The effect of diabetes mellitus on the regulation of urinary bladder smooth muscle contraction was studied. Diabetes was induced in the rabbit by alloxan injection followed by 16 weeks of housing. The bladder was harvested and strips of wall devoid of both mucosa and serosa were examined. Intact strips of bladder smooth muscle from diabetic animals produced less stress in response to membrane depolarization than muscle from control animals; sensitivity to KCl was not changed. Carbachol responses were similar in muscle strips from the two animal groups. Basal myosin light chain (MLC) phosphorylation levels were significantly elevated in response to most stimuli in muscle strips from diabetic animals, although levels of stress were either unchanged or lower. Alpha toxin permeabilized strips which allow for control of the intracellular environment while maintaining excitation-contraction coupling showed increased levels of MLC phosphorylation but decreased sensitivity to activator Ca²⁺ in smooth muscle from diabetic animals. MLC phosphatase contents were similar in smooth muscle from the two animal groups; however MLC phosphatase activity was greater in muscle from control as compared to diabetic animals. These results suggest that diabetes mellitus uncouples basal MLC phosphorylation from force in the bladder smooth muscle cell.