Autonomic nervous system (ANS) dysfunction plays a role in atrial fibrillation (AF) initiation. Cardiorespiratory fitness modulates ANS function and is inversely associated with resting heart rate (RHR) and risk of AF. Thus, we sought to study the association between RHR, as a surrogate for ANS function, and incident AF independent of exercise capacity (EC). We analyzed 51,436 subjects without previous AF who underwent a clinically indicated exercise stress test. Incident AF was ascertained through claims files. RHR was measured before stress testing, and EC was estimated by peak metabolic equivalents of task. We studied the association between RHR categories (<70, 70 to 85 [reference], and >85 beats/min) and incident AF using Cox models adjusted for risk factors and for EC. We tested for interaction between RHR and age, gender, smoking, and EC. Mean (SD) age was 53 (13) years, 53% were men, and 28% were black. Participants with RHR <70 beats/min were older, more likely to be men, have higher EC, and more likely to smoke but less likely to have diabetes and hypertension. Over a median of 5.5 years, RHR <70 beats/min was associated with 14% increased risk of AF (95 CI 6% to 25%) in fully adjusted models, whereas RHR >85 beats/min was not associated with AF risk after adjusting for EC. Results for RHR analyzed continuously and by quartile were similar. No interaction was seen. In conclusion, subjects with low RHR at all levels of EC are at increased risk of AF and may benefit from heart rhythm surveillance, particularly in the presence of other AF risk factors.