Combined glyceraldehyde‐3‐phosphate dehydrogenase/phosphoglycerate kinase in catecholamine‐stimulated guinea‐pig cardiac muscle: Comparison with mass‐action ratio of creatine kinase

The steady‐state reactant levels of triose‐phosphate isomerase and the glyceraldehyde‐3‐phosphate dehydrogenase/phosphoglycerate kinase system were examined in guinea‐pig cardiac muscle. Key glycolytic intermediates, including glyceraldehyde 3‐phosphate were directly measured and compared with those of creatine kinase. Nonworking Langendorff hearts as well as isolated working hearts were perfused with 5 mM glucose (plus insulin) under normoxia conditions to maintain lactate dehydrogenase near‐equilibrium. The cytosolic phosphorylation potential ([ATP]/([ADP]·[P_i]) was derived from creatine kinase and the free [NAD⁺]/(NADH]·[H⁺]) ratio from lactate dehydrogenase. In Langendorff hearts glycolysis was varied from near‐zero flux (hyperkalemic cardiac arrest) to higher than normal flux (normal and maximum catecholamine stimulation). The triose‐phosphate isomerase was near‐equilibrium only in control or potassium‐arrested Langendorff hearts as well as in postischemic ‘stunned’ hearts. However, when glycolytic flux increased due to norepinephrine or due to physiological pressure‐volume work the enzyme was displaced from equilibrium. The alternative phosphorylation ratio [ATP]‘/([ADP]’·[P_i]) was derived from the magnesium‐dependent glyceraldehyde‐3‐phosphate dehydrogenase/phosphoglycerate kinase system assigning free magnesium different values in the physiological range (0.1–2.0 mM). As predicted, [ATP]/([ADP]·[P_i]) and [ATP]'/([ADP]'·[P_i]') were in excellent aggreement when glycolysis was virtually halted by hyperkalemic arrest (flux ∼ 0.2 μmol C₃· min^‐1· g dry mass^‐1). However, the equality between the two phosphorylation ratios was not abolished upon resumption of spontaneous beating and also not during adrenergic stimulation (flux ∼ 5–14 μmol C₃· min^‐1· g dry mass^‐1). In contrast, when flux increased due to transition from no‐work to physiological pressure‐volume work (rate increase from ∼ 3 to 11 μmol C₃· min^‐1· g dry mass^‐1), the two ratios were markedly different indicating disequilibrium of the glyceraldehyde‐3‐phosphate dehydrogenase/phosphoglycerate kinase. Only during adrenergic stimulation or postischemic myocardial ‘stunning,’ not due to hydraulic work load per se, glyceraldehyde‐3‐phosphate levels increased from about 4 μM to > 16 μM. Thus the guinea‐pig cardiac glyceraldehyde‐3‐phosphate dehydrogenase/phosphoglycerate kinase system can realize the potential for near‐equilibrium catalysis at significant flux provided glyceraldehyde‐3‐phosphate levels rise, e.g., due to ‘stunning’ or adrenergic hormones.