Both personality traits and autonomic functioning show as gender differences, but their relationship is not well understood. Medically unexplained symptoms are related to personality features and can be assessed by autonomic measurement. The patterns are hypothesised to identify gender differences. We recruited 30 male and 30 female healthy volunteers. All participants completed the Tridimensional Personality Questionnaire (TPQ) and heart-rate variability (HRV) measurement. Correlation analysis was performed to identify the relationships between TPQ scores and HRV parameters. For the subjects as a whole, the subdimension harm avoidance 4 (HA4, fatigability and asthenia) was found to be negatively correlated with low-frequency (LF) power, high-frequency (HF) power and total power (TP) of HRV. Novelty seeking 1 (NS1, exploratory excitability) was found to be positively correlated with LF power and TP. Multiple linear regression analysis revealed that the interactions exploratory excitability x gender and fatigability x gender are predictors of LF and HF power, respectively. Our result supports the hypothesis that personality features such as exploratory excitability and fatigability are associated with autonomic functioning and that gender is a moderator in these relationships.