Figure 2.

An illustration of the days on which differential handling was done and the daily data collection schedule for the first 4 cages (3 study weeks).Two cages of 4 males and 2 cages of female mice entered the study each week, alternating sex between Mondays and Wednesdays. PreA (prehandling acclimation anxiety testing) was followed by 7 d of tail or tunnel handling of the mice in each cage (Acc1 through 7). Each cage then underwent Post (handling) Acclimation anxiety and baseline pain scoring (PA) before being tattooed or restrained (Tat/Res), and then the postprocedural anxiety and pain assessments (PP) were undertaken. The mice were imaged (I) the same evening, and the PP and imaging data collection was repeated at 24 h. Mice that had undergone restraint were ear tagged 2 wk later, immediately after which the PP recordings were repeated.