The effect of Mg2+ on the contraction of glycerinated smooth and skeletal muscles was examined. In the absence of Ca2+, glycerinated smooth muscle contracted slowly when exposed to 5 mM adenosine triphosphate (ATP) and Mg2+. The magnitude of this contraction depended on the concentration of Mg2+ in a range of 1--20 mM. When 20 mM Mg2+ was used with ATP, glycerinated smooth muscle contracted almost maximally in a Ca2+ free environment, and it did not contract further on subsequent addition of 0.1 mM of free Ca2+ (EGTA-CaEGTA buffer). In contrast, in skeletal muscle a Ca2+ dependent contraction could always be elicited in the presence of 1--20 mM Mg2+ and 5 mM ATP. Tension development in smooth muscle induced by Mg2+ and ATP was not due to a contamination by Ca2+ or the removal of a Ca2+ sensitive protein. In the present experimental conditions, glycerinated smooth muscle developed a Mg2+-dependent contraction in a Ca2+ free medium, but glycerinated skeletal muscle did not.