Rapid turnover of the AMP-adenosine metabolic cycle in the guinea pig heart.

Abstract

The intracellular flux rate through adenosine kinase (adenosine-AMP) in the well-oxygenated heart was investigated, and the relation of the AMP-adenosine metabolic cycle (AMPw±adenosine) to transmethylation (S-adenosylhomocysteine [SAHI -*adenosine) and coronary flow was determined. Adenosine kinase was blocked in isolated guinea pig hearts by infusion of iodotubercidin in the presence of the adenosine deaminase blocker erythro-9-(2-hydroxy-3-nonyl)adenine (5 ,umol/L).lodotubercidin (1 nmol/L to 4 ,umol/L) caused graded increases in venous effluent concentrations of adenosine, from 8±3 to 145±32 nmol/L (mean±SEM, n=3), and in coronary flow, which increased to maximal levels. Flow increases were completely abolished by adenosine deaminase (5 to 10 U/mL). Interstitial adenosine concentrations, estimated using a mathematical model, increased from 22 nmol/L during control conditions to 420 nmol/L during maximal vasodilation. The possibility that iodotubercidin caused increased venous adenosine by interfering with myocardial energy metabolism was ruled out in separate 31P nuclear magnetic resonance experiments. To estimate total normoxic myocardial production of adenosine (AMP-adenosine*-SAH), the time course of coronary venous adenosine release was measured during maximal inhibition of adenosine kinase with 30 ,umol/L iodotubercidin. Adenosine release increased more than 15-fold over baseline, reaching a new steady-state value of 3.4±0.3 nmol* min-' g`-(n=5) after 4 minutes. In parallel experiments, the relative roles of AMP hydrolysis and transmethylation (SAH hydrolysis) were determined, using adenosine dialdehyde (10 ,umol/L) to block SAH hydrolase. In these experiments, adenosine release increased to similar levels of 3.4±0.5 nmol min' * g1 (n=6) during inhibition of adenosine deaminase and adenosine kinase. It is concluded that (1) maximal increases in coronary flow are elicited by increases in interstitial adenosine concentration to approximately 400 nmol/L, (2) more than 90% of the adenosine produced in the heart is normally rephosphorylated to AMP without escaping into the venous effluent, (3) AMP hydrolysis is the dominant pathway for cardiac adenosine production under normoxic conditions, and (4) the high rate of adenosine salvage is due to rapid turnover of a metabolic cycle between AMP and adenosine. Rapid cycling may serve to amplify the relative importance of AMP hydrolysis over transmethylation in controlling cytosolic adenosine concentrations. (Circ Res. 1993;73:846-856.) KEY WoRDs * coronary flow * S-adenosylhomocysteine * iodotubercidin * adenosine kinase T o further understand the role of adenosine as a local humoral agent in the heart, it is necessary to improve the knowledge of the pathways for adenosine metabolism and transport and to define more precisely the coronary sensitivity for endogenous adenosine. In particular, there are questions concerning the mechanisms of myocardial adenosine production during normoxia and the relative rates of venous adenosine efflux and intracellular adenosine...

Quantification of Extracellular and Intracellular Adenosine Production

“…Global production rate would have been underestimated by 31% if inferred from direct measurement of the release rate (1511 pmol ⅐ min Ϫ1 ⅐ g Ϫ1 ). The present estimate of the global production rate is 67% of those reported before, 5,6 which did not take the effects of increased substrate concentrations into account.…”

“…For measurement of cardiac adenosine and inosine, release of the coronary effluent perfusate was collected over 1 to 2 minutes. Individual pathways of adenosine metabolism or membrane transport were inhibited by use of 5 mol/L erythro-9-hydroxy-nonyl-adenine (EHNA) for adenosine deaminase, 10 mol/L iodotubericidine (ITU) for adenosine kinase, 6,11 and 1 mol/L nitrobenzylthioinosine (NBTI), dipyridamole (DIP), or draflazine (DRA) for membrane transport. 12 Blockers dissolved in physiological saline (except NBTI, which was dissolved in dimethylsulfoxide) were infused at rates of 0.1% (NBTI) or 1% (all others) of coronary flow.…”

“…To test whether coronary flow changes were induced by adenosine, purified 6 adenosine deaminase from calf intestinal mucosa (Boehringer Mannheim) was used. In control experiments, adenosine deaminase was infused during coronary dilation with bradykinin 0.1 mol/L.…”

“…1 However, because of rapid metabolism 2 and a high degree of compartmentalization, 3,4 the precise interstitial adenosine concentration that evokes these effects is unknown. Traditionally, it is assumed that the major site of adenosine production is the cytosol, from which the nucleoside diffuses into the interstitial space to finally reach the vascular region, from which a small fraction 5,6 is washed out with the coronary flow. This concept, however, cannot explain the effects of membrane transport blockers, which increase the vascular adenosine concentration.…”

Compromised Energetics in the Adenylate Kinase AK1Gene Knockout Heart under Metabolic Stress

“…In addition to an energetic function, AK has a distinct signaling role through generation of AMP and activation of AMPdependent processes (9,38,39), including opening of ATPsensitive potassium (K ATP ) channels (7) and adenosine production (40,41). This is of significance in view of the role that these AK-catalysis dependent events play in protecting the myocardium under hypoxic insult (37).…”