This study investigated factors contributing to differences between mean alveolar pressure (PA) and mean pressure at the airway opening (Pao) during high-frequency oscillatory ventilation (HFOV). The effect of the inspiratory-to-expiratory time (I/E) ratio and amplitude of oscillation on the magnitude of - Pao (Pdiff) was examined by using the alveolar capsule technique in normal rabbit lungs (n = 4) and an in vitro lung model. The effect of ventilator frequency and endotracheal tube (ETT) diameter on Pdiff was further examined in the in vitro lung model at an I/E ratio of 1:2. In both lung models, fell below Pao during HFOV when inspiratory time was shorter than expiratory time. Under these conditions, differences between inspiratory and expiratory flows, combined with the nonlinear relationship between resistive pressure drop and flow in the ETT, are the principal determinants of Pdiff. In our experiments, the magnitude of Pdiff at each combination of I/E, frequency, lung compliance, and ETT resistance could be predicted from the difference between the mean squared inspiratory and expiratory velocities in the ETT. These observations provide an explanation for the measured differences in mean pressure between the airway opening and the alveoli during HFOV and will assist in the development of optimal strategies for the clinical application of this technique.