H+ Changes Associated with Divalent Cation Uptake by Mouse Liver Mitochondria

The effect of concentration of Ca ++ and mitochondria on the stoichiometry of H + release associated with Ca ++ uptake supported by succinate oxidation was examined. Ratios less than unity were observed with low concentrations of Ca ++ , and higher ratios were obtained with intermediate concentrations of Ca ++ . The highest values (greater than unity but less than 2) could be obtained with increased mitochondrial concentrations. The concentrations of Ca ++ at which the highest ratios were obtained were shifted to higher levels as the mitochondrial concentration was raised. The low ratios observed with low levels of divalent cations are attributed in part to the metabolism-independent binding of Ca ++ , since 45 Ca ++ was observed to be bound by mitochondria in the presence of antimycin and oligomycin. This energy-independent uptake of Ca ++ was also dependent on the concentration of Ca ++ and mitochondrial protein. In contrast to the energy-dependent process, the release of H + was small or absent when antimycin was present. Added NaCl or KCl, but not further additions of mannitol or sucrose, completed for this Ca ++ -accessible space. These findings suggested that H + :Ca ++ ratios of unity may be underestimated, and those ratios which are based on the assumption that all of the added calcium ions are taken up by an energy-linked mechanism may have to be corrected. Under conditions where the ratio was corrected for the energy-independent binding of Ca ++ , the (K + + H + ):Ca ++ ratio approached 2.