Background: Neurally adjusted ventilatory assist (NAVA) is a novel mode of ventilation that has been demonstrated to improve infant-ventilator interaction, compared to the conventional modes in retrospective and short-term studies. Objectives: To prospectively evaluate the physiologic effects of NAVA in comparison with pressure-regulated volume control (PRVC) in two nonrandomized 12-hour periods. Methods: We studied 14 consecutive intubated preterm neonates receiving mechanical ventilation for acute respiratory failure. Peak airway pressure (Pawpeak), diaphragm electrical activity (EAdi), tidal volume (VT), mechanical (RRmec) and neural (RRneu) respiratory rates, neural apneas, and the capillary arterialized blood gases were measured. The RRmec-to-RRneu ratio (MNR) and the asynchrony index were also calculated. The amount of fentanyl administered was recorded. Results: Pawpeak and VT were greater in PRVC (p < 0.01). Blood gases and RRmec were not different between modes, while RRneu and the EAdi swings were greater in NAVA (p = 0.02 and p < 0.001, respectively). MNR and the asynchrony index were remarkably lower in NAVA than in PRVC (p = 0.03 and p < 0.001, respectively). 1,841 neural apneas were observed during PRVC, with none in NAVA. Less fentanyl was administered during NAVA, as opposed to PRVC (p < 0.01). Conclusions: In acutely ill preterm neonates, NAVA can be safely and efficiently applied for 12 consecutive hours. Compared to PRVC, NAVA is well tolerated with fewer sedatives.