Studies have indicated that adult patients with mitral valve prolapse (MVP) may have autonomic dysfunction. The purpose of this study was to evaluate heart rate variability (HRV) in children with MVP. Sixty-seven children with MVP (ages 6 to 18 years; 30 boys and 37 girls) were consecutively studied and subdivided into those with or without symptoms. Thirty-seven normal age-matched children (17 boys and 20 girls) were studied as controls. The patients were further divided into 4 age subgroups. HRV was measured using a 24-hour Holter electrocardiogram (ECG) system (Laser SXP Holter Analysis System, Marquette Electronics, Milwaukee, WI). The ECGs were analyzed in both time domain and frequency domain (spectral analysis). Symptomatic and asymptomatic children with MVP were combined for analysis because they were not significantly different in terms of their HRV. All time-domain indices, with the exception of SDANN (SD of the mean of RR intervals in all 5-minute segments of the 24-hour ECG), were significantly lower in children with MVP than in controls. Children with MVP showed significantly lower spectral power of the high frequency (HF) and low frequency (LF) components when compared with controls. The ratio of LF to HF (LF/HF) was significantly higher in children with MVP. Similar differences were observed in the 4 age subgroups. Sex differences in HRV were observed when girls had lower HRV compared with boys. Lower time-domain and frequency-domain indices of HRV in children with MVP are suggestive of decreased parasympathetic activity and a shift in sympathovagal balance. Various factors including differences in heart rate may contribute to sex differences in HRV.