The study examined the relationship of individual differences in prefrontal brain asymmetry, measured by the EEG in resting conditions, to the individual's responsivity in the context of humor (n = 42). Several weeks after the EEG recording, immediate cardiovascular responses to the perception of humor and behavioral indicators of humor processing were obtained in an experimental paradigm involving non-verbal cartoons. Relatively greater resting activity in the left than right prefrontal cortex, particularly at the ventrolateral positions, was associated with faster detection of humor, a more pronounced cardiac response to the perception of humor (heart rate and cardiac output), and more accessible internal positive affective states (indicated by faster reports of amusement levels). The study confirms and extends findings of the relevance of prefrontal brain asymmetry to affective responsivity, contributing evidence in the domain of positive affect and humor, and demonstrating relationships to the immediate cardiovascular response pattern to an emotional event.