Constant monitoring of behavioral patterns shapes successful social interactions. Factors such as interpersonal distance and emotional expressions prompt specific responses to facilitate functional social exchanges. Despite extensive research on these topics, studying how these features influence human physiological responses in an ecological yet controlled environment has often proven challenging. Moreover, the relation between a possible dysregulation of these physiological reactions and social anxiety remains unclear. Using a 3D virtual setting, we found increased physiological responses (electrodermal activity, EDA) during emotionally salient scenarios when compared to neutral ones. Remarkable differences in subjects’ EDA resulted also from interactions with displays of strong emotions, namely anger and fear. Angry virtual agents elicited higher physiological activity, whereas displays of fear showed lower levels of EDA. Additionally, interpersonal distance in given emotional contexts had different impacts on EDA responses. In neutral scenarios, the invasion of personal space provoked an increase in arousal, which anger amplified, and fear reduced. In a following study, we found that participants with high social anxiety displayed similar levels of EDA across all scenarios. Also, these subjects registered stronger autonomic responses against counterparts with low social anxiety. Our findings show how the interaction of different variables in complex social contexts modulates our physiological reactions. Furthermore, they also indicate that social anxiety induces greater arousal, which hinders situational discernment of different types of interactions. In conclusion, these outcomes lay the groundwork for developing innovative training programs to rehabilitate and enhance social skills.