The Ca2+ Permeability of Sarcoplasmic Reticulum Vesicles. I. Ca2+ Outflow in the Non-energized State of the Calcium Pump

Passive Ca2+ permeability of sarcoplasmic reticulum vesicles has been studied after maximal loading with Ca2+ (150-200 nmol/mg protein) in the presence of Ca2+, MgATP and an ATP generating system of limited capacity. Outflow of accumulated Ca2+ in the non-energized state of the system was studied by depletion of the medium of one of the substrates, either MgATP (by complete consumption) or Ca2+ (by complexation with EGTA). It was found that Ca2+ outflow under these conditions is relatively slow and independent of the medium concentration of Ca2+ (5 X 10(-9)-5 X 10(-5) M) or MgATP (0.7-730 microM). Outflow curves were steep at the beginning of the outflow phase (30-60 nmol/min per mg protein), and outflow proceeded at a much lower rate below 100 nmol Ca2+/mg protein. Outflow could be completely inhibited by La3+. The Ca2+ release curves are not compatible with simple diffusion, and cannot be accounted for by Ca2+ binding inside the vesicles. Neither are our observations consistent with permeation mediated via the Ca2+ translocation sites involved in active transport. We suggest that non-energized Ca2+ outflow may proceed by a process of ion-exchange through negatively charged, water-filled channels in the membrane, the properties of which are altered by a high intravesicular concentration of Ca2+.