnal periods for rest of day.
2-3	incubator (diaper)	none	At 8 a.m. on the 2nd day, turn incubator temperature down to 87°-88°. Check infant's temperature at noon and remaining diurnal periods for rest of day.
3-4	incubator (diaper)	none	At 8 a.m. on the 3rd day, turn incubator temperature down to 84°-85°. Check infant's temperature at noon and remaining diurnal periods for rest of day.
4-10	small cage, nursery (diaper)	heating pad on	After 8 a.m. feeding on the 4th day, remove the infant from the incubator and place it with heating pad in a small cage in the nursery. Turn heating pad on. Check the infant's temperature at noon and remaining diurnal periods for rest of day.
10-14	small cage, nursery (diaper)	heating pad off	At 8 a.m. on the 10th day, turn heating pad off. Check the infant's temperature at noon and remaining diurnal periods for rest of day.
14-28	small cage, quad room (diaper)	none	At 8 a.m. on the 14th day, remove the infant from the nursery and place it without heating pad in a small cage in the quad room. Check the infant's temperature once a day from days 14 to 21.
28-140	large cage (diaper)	none	At 8 a.m. on the 28th day, place the infant in a large cage in the quad room.
140-224	large cage	none	At 8 a.m. on the 140th day, remove the diaper from the infant's cage. (Check with WGTA person to see if this instruction is inappropriate.)
224 and after	large cage	none	At 8 a.m. on the 224th day, move the infant from the quad room into the back room (RR037).|