Abstract Background Mechanical ventilation is applied to unload the respiratory muscles, but knowledge about transpulmonary driving pressure (Δ P L ) is important to minimize lung injury. We propose a method to estimate Δ P L during neurally synchronized assisted ventilation, with a simple intervention of lowering the assist for one breath (“lower assist maneuver”, LAM). Methods In 24 rabbits breathing spontaneously with imposed loads, titrations of increasing assist were performed, with two neurally synchronized modes: neurally adjusted ventilatory assist (NAVA) and neurally triggered pressure support (NPS). Two single LAM breaths (not sequentially, but independently) were performed at each level of assist by acutely setting the assist to zero cm H2O (NPS) or NAVA level 0 cm H2O/uV (NAVA) for one breath. NPS and NAVA titrations were followed by titrations in controlled-modes (volume control, VC and pressure control, PC), under neuro-muscular blockade. Breaths from the NAVA/NPS titrations were matched (for flow and volume) to VC or PC. Throughout all runs, we measured diaphragm electrical activity (Edi) and esophageal pressure ( P ES ). We measured Δ P L during the spontaneous modes ( P L _ P ES ) and controlled mechanical ventilation (CMV) modes ( P L _ CMV ) with the esophageal balloon. From the LAMs, we derived an estimation of Δ P L (“ P L_LAM ”) using a correction factor (ratio of volume during the LAM and volume during assist) and compared it to measured Δ P L during passive (VC or PC) and spontaneous breathing (NAVA or NPS). A requirement for the LAM was similar Edi to the assisted breath. Results All animals successfully underwent titrations and LAMs for NPS/NAVA. One thousand seven-hundred ninety-two (1792) breaths were matched to passive ventilation titrations (matched Vt, r  = 0.99). P L_LAM demonstrated strong correlation with P L _ CMV ( r  = 0.83), and P L _ P ES ( r  = 0.77). Bland–Altman analysis revealed little difference between the predicted P L _ LAM and measured P L _ CMV (Bias = 0.49 cm H2O and 1.96SD = 3.09 cm H2O). For P L _ P ES , the bias was 2.2 cm H2O and 1.96SD was 3.4 cm H2O. Analysis of Edi and P ES at peak Edi showed progressively increasing uncoupling with increasing assist. Conclusion During synchronized mechanical ventilation, a LAM breath allows for estimations of transpulmonary driving pressure, without measuring P ES , and follows a mathematical transfer function to describe respiratory muscle unloading during synchronized assist.