Marble burying test

Marble Burying Protocol

References:

(1) Broekkamp CL, Rijk HW, Joly-Gelouin D, and Lloyd KL. Major tranquillizers can be distinguished from minor tranquillizers on the basis of effects on marble burying and swim-induced grooming in mice. Eur J Pharmacol. 126(3):223-9, 1986.

(2) Njung'e K and Handley SL. Evaluation of marble-burying behavior as a model of anxiety. Pharmacol Biochem Behav. 38(1):63-7, 1991.

(3) Thomas A, Burant A, Bui N, Graham D, Yuva-Paylor, LA, and Paylor R.  Marble burying reflects a repetitive and perseverative behavior more than novelty-induced anxiety. Psychopharmacology 204(2):361-73, 2009.

Before the Experiment / Setting up Cages

1. In the testing room, set the white noise at ~60 dB and illumination at 150 Lux.

2. Bring the mice into the testing room, open the air filter, mark their tails and then allow them to acclimate for at least 30 min.

* Make sure the cages are not stacked on top of each other during habituation. 

* Put aside 2-3 tables/carts.  One or two for transporting clean and dirty cages.  

   One for placing the finished cages of mice on after they have been tested.

3. Prepare 8 – 10 cages at a time for testing by sink:

1. The cages should be filled up to the bottom line (on the corner or the cages) of the cages

1. Make sure the cages are at the correct level before you use them
2. Take a half of hand full bedding from home cage and spread into testing cage
3. Change gloves between male cage and female cage

4. Shake and re-level cages

    

4. Place 20 marbles in the premade cages in a 4 X 5 matrix leaving some space on one end to place the animal.

Starting Test

1. Arrange the table with premade cages and marble in the room for testing
2. When introducing the mice place them in the space without the marbles. 
3. Place the flat tops on top. 
4. When you leave the room start the timer for 30 minutes
5. When 30 minutes are up remove the mice from the cage and put them into their original cages.

Scoring MB

1. Count total visible marbles and subtract that from 20 and that gives you marbles that were buried completely (100%).
2. Out of the visible marbles count total ½ buried and ¾ buried.
3. Number of marbles buried will be 100% buried + 50% buried. 

Clean up *Do as quiet as possible

1. Make sure there are 20 marbles in each cage when you take them out
2. Clean them with water and soap in a large plastic container
3. Rinse  them in a strainer 
4. Pour them quietly on a bench pad
5. Wipe them on top with a napkin
6. Spray them with 30%-50% isopropanol and let them air dry over night. Make sure you write your name on a piece of paper towel and leave it on top so that no one disturbs the set up. 
7. Once dried place the marbles in the container and place them back in the metal shelf. 

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