The Effect of Cognitively Demanding Lock Box Training on the Well-Being and Hippocampal Neurogenesis of Laboratory Mice

Refinement research aims to improve the physical and psychological well-being of laboratory animals, promoted by the possibility of interactions with their environment. Freely selectable access to environmental enrichment can be criticized for negatively influencing the standardization of studies. In a previous study researchers found that the phenotype of mice does not change significantly within differing usage of environmental enrichments (1). Building on these findings, in this study we histologically examined brain sections of the dentate gyrus from the same group of animals, to investigate the positive effect of enrichment-circumstances on the complexity of neuroanatomic brain structure. The dentate gyrus is part of the hippocampus, which can be seen as a gateway to the brain, forming learning and memory processes as a part of the limbic system, where emotions are processed. This study assumes that a higher complexity of neuroanatomic structures in the dentate gyrus can be understood as evidence for well-being. Materials and Methods: A total of 36 mice from three differently enriched housing systems (i.e., standard housing, super environmental enrichment, and lockbox enrichment) are analyzed. Lockboxes are cognitively and sensomotorically demanding riddles baited with palatable food rewards. In the standard housing group and lockbox group, nesting material, a tube, and a house were provided. Additionally, super enriched cages contained a mouse igloo with a running plate, further nesting-materials, a tunnel attached to the cage lid, and a second cage-level, but does not contain a lock box. The lockboxes were only presented to the mice of the lockbox enrichment group. The employment of timed injections of the proliferation markers CldU and IdU, in conjunction with subsequent histological staining, facilitates the quantification and phenotyping of newborn neuronal cells at various temporal points. Concurrently, this methodological approach enables the differentiated analysis of the formation and survival of newly generated neuronal cells. The dendritic complexity of neurons is analysed using morphometric analysis of neurons stained with the Golgi-Cox method. Results and Conclusion: Well-being is more extensive than the absence of suffering, additionally ensuing protection claims for animals focusing on their cognitive and intelligent behavior. The aim of this work is to provide housing conditions for laboratory mice that go beyond the legal minimum, while maintaining the ability to compare research results.