Several experimental procedures were used to increase the intracellular concentration of Ca2+ and determine its effects on protein turnover in isolated extensor digitorum longus and soleus muscle. These methods included the use of ionophore A23187, caffeine, dibucaine, thymol and procaine, all agents known to induce the release of calcium by acting either on the sarcolemma and/or on the sarcoplasmic reticulum. Another approach involved varying the external concentration of Ca2+ in the media in which the muscles were incubated. The changes in muscle Ca2+ concentrations after exposure to the various calcium-releasing agents were in keeping with accepted modes of action of these agents on muscle membranes. The findings suggest that increasing the sarcoplasmic concentration of Ca2+ inhibits protein synthesis and enhances protein breakdown. These catabolic effects of Ca2+ are compared with the changes induced in muscle protein turnover after exposure to insulin or cyclic nucleotides, and in myopathic muscle and situations of work overload. Attention is also drawn to some of the difficulties involved in definitively implicating Ca2+ as a factor involved in the normal regulation of protein turnover.