Structural and Functional Characterization of Dog Heart Microsomes

Abstract

Several procedures for the preparation of microsomes from heart tissue were compared. Sucrose was found the most effective addition to preserve the activity of the sarcoplasmic reticulum vesicles. Electron microscopy of sectioned pellets shows uniform vesicles, but negative staining of microsomal suspensions allows recognition of mitochondrial fragments, which are found in cardiac microsomal fractions in greater amounts than in skeletal muscle preparations. The quantity of mitochondrial contaminations in different fractions was estimated by measuring enzymatic activities. A comparison of Ca²⁺ accumulation, Ca²⁺ efflux, and ATPase activity shows identical features in preparations from the heart and from skeletal muscle, although the specific activity is approximately ten times lower in those from the heart. The apparent affinity of sarcoplasmic reticulum for Ca²⁺ in vitro justifies the assumption that sarcoplasmic reticulum lowers the intracellular Ca²⁺ concentration in the in-vivo heart below the level required for contraction. However, in a weight unit of fresh tissue, the ratio between the maximal Ca²⁺ accumulated by SR and the amount of contractile protein is much lower in the heart than in skeletal muscle. Caffeine and, more markedly, propranolol and tetracaine increase efflux of accumulated Ca²⁺ and thereby reduce net uptake by fragmented sarcoplasmic reticulum. Isopropylmethoxamine is not effective at millimolar concentrations.