We conclude that adenosine released during the preconditioning occlusion stimulates cardiac A1 receptors, which leaves the heart protected against infarction even after the adenosine has been withdrawn.

BACKGROUND Recent data from this laboratory indicate that pretreatment with adenosine can protect the heart against infarction via A1-receptors, but because of systemic hypotension, adenosine had to be given into the coronary circulation. METHODS AND RESULTS In this study, we tested whether the protection could be achieved by intravenous administration of the A1-selective adenosine agonists N6-(phenyl-2R-isopropyl)-adenosine (PIA) and 2-chloro-N6-cyclopentyladenosine (CCPA). Nine groups of open-chest anesthetized rabbits were subjected to 30 minutes of regional coronary ischemia and 3 hours of reperfusion. Infarct size was determined by tetrazolium staining. Control hearts receiving no treatment had 38 +/- 4% of the risk zone infarcted. Preconditioning with 5 minutes of ischemia and 10 minutes of reperfusion before ischemia limited the infarct to 8 +/- 4%. Intravenous PIA 15 minutes before 30-minute ischemia also limited infarct size to 6 +/- 2% at the highest dose. CCPA offered similar protection. When the PIA was given at reperfusion, infarct size was 46 +/- 6%, indicating that receptor activation must precede ischemia to protect. Pretreatment with CGS 21680, a selective A2-receptor agonist, caused identical hypotension but failed to limit infarct size (43 +/- 3%), indicating again that the A1-receptor is involved. When rabbits pretreated with PIA were paced at 220 beats per minutes, PIA still limited infarct size (16 +/- 4%), indicating that protection was not the result of bradycardia. CONCLUSIONS These results indicate that stimulation of adenosine A1-receptors causes the heart to become resistant to ischemia and that this protection can be achieved with intravenous administration of A1-selective agents.

We have examined the effects of adenosine on sodium-coupled phosphate and glucose transport in cultured opossum kidney (OK) cells, a continuous cell line that resembles proximal tubule. Adenosine analogues R-(-)-N6-phenylisopropyladenosine (PIA) and 2',5'-dideoxy-adenosine (DDA) were employed as adenosine A1 receptor and P site-selective agonists, respectively. Sodium-dependent phosphate uptake activity (Na-Pi symport) increased by approximately 25% above both basal and parathyroid hormone (PTH)-inhibited levels in cells treated with PIA (0.1, 1 microM) but not in cells treated with DDA (100 microM). Adenosine (PIA) also stimulated sodium-coupled 3-O-methylglucose transport by approximately 40%. Intracellular adenosine 3',5'-cyclic monophosphate (cAMP) content was inversely related to Na-Pi symport activity in cells treated with PIA and PTH. However, changes in Na-Pi symport activity did not consistently relate to changes in intracellular cAMP. Protein kinase C was activated 15 s after treatment of OK cells with 1 microM PIA. Preincubation of cells with 3 microM staurosporine attenuated the effect of 1 microM PIA on phosphate uptake. These data suggest that Na-Pi and Na-glucose symport activities are stimulated by adenosine acting at a receptor coupled to more than one intracellular signal. It is likely that both protein kinases A and C are involved in these actions of adenosine.

