I skeletal muscle, the sarcoplasmic reticulum (SR) serves as the primary organelle involved in the control of cytosolic free calcium (Caf) transients and hence muscle contractility.1 Regulation of Caf by the SR is accomplished via specialized proteins that provide for the control of Ca release, Ca storage, and Ca uptake.2 An alteration in any one of these properties either intrinsically or by changes in the intracellular environment (metabolic byproduct accumulation) could alter Caf levels during exercise and modify muscle contractility.1 It is possible that at least 1 mechanism underlying the high incidence of exercise intolerance and fatigue in chronic congestive heart failure (CHF)3 is a disturbance in SR function. One property of the SR that has received considerable attention in fatigue studies is Ca uptake. Ca sequestration is controlled by Ca adenosine triphosphatase (ATPase), a tripartite membrane protein that contains domains for adenosine triphosphate binding, Ca binding, phosphorylation, and energy transduction, and provides for the transportation of 2 Ca ions into the lumen of the SR per adenosine triphosphate molecule hydrolyzed.2 Studies in humans have found reductions during exercise in intrinsic Ca ATPase activity4,5 and Ca uptake,5,6 measured under optimal “in vitro” conditions. Intrinsic alterations in Ca ATPase activity can also occur via long-term regulatory changes in the enzyme, given the variety of stimuli that can effect changes in expression of the various SR proteins,7,8 some of which may be present in CHF.9 This study compares the differences in SR maximal Ca activated ATPase activity in skeletal muscle between chronic male and female CHF patients and age-matched healthy controls. • • • Volunteers, men and women, recruited from a population of chronic CHF and healthy, age-matched control groups served as subjects in this study. The CHF patients, which included 18 men and 6 women, were patients at both the Duke Medical Center and the Henry Ford Hospital. All CHF patients were free of claudication, rales, and peripheral bruits. Fourteen had ischemic cardiomyopathy and 9 had nonischemic cardiomyopathy. Medications, which had been stabilized for at least 3 months before sampling, included digoxin (23 of 24), diuretics (22 of 24), angiotensinconverting enzyme inhibitors (22 of 24), and longacting nitrates (11 of 24). The exception was digoxin, which had been withdrawn from Duke University patients 7 to 10 days before sampling. No patient with CHF was accepted into the study if they had insulindependent diabetes, clinically significant chronic obstructive pulmonary disease, or peripheral vascular disease. The healthy normals, all recruited from Duke University, had no cardiopulmonary dysfunction or symptoms of ischemic heart disease. As with the CHF patients, all were sedentary. This study, including all procedures, was approved by the institutional review boards of Duke University, the Durham Veterans Administration Medical Centers, and Henry Ford Hospital. Each volunteer was informed of the specifics of each protocol and the potential risks before obtaining written consent and before participation. All subjects, normal and with CHF, had tissue extracted from the vastus lateralis muscle,10 which was quickly frozen in liquid nitrogen and stored at 280°C. Measurements were obtained of both maximal activities of SR Ca ATPase and of 2 enzymes, selected to represent aerobic (citrate synthase) and glycolytic (phosphofructokinase) potential.11 The measurement of maximal Ca ATPase activity was obtained on crude homogenates, prepared at 0°C using a 11:1 (w/v) dilution of buffer containing 200 mM sucrose, 40 mM L-histidine, 1 mM ethylenediaminetetraacetic acid, 10 mM sodium nitrate, and 1 mM dithiothreitol (pH 7.8), according to the procedure of Simonides and van Hardeveld12 and later validated on human tissue by Ruell et al.13 Ca ATPase activity is derived as the difference between total (Ca magnesium ion ATPase) and basal (magnesium ion ATPase) activities. All assays were performed at 37°C using 1 mM the Ca ionophore A23187 (Sigma C-7522). The maximal activities of citrate synthase and phosphofructokinase were performed on homogenates, prepared from frozen tissue that contained 50% glycerol, 20 mM sodium phosphate buffer (pH 7.4), 5 mM b-mercaptoethanol, 0.5 m ethylenediaminetetFrom the Department of Kinesiology, University of Waterloo, Waterloo, Ontario, Canada; Department of Medicine, Division of Cardiology, Duke Center for Living, and Duke University Medical Center, Durham, North Carolina; and Henry Ford Heart and Vascular Institute, Detroit, Michigan. This study was supported by Grant HL 17670 from the National Heart, Lung, and Blood Institute, Bethesda, Maryland. Dr. Green’s address is: Department of Kinesiology, University of Waterloo, Waterloo, Ontario, Canada N2L. E-mail: green@healthy. uwaterloo.ca. Manuscript received July 28, 1999; revised manuscript received and accepted December 29, 1999.
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Jun 17, 2021
Mel Gabriel 
Open Access
