Rabbit myocardial membrane Ca 2+-adenosine triphosphatase activity: Stimulation in vitro by thyroid hormone

Abstract

The in vitro stimulation of human and rabbit erythrocyte membrane Ca 2+-ATPase activity by physiological concentrations of thyroid hormone has recently been described. To extend these observations to a nucleated cell model, Ca 2+-ATPase activity in a membrane preparation obtained from rabbit myocardium has been studied. Activity of 5′-nucleotidase in the preparation was increased 26-fold over that of myocardial homogenate, consistent with enrichment by sarcolemma. Mean basal enzyme activity in membranes from nine animals was 20.8 ± 3.3 μmol P i mg membrane protein −1 90 min −1, approximately 20-fold the activity described in rabbit red cell membranes. Exposure of heart membranes in vitro to l-thyroxine (T 4) (10 −10 m) increased Ca 2+-ATPase activity to 29.2 ± 3.8 μmol P i ( P < 0.001). Dose-response studies conducted with T 4 showed that maximal stimulatory response was obtained at 10 −10 m. Hormonal stimulation was comparable for l-T 4 and triiodo- l-thyronine (T 3) (10 −10 m). Tetraiodothyroacetic acid was without biological activity, whereas triiodothyroacetic acid and d-T 4, each at 10 −10 m, significantly decreased enzyme activity compared to control (basal) levels. The action of l-T 4 on myocardial membrane Ca 2+-ATPase activity was inhibited by trifluoperazine (100 μ m) and the naphthalenesulfonamide W-7 (50–100 μ m), compounds that block actions of calmodulin, the protein activator of membrane-associated Ca 2+-ATPase. Radioimmunoassay revealed the presence of calmodulin (1.4 μg mg membrane protein −1) in the myocardial membrane fraction and 0.35 μg mg −1 in cytosol. Myocardial Ca 2+-ATPase activity, apparently of sarcolemmal origin, is thus thyroid hormone stimulable. The hormonal responsiveness of this calcium pump-associated enzyme requires calmodulin.