TITLE: Pole Test Assessment
AUTHORS: Ian N Krout, Alexandria White, Tim Sampson

[DESCRIPTION]
Pole descent
This test measures the time for mice to descend a 24-inch pole wrapped in mesh. Mice were trained
for two days with three trials: 1) placed head down ⅓ of the height from the
base, 2) placed head down ⅔ of the height from the base, and 3) placed head
down at the top. On testing day, mice were placed at the top of the pole for
three trials and the time to descend the pole was recorded. The time was
recorded when the hindlimbs reached the base, with a maximum time of 60
seconds. Mice that fell or slid were given a score of 60 seconds.
Protocol has been approved by the California Institute of Technology’s Institutional Animal Care
and Use Committee (IACUC).

[STEPS]
SECTION: Pole set-up
2. Find a metal pole 1cm in diameter with a height of ~50-60cm.  This should have a base that is able to fit inside of a standard mouse cage.
SECTION: Training Day 1
7. Determine the order of assessment that you will use for the length of the training and assessment, record this in lab notebook.
SECTION: Assessment (test day)
16. On test day, mice is assessed for their ability to descend the pole.
SECTION: Training Day 2
15. Repeat all steps from training day 1 no more. At this point the mice should be able to perform all trials without issue. 
SECTION: Pole set-up
3. Wrap the pole with an adhesive foam or PVC material (see material for link). This should cover the entire pole and be replaced as it wears down.
SECTION: Pole set-up
4. Tape the top and bottom of the adhesive to secure to the pole.
5. Wipe down the base and pole with 70% EtOH before beginning any trials and between each mouse.
SECTION: Acclimation
6. On each training and assessment day bring the mice up from the vivarium to the behavior room at least 1h prior to assessment to allow for acclimation to the environment prior to assessment.
SECTION: Training Day 1
9. Place a single mouse, now containing the pole apparatus, allowing them to roam freely for 30 seconds.
SECTION: Training Day 1
10. Training day 1 will then consist of three trials for each mouse.
SECTION: Training Day 1
11.1. Trial 2: Place mouse face down ⅔ of the height from the base of the pole and
allow to descend freely. 
SECTION: Training Day 1
11.2. Trial 3: Place mouse face down at the top of the pole and
allow to descend freely. 
SECTION: Training Day 1
12. In between each trial allow the mouse to roam the home cage for 30 seconds.
SECTION: Training Day 1
13. In some instances a mouse may not complete all trials. In this case, remove the mouse from the pole allow to roam free, and start back over at trial 1 until the mouse has completed all 3 trials.
SECTION: Assessment (test day)
17. Place mouse face down at the top of the pole and
allow to descend freely. 
SECTION: Assessment (test day)
18. Begin the stopwatch/timer as soon as you place the mouse on the pole. Stop timing once it fully descends and all 4 paws are touching the ground. 
SECTION: Assessment (test day)
21. Allow the mouse to free roam post descent for 30 seconds.
SECTION: Assessment (test day)
22. Repeat this 2 more times (total of 3 trials) for each mouse.
SECTION: Assessment (test day)
20. Record the time-to-descend (TTD) in your notebook.
SECTION: Assessment (test day)
23. Clean the pole and base between each mouse using 70% EtOH. 
SECTION: Assessment (test day)
24. After all mice in a cage are assessed, replace mice in the cage and place pole (cleaned) into a new home cage.
SECTION: Training Day 1
8. Pole is placed inside a clean cage filled with bedding.
SECTION: Training Day 1
11. Trial 1: Place mouse faced down ⅓ of the height from the base  and
allow to descend freely. 
SECTION: Training Day 1
14. After completion of three trials, each mouse is placed in a clean cage. After all mice from the same cage have completed their trials transfer all mice back to their home cage.
SECTION: Information
1. Experiments involving GF and GF x SPF comparisons were conducted inside germ-free isolators. For comparisons involving only SPF animals, experiments were completed in a biological safety cabinet. All experiments were performed between ZT6 and ZT10 of the light phase and started at the same time each day.
SECTION: Assessment (test day)
19. The time was recorded when the hindlimbs reached the base, with a maximum time of 60
seconds. Mice that fell or slid were given a score of 60 seconds.