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1. A dual effect of the polyamine spermine on Ca2+ uptake by isolated rat liver, brain and heart mitochondria could be demonstrated by using a high-resolution system for studying mitochondrial Ca2+ transport. Depending on the experimental situation, spermine had an inhibiting or accelerating effects on mitochondrial Ca(2+)-uptake rate, but invariably increased the mitochondrial Ca2+ accumulation. 2. Both effects were concentration-dependent and clearly discernible on the basis of their different kinetic characteristics. For mitochondria from all three tissues the half-maximally effective concentration for inhibition of the initial rate of Ca2+ uptake was approx. 180 microM, whereas that for the subsequent stimulation of Ca2+ accumulation was approx. 50 microM. 3. Acceleration of the initial uptake rate could be seen when the mitochondria were preloaded with spermine during a 2 min preincubation period and thereafter incubated in a medium without spermine. 4. When such spermine-preloaded mitochondria were incubated in a spermine-containing medium, the increase in Ca(2+)-accumulation capacity was maintained in spite of an unchanged rate of Ca2+ uptake. 5. Mg2+ interacted with the effects of spermine in a differential manner, enhancing the initial inhibition of the rate of mitochondrial Ca2+ uptake and diminishing the subsequent stimulation of mitochondrial Ca2+ accumulation. 6. This dual effect of spermine on mitochondrial Ca2+ transport resolves the apparent paradox that a polycationic compound can act as a stimulator of Ca2+ uptake.

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Alloxan at millimolar concentrations slightly inhibited the velocity of Ca2+ uptake by isolated rat liver mitochondria irrespective of the free Ca2+ concentration between 1 and 10 microM and was an effective concentration-dependent stimulator of mitochondrial Ca2+ efflux. Ninhydrin also slightly inhibited the velocity of mitochondrial Ca2+ uptake but only at free Ca2+ concentrations above 5 microM. However, ninhydrin was a strong stimulator of mitochondrial Ca2+ efflux even at micromolar concentrations, 10-50 times more potent than alloxan. The mitochondrial membrane potential was reduced 10-20% at most by alloxan and ninhydrin. Alloxan and ninhydrin also stimulated Ca2+ efflux from isolated permeabilized liver cells. When isolated intact liver cells had been pre-incubated with alloxan or ninhydrin before permeabilization of the cells the ability of spermine to induce mitochondrial Ca2+ uptake was abolished. Glucose provided the typical protection against the effects of alloxan on mitochondrial Ca2+ transport only in experiments with intact cells but not in experiments with permeabilized cells or isolated mitochondria. Therefore glucose protection is apparently due to inhibition of alloxan uptake into the cell. Glucose provided no protection against effects of ninhydrin under any of the experimental conditions. Thus both alloxan and ninhydrin are potent stimulators of Ca2+ efflux by isolated mitochondria but very weak inhibitors of the velocity of mitochondrial Ca2+ uptake. The direct effects of ninhydrin on mitochondrial Ca2+ efflux may contribute to the cytotoxic action of this agent whereas the direct effects of alloxan on mitochondrial Ca2+ transport require concentrations which are too high to be of relevance for the induction of the typical pancreatic B-cell toxic effects of alloxan. However, the effects on mitochondrial Ca2+ transport during incubation of intact cells which may result from the generation of cytotoxic intermediates during alloxan xenobiotic metabolism may well contribute to the pancreatic B-cell toxic effect of alloxan.