Eco-HAB: New quality in research on neuronal basis of social behavior

Therapy is really effective only when it is based on the full understanding of the mechanisms responsible for the occurrence of a disease. When it comes to neuronal background of impairments of social behavior, characteristic for disorders such as autism or social phobia, behavioral tests performed in rodents are an important source of knowledge. However, these tests have a serious flaw: scientists are often unable to reproduce data obtained in other laboratories. This notorious issue has just been resolved. A group of Prof. Ewelina Knapska from Nencki Institute of Experimental Biology in Warsaw, Poland, introduced the Eco-HAB, the first automated system for assessment of social behavior in mice. It ensures results reproducibility and the evaluation of components of sociability that are crucial for coping with day-to-day functioning in a given social group. This high-tech device mimics most essential characteristics of natural murine environment and automatically traces complex social interactions among animals. The Eco-HAB, along with open-source software for data analysis and precise description enabling its manufacturing by any interested party, has recently been described in scientific journal eLIFE.

"One may buy many automated systems for behavioral assessment in mice. However, none of them enables testing of social behavior impairments in a manner asserting data reproducibility. The situation has been rather dire: often experiments could not have been replicated not only in other labs, but even in the very lab where they were performed in the first place," says Prof. Knapska.

To perform conventional behavioral tests of social behavior, a mouse must be taken out of its housing cage and put into the testing environment. Such testing arenas are designed to predispose an animal to a particular behavior. The most popular apparatus is a rectangular box divided into three parts: two side-chambers connected by a center chamber. In one of the side-chambers a social stimulus (e.g. an unknown mouse, trapped under a wire-cup) is presented. The other side-chamber contains a non-social stimulus (e.g. a toy brick, also under the cup). The tested animal is allowed to move freely and decide whether to explore the social or the non-social stimulus.

Such testing arrangement evokes a significant degree of stress in subjects, thus confounding the obtained results. It causes animals to behave in an unpredictable way. Additionally, social isolation from cagemates, enforced by the experimental design, seems to be especially stressful for the subjects. Further, even the type of presented social stimulus may affect the behavioral responses. Come to think of it -- it should not be expected that a stressed mouse would be interested in interacting with another agitated conspecific.

Problems are further exacerbated by the lack of unified experimental standards. The same behavioral assay may differ in terms of execution between laboratories. Moreover, there are no commonly agreed-upon norms concerning such behavioral evaluation. Variables include the sex of the experimenters, light and temperature conditions, and most importantly, there is no consistency when it comes to data analysis.

"Lack of standardization of behavioral testing is a serious problem. It would be very hard to even catalog all the factors that might potentially confound measurements of social interactions. That is why we designed our system to reduce experimental stress in subjects to absolute minimum. Besides asserting group-housing of animals, the Eco-HAB mimics most of the important features of natural murine habitats as faithfully as it is possible in the experimental set-up. Such environment elicits innate behaviors and allows for measuring of spontaneous social interactions. Another bonus is that Eco-HAB processes data automatically, which is very useful from the experimenters' point of view," says Dr. Alicja Puścian, who designed the Eco-HAB in the course of her PhD program.

Mice are very social animals, but at the same time they are very territorial. In the wild they regularly patrol their habitats, often traveling as far as kilometers from its center. In order to allow for spontaneous social interactions and unhampered activity, the Eco-HAB was equipped with elements resembling burrows connected by underground corridors. In the course of a particular experiment the system is capable of tracking the behavior of dozen or so animals. This allows for observation of complex social interactions between particular conspecifics.

Furthermore, Eco-HAB enables assessment of interest in novel social stimuli. Lack of drive to interact with new people is one of the core symptoms of autism, so it is crucial to be able to assess such behaviors in mouse models. However, rodents show their interest in social novelty differently than humans.

"Olfaction is the dominant sense in rodents. That is why we use scent of an unknown conspecific rather than its presence as a social stimulus. In the wild, unfamiliar mice, especially males, tend to avoid each other, because every such encounter carries a risk of aggression. Still, they are always interested in a scent of another mouse. Just like dogs, mice get acquainted through scent marking," explains Dr. Puścian.

To track the mice in the Eco-HAB environment the scientists used RFID (radio-frequency identification) antennas, which register the passing of microchips located under the skin of each mouse (similar chips are used by veterinarians to protect our pets from getting lost). Based on that information, the analyzing software, written in Python, not only records the activity, but also runs appropriate analysis of the social behavior. Customized behavioral parameters have been chosen to guarantee reproducibility of the obtained results.

The Eco-HAB enables non-invasive and reliable analysis of many aspects of murine sociability. It can be used not only to test whether mice are interested in a given social stimulus, but also to establish if particular pairs of animals follow or avoid each other. To standardize the system, group of Prof. Knapska tested it in different laboratories and using various mouse models presenting diverse social behaviors.

"In our article we provide full documentation of the Eco-HAB, dimensions of all elements, list of utilized materials, electronic schemes, and the source code of the software. In that way, anyone interested in building the system in their laboratory should be able to customize the software to fit their needs. They can run their experiments, assured that the results can be verified by other labs," says Prof. Knapska and emphasizes another important asset of the Eco-HAB -- its price. The complete cost of construction is no higher than 2000 euro, thus making the apparatus more than affordable in comparison with other equipment designed for behavioral research in rodents.

Group of scientists lead by Prof. Knapska together with the team of Dr. Grzegorz Kasprowicz from Faculty of Electronics and Information Technology of Warsaw University of Technology, supported by National Science Center grant SYMFONIA and Polish-Swiss Program, have been working on more technologically advanced version of Eco-HAB, which has already been patented.

The Nencki Institute of Experimental Biology of the Polish Academy of Sciences has been established in 1918 and is the largest non-university centre for biological research in Poland. Priority fields for the Institute include neurobiology, neurophysiology, cellular biology and biochemistry and molecular biology -- at the level of complexity from tissue organisms through cellular organelles to proteins and genes. There are 31 labs at the Institute, among them modern Laboratory of Confocal Microscopy, Laboratory of Cytometry, Laboratory of Electron Microscopy, Behavioural and Electrophysiological Tests. The Institute is equipped with state-of-the-art research equipment and modernized animal house, where lab animals are bred, also transgenic animals, in accordance with the highest standards. Quality of experiments, publications and close ties with the international science community, place the Institute among the leading biological research centres in Europe.