The end-diastolic pressure-volume relationship (EDPVR) is routinely used to determine passive left ventricular stiffness, although the diastatic pressure-volume relationship (D-PVR) has also been measured. Based on the physiologic difference between diastasis (LV and the atrium are relaxed and static), and end-diastole, (LV volume increased by atrial systole and atrium is contracted), we hypothesized that although both D-PVR and EDPVR include LV chamber stiffness information, they are two different, distinguishable PV relations. Cardiac catheterization determined LV pressures and conductance volumes in 31 subjects were analyzed. Physiologic, beat-to-beat variation of the diastatic and end-diastolic P-V points were fit by linear and exponential functions to generate the D-PVR and EDPVR. The extrapolated exponential D-PVR underestimated LVEDP in 82% of the heart beats (p<0.001). The extrapolated EDPVR overestimated pressure at diastasis (P_diastasis) in 84% of the heart beats (p<0.001). If each subject's diastatic and end-diastolic P-V data are combined to form a continuous data-set to be fit by one exponential, the goodness-of-fit is always worse than if the diastatic and end-diastolic data are grouped separately and fit by two distinct exponential relations. Diastatic chamber stiffness was less than EDPVR stiffness (defined by the slope of PVR), for all 31 subjects (0.16±0.11mmHg/ml vs. 0.24±0.15mmHg/ml, p<0.001). We conclude that the D-PVR and the EDPVR are distinguishable. Because it is not coupled to a contracted atrium, the D-PVR conveys passive LV stiffness better than the EDPVR. Additional studies that fully elucidate the physiology and biology of diastasis in health and disease are in progress.