We tested the hypothesis that loss of mitochondrial
adenine nucleotides during
myocardial ischemia is induced by the accumulation of
inorganic phosphate (Pi) and a decrease in cytosolic
ATP. In the isolated perfused rat heart, loss of mitochondrial
adenine nucleotides (
ATP +
ADP +
AMP) was preceded by the rise in tissue Pi and the loss of tissue
ATP. After 30 min
ischemia, the average rate of loss of mitochondrial
adenine nucleotides was c. 1.5% of the initial pool/min. In isolated heart mitochondria, there are two pathways for
adenine nucleotide release: a 'fast',
phosphate-dependent pathway, which is inhibited by
atractyloside; and a 'slow',
phosphate-independent pathway, which is insensitive to
atractyloside. Decreasing the pH from 7.4 to 6.5 significantly decreased the rate of release by the
phosphate-dependent pathway (but not the
phosphate-independent pathway). Analysis of release rates indicated that HPO4-2 is responsible for the
phosphate-induced release; Vmax = 53.8% of the pool/per minute, Km = 7.5 mM. In vitro, extramitochondrial
ATP inhibited
adenine nucleotide release in the presence of Pi such that the rate of release was inversely proportional to the extramitochondrial [
ATP]; extrapolation to zero
ATP indicated a release rate of 2 to 3% of the pool/per minute, which is approximately equal to the rate of the 'slow'
phosphate-independent pathway. Moreover, increasing the Pi concentration did not increase the rate of
adenine nucleotide release in the presence of extramitochondrial
ATP. Accumulation of mitochondrial
adenine nucleotides was observed when the mitochondria were incubated in the presence of 4 mM or greater
ATP. The results suggest that the rise in intracellular Pi during
myocardial ischemia does not induce the loss of
adenine nucleotides from the mitochondrial compartment, but rather that degradation of cytosolic
ATP results in a slowing of
ATP influx such that the rate of efflux (
phosphate-independent) exceeds the rate of influx.