Magnesium (Mg2+) is one of the most abundant ions in the body. In the human body, Mg2+ plays important roles including cofactors in many crucial enzyme systems, especially those involving energy transfer, storage and utilization. Alteration of the concentration of Mg2+ may cause neuromuscular hyperactivity, psychiatric disturbances, calcium/potassium abnormalities, and overactivity of cardiac muscle. Most information on the effect of Mg2+ on muscle contraction has been obtained from studies on cardiac, skeletal, and vascular muscle; much less is known about the effect of Mg2+ in other smooth muscle systems. In the current study, we investigated the effect of Mg2+ on the contraction and intracellular free calcium of rabbit urinary bladder detrusor muscle in response to carbachol and transmural field stimulation (FS). The results can be summarized as follows: (1) Reduction of the concentration of magnesium [Mg2+] from normal Ty-rode’s solution enhanced the spontaneous basal activity, whereas addition of Mg2+ gradually abolished this spontaneous activity. (2) Muscle contraction induced by FS or carbachol was enhanced in Mg2+-free Tyrode’s solution. Addition of Mg2+ inhibited the response to both forms of stimulation in a dose-dependent manner. (3) Inhibitory effects of Mg2+ were potentiated when the Ca2+ concentration in the Tyrode’s solution was reduced to 0.6 mM, whereas increasing the extracellular concentration of Ca2+ (5.4 mM) reduced the inhibitory effects of Mg2+. (4) Using FURA-2 to monitor intracellular free calcium simultaneous with contractile tension, we demonstrated that the alterations in the contractile responses observed at different concentrations of extracellular Mg2+ correlated with similar changes in intracellular free calcium. We conclude from this study that Mg2+ can significantly alter the contractile responses to FS and muscarinic stimulation by alteration of calcium influx during contraction.