Interaction of Ca2+ with Blowfly Flight Muscle Mitochondria

Abstract

Abstract Blowfly flight muscle mitochondria differ markedly from mammalian mitochondria in their interaction with Ca2+. State 4 respiration of blowfly mitochondria is not stimulated by Ca2+, nor does Ca2+ induce a reversible shift in the oxidation-reduction state of cytochrome b. Nevertheless, Ca2+ is accumulated by respiring blowfly mitochondria, but with a low efficiency; the Ca2+: ∼ accumulation ratio is about 0.6 with pyruvate plus proline, and about 0.2 with glycerol 1-phosphate, compared with values of about 2.0 observed in mammalian mitochondria. Ca2+ uptake is blocked by respiratory chain inhibitors and by uncouplers, but not by oligomycin. The presence of permeant anions in the medium is essential for Ca2+ uptake by blowfly mitochondria. At high levels of Ca2+, phosphate supports Ca2+ uptake better than acetate, but at low Ca2+ levels, phosphate and acetate are equally effective. Since acetate penetrates blowfly mitochondria passively it is probable that respiration-coupled uptake of Ca2+ in blowfly mitochondria must be accompanied by passive entry of matching anions. The accumulation of Ca2+ by blowfly mitochondria is very slow and does not exhibit saturation kinetics; moreover, no high affinity Ca2+ sites characteristic of a specific Ca2+ carrier could be detected. For these reasons it is concluded that the respiration-dependent accumulation of Ca2+ in blowfly mitochondria is not mediated by a specific carrier, but occurs through slow, concentration-dependent, physical diffusion through the membrane, in response to an electrochemical gradient generated by electron transport. The number of low affinity Ca2+-binding sites is surprisingly low, about one-tenth of that found in mammalian mitochondrial, and has been related to the peculiar phospholipid composition of these mitochondria. The findings described strongly suggest that the mitochondria of blowfly flight muscle do not actively segregate Ca2+ at the low concentrations at which it is presumed to occur in the sarcoplasm in vivo.