—During triggered propagated contractions, Ca²⁺ waves travel along cardiac trabeculae with a constant velocity (V_prop) ranging from 0.34 to 5.47 mm/s. To explore the determinants of V_prop, we studied (1) the relationship between [Ca²⁺]_i and V_prop and (2) the effect of low concentrations of caffeine on V_prop. Trabeculae were dissected from the right ventricle of rat hearts. [Ca²⁺]_i was measured using electrophoretically injected fura-2 and an image-intensified CCD camera. Force was measured using a silicon strain gauge, and sarcomere length was measured using laser diffraction techniques. After induction of reproducible Ca²⁺ waves by trains of electrical stimuli (2.5 Hz) at 21.9±0.2°C, the number of stimuli or [Ca²⁺]_o was varied in 9 trabeculae. In 5 trabeculae, the effects of caffeine (0.1 to 1.0 mmol/L) at [Ca²⁺]_o of 2.2±0.3 mmol/L were determined. All images were recorded under stable conditions of wave propagation. The increment in [Ca²⁺]_i during the last electrically stimulated transient (ΔCa_T) and [Ca²⁺]_i just before onset of the Ca²⁺ waves (Ca_D) were used to estimate the Ca²⁺ loading of the sarcoplasmic reticulum (SR) and the myoplasm, respectively. The ratio (ΔCa_W/ΔCa_T) of the [Ca²⁺]_i increment during the waves (ΔCa_W) to ΔCa_T was used to estimate the probability of opening of the SR-Ca²⁺ release channel during wave propagation. As a result of an increase of the number of stimuli or [Ca²⁺]_o, V_prop increased in proportion to (1) ΔCa_T (r=0.82); (2) Ca_D (r=0.88); (3) ΔCa_W (r=0.85); and (4) ΔCa_W/ΔCa_T (r=0.74). The addition of caffeine (≤0.3 mmol/L) increased V_prop for any ΔCa_T and any ΔCa_W, revealing an increased sensitivity of V_prop to ΔCa_T and ΔCa_W. In contrast, caffeine had little effect on the relationship between V_prop and Ca_D and no effect on that between V_prop and ΔCa_W/ΔCa_T. These results suggest that both the cellular Ca²⁺ loading and open probability of the SR-Ca²⁺ release channels determine the velocity of propagation of Ca²⁺ waves.