To behaviorally characterize rodents and also to assess drug effects in these animals, tests of spontaneous unconditioned behaviors are often used. In these tests locomotor behavior plays a crucial role as it seems to be difficult for instance to reliably dissociate locomotion from indices of anxiety or exploration. To differentially investigate behavioral dimensions in rodents, behavioral data obtained from a series of different independent tests have to be correlated, resulting in cost-intensive and time-consuming procedures. Moreover, it has to be considered that an animal's behavior is context-specific and, thus, independent behavioral dimensions might be produced by different test paradigms themselves. Behavioral tests which allow to investigate a wide range of behaviors and which are focussed on a more detailed ethological analysis of experimental animals in a single complex paradigm may overcome these disadvantages.

We designed a modified hole board (mHB) paradigm which enables the investigator to perform more complex ethological observations than other tests of unconditioned behavior. The mHB comprises the characteristics of the classical hole board, which is especially used to evaluate exploration and activity, and the open field, which is used to investigate anxiety and locomotor activity. Additionally, by allowing visual and olfactory contact among the animals to be tested, the design of the mHB circumvents the stressful factor of social isolation during the test and enables the assessment of the social affinity among group mates. Therefore, this new test paradigm enables to investigate a wide range of behaviors, including anxiety-related behavior, risk assessment behavior, exploration, locomotor activity, and social affinity.

Wistar rats that have been selectively bred over the past decade for high anxiety-related behavior (HABs) show an increased innate emotionality and more passive stress coping strategies than the line of low anxiety-related behavior rats (LABs). Thus, HABs represent a unique psychopathological animal model of innate anxiety with face and predictive validity. In the present study the mHB behavior of untreated HABs and LABs revealed clear differences in anxiety-related behavior. Furthermore, HABs compared with LABs showed no differences in exploration of the protected area (i.e. rearings in the box) or social affinity. The same behavioral characteristics were found in vehicle-treated animals in two pharmacological experiments.

These results demonstrate that, using the mHB test, it is possible to differentially analyze the basal behavior in rodents and to evaluate behavioral effects induced by different pharmacological treatments under stress-reduced conditions. Behaviors such as anxiety, exploration, and locomotor activity can be dissociated and investigated by thorough monitoring and analysis of behavioral parameters obtained in this behavioral test. Hence, the mHb represents an alternative to the common practice, namely performing a series of more specific tests. Finally, due to its stress-reduced characteristic by avoiding social separation, the mHB test enables the investigator to evaluate subtle behavioral modulations such as mild changes in anxiety-related behavior or changes in the exploration of unprotected areas. Our results confirm that the mHB represents an effective tool for behavioral characterization in rodents and for high throughput-screenings for potential therapeutic agents in preclinical research.

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