TW Innate Odor Aversions in 14-3-3 FKO mice

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Innate odor aversion in 14-3-3 FKO mice expressed as locomotor activity in response to TMT or 2PE in Odor Arena, scored by Tracker.

Adam Kimbrough, Charles E. Houpt, Yuying Wu, Dr. Thomas A. Houpt, Dr. Yi Zhou, Dr. Michael Meredith

Procedure

Male 14-3-3 FKO mice (n=6) and wildtype C57 mice (n=6), singly housed with ad lib access to food and water. Habituated to Odor Arena (without stimulus dish) for 5 min per day for 4 days.

Trials 1 - 3: response to water, TMT, or 2PE

All mice exposed to an odor stimuli for 5 min, 1 stimulus/day, for 3 consective days.

Odor stimuli are either 5 ul distilled water, 5 ul TMT (1% in water), or 1.25 ul 2PE (neat). Stimuli pipetted onto filter paper (#1 Whatman, 25mm diameter disc) and placed in small glass petri dish. Stimuli were prepared prior to bringing mouse home cage into test room. Stimulus was placed in random corner of the 29cm x 29cm arena immediately prior to mouse being placed in center of arena. Each stimulus was presented once, in random order for each mouse.

Locomotor activity video was captured with a Logitech Quick Pro webcam and an Apple iMac computer into Quicktime Player at 640x480 pixel at 15 fps. Video was processed by Tracker program. The position of the mouse, defined as the center of the mouse body as detected by Tracker to within ~2 mm, was recorded 3 times per second. From the position data, several variables were calculated: total distance traveled during the 5 min test, distance from the stimulus, etc.

Between mice, the arena was rinsed with soapy water (Alconox) and 70% isopropanol. Stimulus filter paper and pipette tips were disposed into sealed container of bleach.

Trial 4: contrasting response to water vs. TMT

All mice exposed to H20 stimulus for 5 min, then odor stimulus was replaced with TMT for 5 min to evaluate contrasting responses to water & TMT back to back.

Stimuli

TMT (ConTech, Inc) received April 1, 2013

  • Product code 3000000368
  • Lot number 11125
  • Manufactured May 5, 2011
  • Expiry May 2014

2PE (Sigma-Aldrich) received March, 2013

  • Item #77861
  • Lot# BCBH9821V

Solution Prep

made 50% TMT in water: 50 ul TMT + 50 ul water

made 1% TMT in water: 10 ul TMT + 990 ul water

aliquoted 1000 ul 2PE

aliquoted 50 ml distilled water

Random Trial Schedule

Mouse Genotype Trial 1 Trial 2 Trial 3
TW01 Tg 2PE, br TMT, tl H20, tr
TW02 Tg H20, bl TMT, br 2PE, tl
TW03 Tg 2PE, br H20, tl TMT, br
TW04 Tg TMT, bl 2PE, tl H20, tl
TW05 Tg TMT, br H20, br 2PE, tr
TW06 Tg H20, br 2PE, bl TMT, tr
TW07 wt H20, tl TMT, bl 2PE, tr
TW08 wt TMT, tl H20, br 2PE, br
TW09 wt TMT, br H20, tl 2PE, br
TW10 wt H20, bl TMT, br 2PE, br
TW11 wt H20, tr TMT, bl 2PE, tl
TW12 wt H20, tr 2PE, tl TMT, bl

tl, tr, bl, br = corner of odor arena.