Model Systems for Modulating the Free Energy of ATP Hydrolysis in Normoxically Perfused Rat Hearts

This study has two objectives; first, to develop perfusion conditions that decrease the free energy of ATP hydrolysis,ΔGATP, in isolated hearts; and, second, to modulate theΔGATPin these perfused hear models. To accomplish the first goal, a series of inhibitors was employed to restrict acetyl-CoA oxidation. The second goal was accomplished by increasing work demand. Rat hearts were perfused with Krebs–Henseleit solution containing glucose and either; (i) no inhibitors (G group hearts); (ii) 0.3 m bromo-octanoate (BrO), an inhibitor ofβ-oxidation (GB group); (iii) 0.4 m amino-oxyacetate (AOA), an inhibitor of the malate-aspartate shuttle (GA group); (iv) BrO ands AOA (GBA group hearts); or (v) BrO, AOA, and 4 m butyrate, an alternate substrate (GBA-Bu). Pacing hearts at 300 beats per min (beats/min), at 450 beats/min, and at 450 beats/min in the presence of 80μg/l dobutamine allowed three increasing levels of work demand to be attained. TheΔGATPvalues of the five groups of hearts were calculated for each workstate using the concentrations of high energy phosphate metabolites measured by31P NMR spectroscopy. At the highest levels of workload demand, the G, GB, and GBA-Bu group hearts hadΔGATPvalues ≥−53 kJ/mol ATP. At the highest levels of workload demand, the GA and GBA hearts hadΔGATPvalues ≤−49 kJ/mol ATP. The values ofΔGATPand the heart work output during any perfusion conditions were stable for periods of >20 min. The G, GB, and GBA-Bu hearts attained RPPs of ≥54×103mmHg/min at the highest levels of workload demand. The GA and GBA hearts attained RPPs of ≤30×103mmHg/min at the highest levels of workload demand. This study establishes stable, non-ischemic, isolated perfused heart preparations with whichΔGATPcan be easily varied across the physiological relevant range of −56 to −47 kJ/mol ATP.