Molenhuis1 project protocol

Social interaction and locomotor activity, digging, and grooming in males of 52 Collaborative Cross strains of mice   (2018)

Molenhuis RT, Iraqi FA, Mott RF, Kas MJ
With: Bruining H, Brandt MJV, van Soldt PE, Abu-Toamih Atamni HJ, Burbach JPH




Project protocol - Contents

Workflow and sampling

Step Description Equipment Data collected
1 Social interaction and discrimination capacity determined Monitoring system Time exploring unfamiliar conspecific mouse, preference for novel versus familiar conspecific mouse
2 Locomotor activity and exploratory behaviors assessed with four novel objects Monitoring system Distance traveled, time digging, time grooming
3 Mice weighed and euthanized; brains harvested and weighed Balance Body weight, brain weight

Procedures

Procedure 1: Social interaction and discrimination capacity

Equipment, software, and supplies

  • Clean and transparent mouse cage, with wood chip bedding
  • The Observer XT (Noldus Information Technology, Wageningen, The Netherlands)

Environmental Conditions

Testing Area

Acclimation Period: Test animals are allowed to habituate for 5 min.

Steps

  1. On day 1 the mice are exposed to an unfamiliar age- and gender-matched conspecific animal (A/J) for 2 min (T0).
  2. An inter-trial interval (ITI) of 5 min is carried out, after which the mice are re-introduced to both the familiar conspecific animal, and a first novel conspecific animal for 2 min (short-term memory, T1).
  3. On day 2, 24 h after the ITI, mice are re-exposed to the same familiar intruder of day 1, and to a different novel intruder from a different cage and housing room than the intruder of day 1. Test mice are exposed for 2 min (long-term memory test, T24). The time spent exploring the stimulus animals is recorded.
  4. Discrimination capacity is calculated as the preference for the novel over the familiar animal using the formula: tN/(tN + tF). Where tN is the time spent exploring the novel animal and tF is the time spent exploring the familiar animal. A ratio value above 0.5 indicates successful discrimination between novel and familiar animal.

Procedure 2: Locomotor activity and exploratory behavior

Equipment, software, and supplies

  • Arena of 26.5 x 17 x 11.4 cm, with wood chip bedding and four novel objects
  • The Observer XT (Noldus Information Technology, Wageningen, The Netherlands)
  • Overhead camera with video tracking software (EthoVision XT) (Noldus Information Technology, Wageningen, The Netherlands)
  • Theme software version 5 (Noldus Information Technology, Wageningen, The Netherlands)

Steps

  1. On day 3, locomotor trajectories are recorded in the presence of four novel objects, which are used to determine the total distance covered per animal in a 10 min test. The objects used are made of glass, metal or plastic and are of sufficient height to prevent mice from climbing on to the objects (from Molenhuis et al. 2014).
  2. During the same 10 min test, time spent digging and time spent grooming are manually recorded.

Procedure 3: Body weight and brain weight

Equipment, software, and supplies

  • Balance

Steps

  1. On day 4, mice are weighed and then euthanized, after which the brains are harvested and weighed.

References

Primary References

Molenhuis RT, Bruining H, Brandt MJV, van Soldt PE, Abu-Toamih Atamni HJ, Burbach JPH, Iraqi FA, Mott RF, Kas MJH. Modeling the quantitative nature of neurodevelopmental disorders using Collaborative Cross mice. Mol Autism. 2018 Dec 13;9:63. doi: 10.1186/s13229-018-0252-2. eCollection 2018.   PubMed 30559955     FullText

Protocol References

Molenhuis RT, de Visser L, Bruining H, Kas MJ. Enhancing the value of psychiatric mouse models; differential expression of developmental behavioral and cognitive profiles in four inbred strains of mice. Eur Neuropsychopharmacol. 2014 Jun;24(6):945-54. doi: 10.1016/j.euroneuro.2014.01.013. Epub 2014 Jan 18.   PubMed 24491952