This study aimed at examining the associations between weight status, body fat mass, and heart rate variability in prepubescent children, adjusting for physical activity levels. A cross-sectional investigation in which a total of 50 Caucasian pre-pubertal children (21 normal weight; 8 overweight; 21 obese), aged 6-10years (8.33±1.14), including both boys (n=24) and girls (n=26), were recruited from local schools. Total body fat and trunk fat were evaluated through dual-energy X-ray absorptiometry. Free-living physical activity levels were evaluated by accelerometer. Short-term heart rate variability acquisition was performed; time- and frequency-domain parameters were analysed. Logarithmic transformations of the low-frequency (LnLFnu), high-frequency (LnHFnu) normalized units and low-frequency/high-frequency (LnLFnu/HFnu) ratio were computed. Adjusting for age, Tanner stage, and moderate to vigorous physical activity levels, obese children compared to normal weight children showed a significant decreased LnHfnu (3.8±0.2 vs 4.1±0.2%) and both higher LnLFnu (4.0±0.4 vs 3.7±0.3%) and LnLFnu/LnHFnu ratio (1.1±0.1 vs 0.9±0.1). LnHFnu showed significant negative correlation with waist circumference (r=-0.598; P=0.000), total body fat (r=-0.409; P=0.011) and trunk fat (r=-0.472; P=0.003). Both LnLFnu and LnLFnu/LnHFnu ratio showed positive correlations with waist circumference (r=0.455; r=0.513) and trunk fat (r=0.370; r=0.415). A higher amount of body fat mass, particularly central fat, was shown to be related to decreased parasympathetic modulation in time-domain heart rate variability. This finding highlights the potential cardiovascular risk that excessive fat mass may represent even at very young age.