Long-chain n-3 PUFA from fish have been associated with lower risk of CVD. Fish may also contain methylmercury, which may attenuate the inverse associations of the long-chain n-3 PUFA. However, the mechanisms underlying these associations are not fully known. We evaluated the associations of the serum long-chain n-3 PUFA (EPA, DPA and DHA) and hair Hg with resting heart rate (HR), peak HR during cycle ergometer exercise and HR recovery after exercise. A total of 1008 men from the population-based Kuopio Ischaemic Heart Disease Risk Factor Study, aged 42–60 years and free of CVD, were studied. After multivariate-adjustments in ANCOVA, higher serum total long-chain n-3 PUFA concentration was associated with lower resting HR (extreme-quartile difference 2·2 beats/min; 95 % CI 0·2, 4·1, P
trend across quartiles=0·02), but not with peak HR or HR recovery. Associations were generally similar when EPA, DPA and DHA were evaluated individually, except for DPA, which was also associated with better HR recovery after exercise (extreme-quartile difference 2·1 beats/min; 95 % CI 0·1, 4·2, P
trend=0·06). Higher hair Hg content had a trend towards lower peak HR after adjusting for the long-chain n-3 PUFA (P
trend=0·05), but it only slightly attenuated the associations of the serum long-chain n-3 PUFA with HR. These findings suggest that higher serum long-chain n-3 PUFA concentrations are associated with lower resting HR in middle-aged men from Eastern Finland, which may partially explain the potential cardioprotective effect of fish intake.