Characterization of sarcoplasmic reticulum in skinned heart muscle cultures

Abstract

The plasma membranes of rat heart muscle, grown in cell culture, were made permeable with saponin in a Ca-free solution. The cells were then supplied with a medium resembling the cytosol, and the adenosine triphosphate (ATP)-dependent Ca2+ sequestration was measured in the presence of oxalate. The nonmitochondrial component accounts for about 50% of the total Ca2+ uptake. The nonmitochondrial accumulation of Ca2+ within myocardial cells was found to be reversible by addition of the Ca2+ ionophore A23187. On the other hand, the Ca2+ antagonist D-600 (50 microM) had almost no effect on Ca2+ accumulation. Caffeine reduced Ca2+ accumulation in the skinned cardiomyocytes in a concentration-dependent manner. In addition, the anticalmodulin drug trifluoperazine (TFP) reduced Ca2+ accumulation in the skinned cells. Because of the analogy between nonmitochondrial ATP-dependent Ca2+ accumulation and the sarcoplasmic reticulum (SR) function with regard to the influence of various agents, it is assumed that we actually measure Ca2+ accumulation in the SR. The rate of Ca2+ accumulation into the SR measured during the development of the cardiomyocytes in culture shows an almost linear increase as a function of culture age. Amiodarone, a potent antiarrhythmic agent, and its metabolite, desethylamiodarone, inhibited Ca2+ accumulation into SR, which may explain their therapeutic effect.