The energized uptake of low levels of Ca2+ in the presence and absence of phosphate by isolated rat liver mitochondria, and the perturbation effected by this activity on ultrastructural and metabolic parameters of mitochondria have been investigated. In the presence of phosphate, low levels of Ca2+ are taken up by mitochondria and result in various degrees of ultrastructural expansion of the inner mitochondrial compartment. This indicates that low levels of Ca2+ in the presence of phosphate, are accumulated in an osmotically active form into the water phase of the inner compartment. The first clearly observable quantitative increase in the volume of the inner compartment occurs after the accumulation of 100 nmoles Ca2+/mg protein. An accumulation of 150–200 nmoles Ca2+/mg protein, which is equivalent to the osmolar concentration of endogenous K+, is required to effect a doubling of the volume of the inner compartment. This degree of osmotic perturbation occurs as mitochondria transform from a condensed to an orthodox conformation. The osmotically induced orthodox conformation differs from the mechanochemically induced orthodox conformation previously described, in that its development is concomitant with a marked decrease in acceptor control and oxidative phosphorylation efficiency and it fails to transform to a condensed conformation in response to addition of ADP. In the absence of added phosphate, a maximum of 190 nmoles Ca2+/mg protein was found to be taken up by mitochondria (state 6). Ca2+ is apparently bound under state 6 conditions since the uptake does not effect an ultrastructural expansion of the inner compartment. Phosphate added after state 6 Ca2+ binding, however, results in an immediate ultrastructural expansion of the inner compartment. The addition of phosphate to mitochondria in the absence of exogenous Ca2- fails to effect an osmotic ultrastructural transformation. Under state 6 conditions, the binding of between 40 and 190 nmoles Ca2+/mg protein results in the formation of dense matrix inclusions which appear to be composed of tightly packed, concentrically oriented membranes. Under conditions in which the bound Ca2+ is subsequently released, there is a concomitant loss in the density of these matrix inclusions, leaving behind morphologically distinct membrane whorls in the mitochondrial matrix.
Fluorescence depolarization measurements on oriented membranes
