A sustained first inflation (SI) at birth may aid lung liquid clearance and aeration, but the impact of SI duration relative to the volume-response of the lung is poorly understood. We compared three SI strategies: 1) variable duration defined by attaining volume equilibrium using real-time electrical impedance tomography (EIT; SI plat); 2) 30 s beyond equilibrium (SI long); 3) short 30-s SI (SI30); and 4) positive pressure ventilation without SI (no-SI) on spatiotemporal aeration and ventilation (EIT), gas exchange, lung mechanics, and regional early markers of injury in preterm lambs. Fifty-nine fetal-instrumented lambs were ventilated for 60 min after applying the allocated first inflation strategy. At study completion molecular and histological markers of lung injury were analyzed. The time to SI volume equilibrium, and resultant volume, were highly variable; mean (SD) 55 (34) s, coefficient of variability 59%. SI plat and SIlong resulted in better lung mechanics, gas exchange and lower ventilator settings than both no-SI and SI30. At 60 min, alveolar-arterial difference in oxygen was a mean (95% confidence interval) 130 (13, 249) higher in SI30 vs. SIlong group (two-way ANOVA). These differences were due to better spatiotemporal aeration and tidal ventilation, although all groups showed redistribution of aeration towards the nondependent lung by 60 min. Histological lung injury scores mirrored spatiotemporal change in aeration and were greatest in SI 30 group (P Ͻ 0.01, Kruskal-Wallis test). An individualized volume-response approach to SI was effective in optimizing aeration, homogeneous tidal ventilation, and respiratory outcomes, while an inadequate SI duration had no benefit over positive pressure ventilation alone. sustained inflation; neonatal resuscitation; lung mechanics; lung volume; variability; electrical impedance tomography; lung injury THE MAJORITY OF EXTREMELY preterm infants require respiratory assistance in the delivery room (41). In part this is because many of these infants do not have the ability to generate the initial prolonged high transpulmonary pressures required to drive lung fluid from the main airways, allow alveolar aeration, establish functional residual capacity (FRC), and then maintain it during tidal ventilation, essential processes for efficient gas exchange and lung protection (19,31). Recently, applying an initial sustained inflation (SI) at birth, consisting of an elevated pressure applied for longer than needed for usual tidal inflation, followed by sufficient positive end-expiratory pressure (PEEP), has been proposed as a method of generating the initial transpulmonary pressure needed at birth (10,18,20). SI has been extensively investigated in preterm animals (15,26,29,32,33,(35)(36)(37)(38) and humans (10, 18, 34) with conflicting results. Some studies suggested SI improved aeration, FRC, and cerebral oxygen delivery (29,32,33), while others failed to demonstrate any benefit over standard respiratory support with sufficient PEEP (26,(35)(36)(37). SI was associated wi...