We investigated theoretically and experimentally the role of Rho kinase (RhoK) in Ca²⁺-contraction coupling in rat airways. Isometric contraction was measured on tracheal, extra- and intrapulmonary bronchial rings, in response to carbachol and 50 mM external KCl (n=6 to 8). [Ca²⁺]_i was recorded in freshly isolated tracheal myocytes using Indo-1 (n=19 to 25). Values are expressed as mean ± SEM. Statistical comparisons were done using a Student’s t test. Theoretical modeling consisted in a four-state model of the contractile apparatus coupled with a model of Ca²⁺-dependent MLCK activation and RhoK-dependent MLCP inactivation. Analysis of the time course of contraction to carbachol (0.3 and 10 µM) showed that force development occured in two phases: (i) a short-time, Hill-shaped contraction obtained within 90 s, (ii) followed by a maintained or an additional delayed contraction. Hill fitting of the first phase showed that the short-time maximal contraction (stFmax) of tracheal rings to 10 µM carbachol was 77,8±2.6% of total maximal force, and the time to obtain half-stFmax was 17.4±1.6 s. Values of similar range were obtained in bronchial rings. [Ca²⁺]_i responses to 10 µM acetylcholine (ACh) consisted in a fast peak followed by a plateau and, in 42% of the cells, superimposed Ca²⁺ oscillations. Exposure to the RhoK inhibitor Y27632 (10 µM) did not alter the resting [Ca²⁺]_i nor the parameters of the ACh-induced Ca²⁺ response. Whatever the concentration of carbachol and the location along the airway tree, Y27632 did not modify the basal tension but decreased the amplitude of the short-time response, without altering the additional delayed contraction. Calyculin A (MLCP inhibitor) increased the basal tension, and abolished the effect of RhoK inhibition on carbachol-induced contraction. Stimulation by 50 mM external KCl solution in the presence of atropine induced a short-time contraction followed by a sustained tension, both depending of the presence of external Ca²⁺. As with carbachol-induced contraction, Y27632 decreased the amplitude of the short-time response, without altering the delayed tension. KN93 (CamK II inhibitor) and DIDS (Ca²⁺-activated Cl^– channels) had not influence on the effect of RhoK inhibition. We conclude that Ca²⁺-dependent but CamK II-independent RhoK activation contributes to the early phase of the contractile response via MLCP inhibition. On these bases, we inplemented our previously published model of Ca²⁺-contraction coupling (1). The model explains the time course of the short-time contraction and the role of RhoK by Ca²⁺-dependent activation of MLCK and RhoK, which inactivates MLCP. Oscillatory and non-oscillatory [Ca²⁺]_i responses result in a non-oscillatory contraction which amplitude is encoded by the plateau value and oscillation frequency.