Sodium-calcium exchange in membrane vesicles from Artemia

Abstract

Membrane vesicles were prepared from Artemia nauplii (San Francisco Bay variety) 45 h after hydration of the dry cysts. Na +-loaded vesicles accumulated up to 10 nmol Ca 2+/mg protein when diluted 50-fold into 160 m m KCl containing 15 μ m CaCl 2. Practically no accumulation of Ca 2+ was observed if the vesicles were diluted into 160 m m NaCl instead of KCl, or if they were treated with monensin, a Na + ionophore, for 30 s prior to addition of CaCl 2 to the KCl medium. These observations indicate that the Artemia vesicles exhibit Na-Ca exchange activity. The velocity of Ca 2+ accumulation by the vesicles in KCl was stimulated 2.6-fold by the K + ionophore valinomycin, suggesting that the exchange system is electrogenic, with a stoichiometry greater than 2Na + per Ca 2+. K m,Ca and V max values were 15 μ m and 7.5 nmol/mg protein·s, respectively. Exchange activity in the Artemia vesicles was inhibited by benzamil (IC 50 ≅ 100 μ m) and by quinacrine (IC 50 ≅ 250 μ m), agents that also inhibit exchange activity in cardiac sarcolemmal vesicles. Unlike cardiac vesicles, however, exchange activity in Artemia was not stimulated by limited proteolysis, redox reagents, or intravesicular Ca 2+. This indicates that the two exchange systems are regulated by different mechanisms. Vesicles were prepared from Artemia at various times after hydration of the dry cysts and examined for exchange activity. Activity was first observed at approximately 10 h after hydration and increased to a maximal value by 30–40 h; hatching of the free swimming nauplii occurred at 18–24 h. The results suggest that hatching Artemia nauplii might be a particularly rich source of mRNA coding for the Na +Ca 2+ exchange carrier.