Total power and high frequency power heart rate variability (HRV) have been reported to decline at moderate exercise intensities when cycling (Arai et al., 1989). This may be indicative of the withdrawal of vagal input to the sino-atrial node (Arai et al., 1989). Similarly, heart rate (HR) has been shown to increase with cycling cadence, at a constant power output (Gotshall et al., 1996). However, the effect of different cycling cadences on measures of HRV has not previously been examined; this may have important implications for studies measuring autonomic responses during cycling. The aim of the present study was to examine the effect of cadence on HRV during cycling at a constant power output. It was hypothesised that HRV would be similar when cycling at different cadences. Following receipt of ethical approval, sixteen males performed three, ten-minute periods of unloaded cycling, on an electronically braked recumbent cycle ergometer, consisting of: 1) a freely chosen cadence (FCC) whilst blinded to the pedalling rate; 2) cadence of 40 revs.min^-1; 3) cadence of 80 revs.min^-1. All conditions were performed on the same day with FCC first and the other conditions counterbalanced; each test was separated by a recovery period. A three-lead ECG trace was recorded for the calculation of total power and high frequency power using Fast Fourier Transformations. A one-way repeated measures ANOVA, was used to investigate between conditions differences in total power and high frequency power. Significant differences were examined post-hoc by pairwise comparisons. Statistical significance was accepted at P<0.05 with data presented as means (SD). The mean(SD) FCC was 57(9) revs.min^-1.Heart rate was higher at FCC and 80 revs.min^-1 than 40 revs.min^-1 (69[9]; 75[4]; 63[6], beats.min^-1, respectively).Total power declined with each increment in cadence (1769[1311]; 1367[997] and 925[761] ms², for 40 revs.min^-1, FCC and 80 rev.min^-1 respectively). Differences in high frequency power were observed between the FCC and 80 revs.min^-1 conditions (832[111] and 337[432] ms² respectively. It is concluded that total power and high frequency power are reduced as a consequence of increasing cycling cadence, therefore the hypothesis is rejected. This may be indicative of vagal withdrawal resulting in an elevated HR whilst cycling at a FCC and 80 revs.min^-1. Studies examining autonomic function during cycling should consider the confounding effects of alterations in cycling cadence.