Acclimation and Blood Sampling: Effects on Stress Markers in C57Bl/6J Mice

Abstract

Blood sampling in rodents is common practice in scientific studies. Some of the refined methods widely used are the puncture of the saphenous vein or tail vein, or even tail docking. The handling needs of these different blood sampling methods are different and can directly affect stress, increasing the variability of the study. Moreover, there is less aversion and stress if the animal is accustomed to the environment, handling and technique. Therefore, our study aimed to assess the influence of these three blood sampling techniques (saphenous puncture, tail vein puncture and tail vein docking) and the use of previous acclimation on different indicators of animal stress, assessing blood glucose concentrations and faecal corticosterone metabolites (FCMs). Twenty-four young adult male and female C57Bl6/J mice were divided in three groups by sampling method: tail docking (TD), saphenous vein puncture (SV) and caudal vein puncture (CV) groups. All mice were studied with and without acclimation, which was performed during 9 consecutive days. The results showed that both males and females present very similar responses to the different handling and sampling methods without significant differences. Nevertheless, acclimation in all sampling methods decreased glucose and FCM levels significantly. The method that obtained the lowest glucose and FCM levels with significance was saphenous vein puncture. Therefore, we can say that it causes less stress when performing prior acclimation, even when this involves greater handling of the animal. Our results contribute to refinement within the 3R concept and could serve researchers to programme and select a good handling technique and a welfare-friendly blood sampling method for their experiments.

Keywords: 3R; acclimation; animal welfare; blood sampling; glucose refinement; mice; stress.

Figure 1

Experimental design timetable. Blood sampling was performed on day 1 and 9. Faecal sampling was performed on day 2 and 10.

Figure 2

Differences between glucose levels pre and postacclimation between sexes. Results showed no differences by sex in all methods used for sampling (TC group p = 0.2; TV group p = 0.69; SV group p = 0.88).

Figure 3

Glucose levels (mg/dL) by technique before (pre-) and after (post-) acclimation.

Figure 4

Differences of glucose levels between methods, without acclimation. Results show differences were statistically significant between the SV and TC groups (p = 0.083), and between the SV and TV groups (p = 0.046).

Figure 5

Differences of glucose levels between methods, post-acclimation period. Results show differences were statistically significant between the SV and TC groups (p = 0.00094) and between the SV and TV groups (p = 0.046).

Figure 6

Differences in FCMs (pg/mL) comparing by methods (p = 0.44).

Figure 7

Differences in FCMs (pg/mL) comparing groups with (mean: 0.6052 pg/mL) and without acclimation (mean: 0.9772 pg/mL).