Gender and age are both known to affect heart rate variability (HRV). Their interaction is not known. HRV, determined from 24-hour Holter recordings, was compared between gender-matched older (15 men and 15 women, aged 67 ± 3 years, range 64 to 76) and younger (15 men and 15 women, aged 33 ± 4 years, range 26 to 42) subjects selected for an age difference of approximately 35 years. HRV for older and younger subjects was compared separately by gender. HRV was also compared by gender within groups. Heart rates were significantly higher, and all time and frequency domain indexes of HRV were significantly lower among the older than among the younger men. Among the women only the shorter term indexes of HRV were significantly lower in the older group. When HRV was compared by gender within age groups, there were no significant differences between men and women in the older group. In the younger group, men had lower heart rates, and all 24-hour time domain indexes of HRV, except those that reflect vagal modulation of heart rate, were significantly higher than those in women. We conclude that HRV is comparable in older men and women. However, HRV is differently affected by age. In men, for whom initial levels of HRV are significantly higher, older age is associated with a global reduction in HRV, reflecting reductions in both sympathetic and parasympathetic modulation and a loss of circadian variability. In women, older age is associated mainly with a decline in shorter term indexes of HRV without significant changes in circadian variability.