Abstract Temperament, as indicated by Cloninger's psychobiological model predicts coronary heart disease risk, but its association with autonomic cardiac regulation, a potential mediating mechanism, is unclear. We examined the associations between temperament traits and autonomic cardiac regulation in a resting situation in 798 women and 580 men derived from a population-based sample. After adjustment for age and sex, harm avoidance was associated with lower level of high-frequency (HF) variation, root mean square successive differences (RMSSDs), the percentage of successive R–R intervals >50 ms (pNN50) and higher heart rate (HR) (all p ≤ 0.005), suggesting that harm avoidance is related to low parasympathetic activity. Additional adjustments for behavioral factors attenuated these associations more than the adjustment for biological risk factors. Novelty seeking was associated with higher RMSSD ( p = 0.007) and pNN50 ( p = 0.012) and lower heart rate ( p < 0.001). With adjustment for behavioral risk factors, the associations with RMSSD ( p = 0.136) and pNN50 ( p = 0.236) attenuated to the null, but adjustment for biological risk factors had little effect. Reward dependence and persistence were unrelated to indices of cardiac regulation.