Larsen, R. A., Peveler, J. L., Klutzke, J. B. et al. 2017. Effects of daily restraint with and without injections on skeletal properties in C57BL/6NHsd mice. Lab Animal 46(7), 299-301.

Studies typically involve groups of animals that are injected with placebo compounds. As there are studies that demonstrate that restraint and injection can affect behavior and corticosteroid levels in rodents, the basis of such placebo injections is to control for any potential effects caused by handling and injecting the experimental animals. But these stressors may not adversely affect all studies equally. While placebo injections make sense for studies that are focused on outcomes which may be directly or indirectly affected by stress hormones, the value of placebo injections is less clear for other studies. If placebo groups are not necessary for some studies, this would reduce both the number of animals used in research and the need to handle/inject a significant number of animals. We tested 60 female C57BL/6NHsd mice that were randomly assigned to one of three experimental groups: animals that were only handled during weekly cage changes (CON, n=20); animals that were restrained but not injected 5 days per week (SHAM, n=20); and animals that were restrained and given an intraperitoneal (IP) injection (0.15 cc 0.9% saline solution) 5 days per week (INJ, n=20). Our data clearly showed no effect of saline injections or animal handling on skeletal morphology or mechanical properties following an 8-week study period. These results suggest that for these tissue/organ level outcome measures there is no need to use saline-injections in control animals. This represents a refinement in experimental design that can result in a reduction of overall animal use in similar studies as well as in investigator time to do sham injections of large numbers of animals.