We have reported the linear relation of myocardial oxygen consumption per beat (VO₂) and systolic pressure-volume area (PVA) in the left ventricle of the cross-circulated rat heart. The VO₂ intercept (PVA-independent VO₂) is primarily composed of VO₂ for Ca²⁺ handling in excitation-contraction coupling and basal metabolism. Recently, we proposed a new index for oxygen cost of contractility obtainable as a slope of a linear relation between PVA-independent VO₂ and left ventricular contractility. This index indicates the Ca²⁺ handling VO₂ per unit contractility change. However, a dependency of this index on heart rate has not yet been investigated. The aim of the present study was to investigate the dependency of oxygen cost of contractility on heart rate. This is a critical point to compare this cost under different heart rates. At first we found no differences of VO₂-PVA relations at 240 and 300 beats/min (bpm). Therefore, after control VO₂-PVA relation at 300 bpm, we gradually enhanced left ventricular contractility by Ca²⁺ at a midrange left ventricular volume and obtained the gradually increased PVA-independent VO₂. At each contractility level, the pacing rate was alternately changed at 240 and 300 bpm. We obtained the two composite VO₂-PVA relation lines and found no significant differences between the slopes of PVA-independent VO₂ and left ventricular contractility relations at 240 and 300 bpm. The present results indicated no dependency of oxygen cost of left ventricular contractility on heart rates within 240-300 bpm. Based on this fact, we concluded that even under the different pacing rates within 240-300 bpm, this oxygen cost is valid for assessing cardiac mechanoenergetics, especially the economy of total Ca²⁺ handling in E-C coupling.