Thermodynamic State of Cytosolic Adenylates in Guinea Pig Myocardium. Energy-Linked Adaptive Changes in Free Adenylates and Purine Nucleoside Release

Abstract

The phosphorylation state of cytosolic ATP ([ATP]/([ADP][Pi)) can be calculated from the measured reactants of creatine kinase, including H+. Measured total tissue ATP/(ADP x Pi) ratio underestimates [ATP]/([ADP][Pi]) by up to two orders of magnitude, mainly because total tissue ADP grossly overestimates the thermodynamic concentration of ADP ([ADP]) in the soluble cytoplasm (cytosol). Both ADP concentration estimates using the creatine kinase equilibrium and published estimates of cytoplasmic binding sites for ADP indicate that, in muscle and heart in particular, ADP is compartmented between cytosol and mitochondria and that most of cytoplasmic ADP is bound to actin. Current data on [ADP] place its concentration between 16 to 60 μM as a function of cardiac energy output which determines oxygen usage (MVO2). Similarly, employing the myokinase equilibrium to estimate the thermodynamic concentration of AMP ([AMP]), the calculated free cytosolic AMP concentrations exhibited highly significant square dependencies on MVO2, [ADP], and the [ADP]/[ATP] ratio, respectively. Calculated [AMP] ranged from approximately 100 to 800 nM as a function of the [ADP]/[ATP] ratio in normal and ischemic hearts. Considering that measured total AMP was in the submillimolar range, the data suggested that most of cardiac AMP was not in catalytic contact with myokinase, i. e., probably located in the mitochondrial compartment. Net release of adenosine plus inosine (V(AR+INO)) was directly related to [AMP] and cellular lactate/pyruvate ratio, respectively. These results suggested that the availability of free AMP may be a determinant of 5′-nucleotidase plus adenylate deaminase activities and that V(AR+INO) was reciprocally linked to moycardial energy state as quantitated by the [ATP]/([ADP][Pi]) ratio. Such energy-linked V(AR+INO) allows for adenosine formation in normoxic heart without assuming a microhypoxia stimulus. According to this model the key trigger for net ATP degradation and V(AR+INO) during both normoxia and ischemia is the actually incurred myocardial energy deficit or the cytosolic ATP potential, not a change in the oxygen supply-demand ratio per se.