Adenosine A1 receptor densities were increased in cultured LLC-PK1 and OK cells by chronic treatment with the adenosine receptor antagonists 1,3,7-trimethylxanthine (caffeine, 1 mM) and 1,3-dimethyl-8-cyclopentylxanthine [cyclopentyltheophylline (CPT), < or = 0.4 mM], respectively. The A1 receptor number per cell was increased twofold by 10-day treatments with 1 mM caffeine or 0.1 mM CPT, and the sodium-coupled glucose uptake was augmented twofold by 1 mM caffeine and sevenfold by 0.1 microM CPT (higher doses of CPT were progressively less stimulatory). Glucose uptake was blocked by acute (2-h) treatment with CPT, adenosine deaminase, or calphostin C. Caffeine (1 mM) or CPT (> or = 0.1 mM) inhibited cell proliferation for the first 10 days, then cell growth assumed a normal proliferative rate despite continued presence of antagonist. Cytosolic protein kinase C (PKC) beta-isoform immunoactivity and PKC-beta II mRNA were elevated at least twofold during 10 days of 0.1 mM CPT or 1 mM caffeine treatment. The sustained elevation in sodium-glucose symport and PKC activity observed with adenosine receptor antagonists was similar to acute (2-h) effects of the adenosine A1 agonist R(-)-N6-phenylisopropyladenosine (R-PIA, 0.1-1 microM). Moreover, cell proliferation was increased by adenosine (0.1 microM R-PIA), whereas Na-K-adenosinetriphosphatase activity was unaltered with chronic antagonist or acute adenosine treatments. Caffeine treatment may have some non-adenosine A1 receptor-mediated actions, because it slightly (30%) augmented protein kinase A activity. It is concluded that chronic exposure of proximal tubule cells to caffeine or CPT augments PKC and sodium-glucose transport but retards cell proliferation mainly via adenosine A1 receptor-mediated mechanisms.

SUMMARY. We employed intracoronary infusion of adenosine deaminase to test the hypothesis that endogenous adenosine contributes to regulation of coronary blood flow following acute reductions in coronary artery pressure. In 16 closed-chest anesthetized dogs, we perfused the left circumflex coronary artery from a pressurized arterial reservoir and measured coronary blood flow following changes in perfusion pressure before and 10 minutes after the start of intracoronary adenosine deaminase, 5 U/min per kg body weight. Parallel studies showed that this dose of enzyme resulted in cardiac lymph adenosine deaminase concentrations of 3.2 ± 0.4 U/ml. Adenosine deaminase abolished the vasodilator response to intracoronary adenosine, 4 and 8 /tg, but had no effect on the vasodilator response to intracoronary papaverine, 200 and 300 ^g, demonstrating enzyme efficacy and specificity. Additional experiments demonstrated that adenosine deaminase reversibly attenuated myocardial reactive hyperemia following 5-and 10-second coronary occlusions by 30% (P < 0.05), evidence that the infused enzyme effectively degraded endogenous adenosine. However, adenosine deaminase did not alter the time course for coronary autoregulation or the steady state autoregulatory flow response over the pressure range between 125 and 75 mm Hg. Further, adenosine deaminase did not alter steady state coronary flow when perfusion pressure was reduced below the range for effective autoregulation (60-40 mm Hg). Such results show that adenosine is not essential for either coronary autoregulation or for the maintenance of coronary vasodilation when autoregulatory vasodilator reserve is expended. (CircRes 56: 517-524, 1985) CORONARY autoregulation is the intrinsic ability of the heart to maintain its blood supply relatively constant in the face of fluctuations in perfusion pressure (Mosher et al., 1964;Cross, 1964). The mechanism underlying this fundamental adaptive flow response is unknown. According to the metabolic hypothesis, coronary autoregulation is initiated by flow-dependent changes in tissue levels of a metabolic substrate or a metabolite (Johnson, 1964;Rubio and Berne, 1975;Belloni, 1979;Feigl, 1983). Several studies have demonstrated increased adenosine production and release during conditions in which oxygen supply is reduced (Katori and Berne, 1966;Rubio and Berne, 1969;Rubio et al., 1974;. In one of these studies , tissue adenosine content and adenosine release increased when perfusion pressure was lowered over the autoregulation range in isolated buffer-perfused guinea pig hearts. As the investigators point out, such a correlation only suggests, but does not prove, that adenosine is the primary determinant of changes in coronary resistance during autoregulation.This study tests the hypothesis that endogenous adenosine is an essential mediator of coronary autoregulation. The hypothesis predicts that destroying interstitial adenosine should abolish autoregulation. Extending this inquiry to pressures below the autoregulation range tests a secon...