Effect of W7 on Ca 2+ uptake and Ca 2+-ATpase activities of the endoplasmic reticulum of rat liver

Abstract

In an initial attempt to use calmodulin antagonists as probes to study the role of calmodulin in the modulation of Ca 2+ uptake activity in the endoplasmic reticulum of rat liver, we noticed that W7 had a differential effect on the Ca 2+ uptake and Ca 2+-ATPase activities. To test the specificity of this effect and explore the underlying mechanism, we examined the effects of W7 on Ca 2+ 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 Ca 2+ accumulation in both preparations in a dose-dependent fashion but the half-maximal inhibitory concentrations were different for Ca 2+ accumulation (90 μ m) and Ca 2+-ATPase activity (500 μ m). Kinetic analysis indicated that the inhibition of both Ca 2+ uptake and Ca 2+-ATPase activity by W7 was noncompetitive with respect to Ca 2+ and ATP. Addition of W7 did not enhance the rate of Ca 2+ efflux from microsomes after Ca 2+ 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 Ca 2+ handling were not the result of increased membrane permeability to Ca 2+ but rather were due to dissociation of Ca 2+ uptake from Ca 2+-ATPase activity.