Interpersonal aspects of teaching have repeatedly been linked to teacher emotions and well-being on a general level. However, it is unclear how teachers’ moment-to-moment interpersonal behavior is associated with their physiological arousal during teaching and how this contributes to their lesson-focused emotional outcomes. Eighty secondary education teachers with a mean age of 43.7 years (SD = 11.5) and 13.4 years of teaching experience (SD = 9.7) participated during one real-life lesson. We coded teacher behavior from an interpersonal perspective on the dimensions of agency (i.e., social influence) and communion (i.e., friendliness). Teachers’ physiology (in terms of heart rate) was measured as a proxy for their affective arousal. Teachers differed widely in their behaviors and in how behavior and physiology were associated from moment to moment. Being generally agentic was associated with higher levels of self-reported positive emotions after the lesson, also when being agentic went together with a high heart rate. In contrast, the stronger and the more positively a teacher’s physiological arousal was associated with displaying communal behavior, the more likely a teacher was to report negative emotions. We conclude that combining moment-to-moment data of teachers’ interpersonal behavior and physiological arousal has the potential to explain differences in teachers’ emotional outcomes. Such an approach might ultimately provide teachers and teacher educators with the fine-grained and personalized information needed to foster teacher well-being.