Effect of W7 on Ca2+ uptake and Ca2+-ATPase activities of the endoplasmic reticulum of rat liver

In an initial attempt to use calmodulin antagonists as probes to study the role of calmodulin in the modulation of Ca2+ uptake activity in the endoplasmic reticulum of rat liver, we noticed that W7 had a differential effect on the Ca2+ uptake and Ca2+-ATPase activities. To test the specificity of this effect and explore the underlying mechanism, we examined the effects of W7 on Ca2+ accumulation and release by endoplasmic reticulum in both permeabilized hepatocytes and a subcellular membrane fraction (microsomes) enriched in endoplasmic reticulum. W7 reduced the steady-state Ca2+ accumulation in both preparations in a dose-dependent fashion but the half-maximal inhibitory concentrations were different for Ca2+ accumulation (90 microM) and Ca2+-ATPase activity (500 microM). Kinetic analysis indicated that the inhibition of both Ca2+ uptake and Ca2+-ATPase activity by W7 was noncompetitive with respect to Ca2+ and ATP. Addition of W7 did not enhance the rate of Ca2+ efflux from microsomes after Ca2+ influx had been terminated. The effect of W7 was apparently not related to its calmodulin antagonist properties as the phenomenon could not be demonstrated with the other more specific calmodulin antagonists, calmidazolium or compound 48/80. A similar observation with W7 has also been reported with the endoplasmic reticulum of pancreatic islets (B. A. Wolf, J. R. Colca, and M. L. McDaniel (1986) Biochem. Biophys. Res. Commun. 141, 418-425). We concluded that the effects of W7 on microsomal Ca2+ handling were not the result of increased membrane permeability to Ca2+ but rather were due to dissociation of Ca2+ uptake from Ca2+-ATPase activity.