Abstract
Calcium uptake by vesicles in a highly enriched sarcolemma preparation from canine ventricle was found to be markedly stimulated by intravesicular calcium. Stimulation of calcium uptake appeared to be a saturable function of intravesicular calcium. Calcium efflux from the vesicles was stimulated by calcium in the reaction medium. Calcium uptake, supported by intravesicular calcium, and calcium efflux, stimulated by extravesicular calcium, were found to correspond on a one-to-one basis. Only small changes in net uptake or efflux were observed to occur in response to chemical gradients of calcium across the membrane. It was concluded, therefore, that under certain conditions, the major means for calcium movement across vesicles in the preparation is via a one-to-one exchange of calcium. Sodium was found to stimulate calcium uptake when present in the intravesicular space and to stimulate calcium efflux when present in the extravesicular space, but the effects of calcium plus sodium were not additive with respect to stimulating either calcium uptake or efflux. The effects of unlabeled calcium, strontium, barium, and magnesium on calcium uptake stimulated by intravesicular calcium and by intravesicular sodium were similar though not identical. The temperature dependence for calcium-stimulated and sodium-stimulated calcium movements was characterized by Q10 values of 1.27 and 2.06, respectively. Previous work has associated the sodium-calcium exchange reaction with the sarcolemma. It is argued that the present study, in turn, provides evidence that the calcium-calcium exchange reaction is also associated with the sarcolemma. In addition, the results of the study are consistent with the hypothesis that one membrane system can promote the exchange of either calcium for calcium or calcium for sodium.
Highlights
Calcium uptake by vesicles ina highly enriched sar- diastole
The effects of unlabeled calcium, stron- niques to intactmyocardial tissue preparations has generated tium, barium, and magnesium on calcium uptake stimth-ree hypotheses about how calciummaycross the sarcoulated by intravesicular calcium and by intravesicular lemma: (a)a voltage-dependent, verapamil-sensitivechannel sodium were similar though not identicaTlh. e temper- [5, 6] that may be the major pathway for net calcium moveature dependence for calcium-stimulated and sodium- ment into themyocardial cell;( b )an exchange reaction which stimulated calcium movements was characterized by couples the movement of calcium into and/or out of the
Inclusion of0.1 to 1.0 m~ calcium in the loading medium markedly increased the uptake of calcium from the reaction medium (Fig. 1).As stated under “Experimental Procedures,” calcium was observed to enter the vesicles from the loading medium and equilibration between the loading medium and the intravesicular space was reached by 12 h
Summary
Aliquots of the sarcolemma-enriched preparatiownere equilibrated overnight at 4-9°C withamediumcontaining 150 mM KC1 and 10 tassium-phosphatase activity, andhigh affinity (-)dihydroalprenolol mM Tris-C1, pH 7.4. After 60 s, the reaction was terminated by the addition of 5 activity, were less in the sarcolemma preparation thanin the homog- ml of an ice-cold medium (“stopping solution”) containing 150 mM enate. Calcium Transport Studies-The following two-stage procedure to a 60-s reaction, whereas Experiment B equates tozero time. Aliquots of the initiation of contact between the membranes and the stopping solusarcolemma-enrichedpreparation wereexposed to a “loading me- tion, the tubeswere placed on ice and thesuspension was filtered 5 to dium” for 12 to 18 h at 4-9°C. The fdters werewashedwithtwo 5-ml aliquots of the ligands to enter the membranveesicles from theloading medium and stopping solution and 4“Cacontent of the filters was determined.
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