A covalent regulatory mechanism involving Ca²⁺-dependent cross-bridge phosphorylation determines both the number of cycling cross bridges and cycling kinetics in mammalian smooth muscle. Our objective was to determine whether a similar regulatory mechanism governed smooth muscle contraction from a poikilothermic amphibian in a test of the hypothesis that myosin regulatory light chain (MRLC) phosphorylation could modulate shortening velocity. We measured MRLC phosphorylation of Rana catesbiana urinary bladder strips at 25°C in tonic contractions in response to K⁺ depolarization, field stimulation, or carbachol stimulation. The force-length relationship was characterized by a steep ascending limb and a shallow descending limb. There was a rapid rise in unloaded shortening velocity early in a contraction, which then fell and was maintained at low rates while high force was maintained. In support of the hypothesis, we found a positive correlation of the level of myosin phosphorylation and an estimate of tissue shortening velocity. These results suggest that MRLC phosphorylation in amphibian smooth muscle modulates both the number of attached cross bridges (force) and the cross-bridge cycling kinetics (shortening velocity) as in mammalian smooth muscle.