BACKGROUND: Premature infants are vulnerable to destructive brain injury and disturbed neurological development. Prematurity may alter maturation of the central autonomic nervous system (ANS). AIMS: To compare ANS function (using heart rate variability; HRV) between preterm infants with normal neuroimaging at term equivalent age and low-risk term controls. Study design, subjects. We performed a case-control study of preterm infants born </=28鈥痺eeks gestational age that had normal brain imaging and archived continuous EKG data at term equivalent age. We documented other factors thought to influence ANS maturation (e.g. infection, ventilation days, and postnatal steroids). Controls were low-risk term gestational age newborns from uncomplicated pregnancies/deliveries. We characterized HRV metrics using frequency-(Welch periodogram) and time-domain (detrended fluctuation) analyses. Sympathetic tone was characterized by alpha(1), root mean square analysis (RMS(1) and RMS(2)), low-frequency (LF) power, and normalized LF (nLF) and parasympathetic tone was characterized by high-frequency (HF) power and normalized HF (nHF). alpha(2) characterized ultraslow changes in heart rate. We used ANCOVA to compare HRV metrics between groups. Outcome measures, results. HRV from 26 preterm infants were compared to 55 controls. Analyzed HRV data for preterm infants were recorded at median (range) gestational age of 39 (36-39) weeks and for controls at 39 (37-41) weeks gestational age. alpha(1), RMS(2), LF and HF were significantly higher in control infants and remained significant after controlling for infection, ventilator days, and postnatal steroids (P鈥?鈥?005). CONCLUSIONS: Autonomic maturation is impaired in a premature extrauterine environment. In the absence of destructive brain injury, our data suggest an important role for disturbed programming in this impaired autonomic development.