Reliability of repeated exposure to the human elevated plus-maze in virtual reality: Behavioral, emotional, and autonomic responses.

Approach-avoidance conflicts are a hallmark of anxiety-related behaviors. A gold standard for assessing anxiety-related behaviors in rodents is the elevated plus-maze (EPM), which was recently translated to humans using immersive virtual reality. Repeated behavioral testing is particularly interesting for clinical and pharmacological research in humans but could be limited by habituation effects. Here, we tested whether comparable strategies that are used in rodents (different environments and inter-trial interval of 28 days) are sufficient to avoid habituation or sensitization effects on the EPM, making it possible to perform repeated measurement of anxiety-related behavior in humans. Moreover, we developed two novel virtual environments for repeated testing to explore whether a scenario resembling the real world is superior to a video game-like EPM in terms of lifelike physiological, emotional, and behavioral responses. On a behavioral level, no significant differences but a high correlation between first and repeated exposure to the human EPM independent of EPM version were found. On a psychophysiological level, salivary alpha-amylase, skin-conductance, and respiratory frequency increased at first and second exposure independent of EPM version. However, at repeated exposure, skin-conductance and heart rate showed indicators for anticipatory anxiety and a small sensitization effect, while no effect of real-world resemblance on these physiological measures was found. This was also reflected in slightly higher subjective anxiety levels at second exposure, although subjective anxiety still correlated strongly between first and second exposure. In conclusion, the human EPM can be used for longitudinal assessments of human anxiety-related behavior when strategies to avoid habituation and sensitization are considered.