Role of adenosine kinase and AMP deaminase in the regulation of cardiac purine release

Abstract

To analyze the relation between cardiac energy status, adenosine formation, and purine release, reliable measurements of the cytosolic concentrations of ATP, ADP, AMP, and adenosine are required. Based on the creatine kinase and myokinase equilibrium, ADP and AMP are determined by 31 P nuclear magnetic resonance spectroscopy, whereas free cytosolic adenosine is measured by the S-adenosyl-homocysteine (SAH) technique. Combining these methods with efflux measurements, selective enzyme blockade and a comprehensive model analysis enables a description of both concentrations and flux rates in purine metabolism. In the well-oxygenated heart, adenosine is predominantly formed intracellularly from AMP, but also from S-adenosyl-homocysteine. Net adenosine formation (2.3 nmol/min per g) exceeds coronary venous release (0.07 nmol/min per g) more than 30-fold, because most of the adenosine formed is rephosphorylated by adenosine kinase. This enzyme maintains a low intracellular adenosine and limits both adenosine release and deamination to inosine. In fact, inosine is mainly formed from IMP (1.8 nmol/ min per g) the product of AMP deaminase. Inosine, hypoxanthine, xanthine, and uric acid (1.1, 0.4, 0.2, 1.4 nmol/min per g) are the main purine catabolites released. In the oxygen-limited heart, energy status is impaired and AMP increased. Under these conditions, a linear relation between AMP (200-3,000 nmol/liter), net adenosine formation, as well as net inosine formation is observed. It is, thus, the AMP substrate concentration that directly controls adenosine formation by cytosolic 5'-nucleotidase and most likely flux through AMP deaminase. Hypoxia-induced inhibition of adenosine kinase shunts adenosine from the salvage pathway to venous release and causes the amplification of small changes in AMP into a major rise in adenosine. This mechanism plays an important role in the high sensitivity of the cardiac adenosine system to impaired oxygenation.