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The intracellular distribution of cobalt was analysed in the myocardium of exposed and unexposed rats. The exposed rats were given a dietary cobalt supplementation of 40 mg CoS04-7 H20/kg body weight for 8 weeks. The mitochondrial fraction showed the greatest relative increase in cobalt: 0.09 ng/mg protein in the unexposed rats to 8.43 ng/mg protein in the exposed rats. In the exposed rats the submitochondrial particles had the highest levels of cobalt: 19.43 ng/mg protein, followed by the sarcoplasmatic reticulum: 12.3 ng/mg protein. The microsomal 44 000g supernatant also showed an increase, although the levels remained low (0.51 ng/mg protein in the exposed animals). Apparently the calcium-storing organelles had the highest levels of cobalt. This could affect calcium flux in myocardial cells and, secondarily, tension development in cardiac muscle.

Cobalt has been shown to accumulate in the myocardium of uraemic patients and has been suggested as a myocardial toxin inhibiting mitochondrial respiration. In order to study the cellular effects of cobalt exposure three groups of rats (n=12 per group) were fed a diet containing 12% protein without supplementation or with 20 mg and 40 mg CoSO4 7 H2O/kg body weight/day respectively. After 8 weeks the hearts and soleus muscles were removed. Cobalt in tissues and in four cell fractions were analysed with neutron-activation analysis (ng/g wet weight and ng/mg protein respectively). Mitochondrial respiration was analysed as ATP-production rate using pyruvate+malate and palmitoyl-carnitine+malate as substrate. The ATP-production from pyruvate+ malate was unchanged in both heart and skeletal muscle in the exposed animals. With palmitate as substrate, the heart muscle showed a slightly lower ATP-production rate (p<0.05) after the 20 mg cobalt dose, but the rate was unchanged in the group with higher cobalt intake. No changes in ATP-production rate from palmitate was observed in soleus muscle.The microsomal (100 000 g) fraction in the myocardial cells contained significantly higher cobalt concentrations compared to the mitochondrial fraction in both the unexposed (1.4 ng/mg protein vs 0.19, p<0.05) and exposed rats (53.4 ng/mg protein vs 13.2, p<0.005).In conclusion, cobalt showed a large accumulation in myocardial cells, without significant effects on mitochondrial ATP-formation rate from oxidation of pyruvate or palmitate and with the highest cobalt content contained in the microsomal (100 000 g) fraction.

A sensitive method for the measurement of the ATP production rate in isolated skeletal muscle mitochondria is presented. Mitochondrial suspensions were prepared by differential centrifugation from approximately 80 mg of soleus muscle. ATP production rates were measured luminometrically, utilizing a reagent based upon firefly luciferase, which emits light proportional to the ATP concentration. In a group of 10 rats the ATP production rates were measured with the following substrate combinations: pyruvate+malate, palmitoyl-L-carnitine+malate, oc-ketoglutarate, succinate+rotenone and succinate alone. The variance of the method including tissue preparation, protein determination and the luminometric determination of ATP production was estimated to be 10-14% for the various substrates. Compared to values in the literature, the present results show a good agreement for the substrates pyruvate+malate and palmitoyl-L-carnitine+malate, but lower rates were obtained in our study for αα-ketoglutarate and succinate+rotenone. The advantage of the luminometric method is its high sensitivity. Only 30-40 mg of tissue is required for a complete determination, compared to 1-2 g for a similar assay of oxygen consumption. The method is intended for use in human subjects and will facilitate studies of mitochondrial respiration both in patients of different age groups and in healthy subjects.