Heart rate asymmetry (HRA) is an index that accounts for an uneven contribution of decelerations and accelerations to the heart rate variability (HRV). Clinical studies indicated that HRA measures have additive clinical value over the more frequently used HRV indexes. Despite the abundance of studies on psychological influences on HRV, little is known whether psychological factors influence HRA. Based on previous research regarding HRA and stress, we expected that negative emotions compared to positive emotions would decrease the contribution of decelerations to HRV. Thirty female participants watched three clips that produced negative emotions, positive emotions, and neutral affect. Besides electrocardiogram, we measured several physiological and behavioral responses to ascertain the affective impact of the clips. Using the RR interval time-series, we calculated HRV and HRA indexes. We found that HRA differentiated between positive emotions and negative emotions reactivity. Positive emotions produced a higher number of decelerations in short-term variability to the total short-term variability (C1d) compared to negative emotions. Moreover, C1d correlated with subjective ratings of affect. In sum, the results of this study indicated that HRA is sensitive to psychological influences. HRA indexes are likely to contribute to a more nuanced physiological differentiation between emotions.