Novel Object Recognition Protocol

Novel Object Recognition Protocol

Novel Object Recognition Protocol

Christine Ann Denny

Updated: 01.12.11

Information

  1. We used a modified version of the novel object recognition task described by Ennaceur and Delacour (1988).

Testing Room Equipment

  1. Two 60 W light bulbs.
  2. Tripod.
  3. Video camera.
  4. Testing arena
  5. White, plastic transport box (55 x 40 x 15 cm).
  6. Divide into two equal halves so that two sessions could occur simultaneously.
  7. The divider is made of the same white plastic as the transport box and measured 15 cm in height.
  8. Subject should not contact or see one another during the exposures.
  9. Subject bedding (wooden bedding, NOT bed-o-cobs).
  10. Luxmeter
  11. The light intensity should be equal in all parts of the arena (approximately 20-25 lx).
  12. Objects (all in triplicate)
  13. (1) a blue, ceramic shoe (diameter 9.5 cm, maximal height 6 cm);
  14. (2) a black, shiny box (8 x 3 x 9.5 cm); and
  15. (3) a clear, plastic funnel (diameter 8.5 cm, maximal height 8.5 cm).

NOR Protocol (129SvEv subjects)

  1. Subjects were transported to the testing room in their home cages.
  2. Five 5-min exposures with 3-min inter-exposure intervals.
  3. Place subject in the center of the arena at the start of each exposure.
  4. Between exposures, subjects are held individually in standard cages, the objects and arenas were cleaned using PDI Sani-Cloth® HB Germicidal Disposable Wipes (Orangeburg, NY, USA), and the bedding was replaced.
  5. Exposures 1-4 were habituation sessions with two objects placed symmetrically on either end of the arena approximately 5 cm away from the wall. In exposure 5, one of the objects was replaced with a novel object. All subjects were returned to their home cages at the end of the 5th exposure.

NOR + Displacement Protocol

  1. Same as above with the exception of 2 additional runs. Subjects are given a 6th run of the two objects.
  2. The 7th run consists of the constant object with the most recent novel object being displaced.

Pre-Exposure Protocol

  1. Subjects are pre-exposed to the NOR arena for a 5-min, given a 3-min inter-exposure interval, and then given a 5-min exposure of the arena plus objects.
  2. A 3-min inter-exposure interval followed and a novel object replaced a constant object in the 3rd exposure.

NOR Protocol (C57Bl/6J subjects)

  1. For NOR experiments utilizing C57Bl/6J subjects, the testing arena was a white, plastic transport box (55 x 40 x 15 cm) but was not divided into two equal halves as previously done with the 129SvEv subjects. The arena size was increased because C57Bl/6J frequently escaped from the smaller arenas used with the 129SvEv subjects.
  2. The other option for arena size is to use the large plastic boxes with very high walls.

Stopwatch+ Scoring

  1. Behavior was scored on videos by an observer blind to treatment or to genotype using the Stopwatch+ program (Center for Behavioral Neuroscience, Atlanta, GA, USA).
  2. Each test session was scored continuously in its entirety. As a proxy for locomotor activity, a grid of 6 boxes was superimposed over the arena, and grid crossings were counted.
  3. Object investigation was defined as orientation of the head toward the object with the nose within 1 cm of the object. Investigation was not scored if the subject was on top of the object or completely immobile.
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