Role Of Adenosine In Regulation Research Articles

The Role of Adenosine in the Regulation of Sleep

This paper presents an overview of the current knowledge about the role of adenosine in the sleep-wake regulation with a focus on adenosine in the central nervous system, regulation of adenosine levels, adenosine receptors, and manipulations of the adenosine system by the use of pharmacological and molecular biological tools. The endogenous somnogen prostaglandin (PG) D(2) increases the extracellular level of adenosine under the subarachnoid space of the basal forebrain and promotes physiological sleep. Adenosine is neither stored nor released as a classical neurotransmitter and is thought to be formed inside cells or on their surface, mostly by breakdown of adenine nucleotides. The extracellular concentration of adenosine increases in the cortex and basal forebrain during prolonged wakefulness and decreases during the sleep recovery period. Therefore, adenosine is proposed to act as a homeostatic regulator of sleep and to be a link between the humoral and neural mechanisms of sleep-wake regulation. Both the adenosine A(1) receptor (A(1)R) and A(2A)R are involved in sleep induction. The A(2A)R plays a predominant role in the somnogenic effects of PGD(2). By use of gene-manipulated mice, the arousal effect of caffeine was shown to be dependent on the A(2A)R. On the other hand, inhibition of wake-promoting neurons via the A(1)R also mediates the sleep-inducing effects of adenosine, whereas activation of A(1)R in the lateral preoptic area induces wakefulness, suggesting that A(1)R regulates the sleep-wake cycle in a site-dependent manner. The potential therapeutic applications of agonists and antagonists of these receptors in sleep disorders are briefly discussed.

Adenosine regulation of microtubule dynamics in cardiac hypertrophy

There is evidence that endogenous extracellular adenosine reduces cardiac hypertrophy and heart failure in mice subjected to chronic pressure overload, but the mechanism by which adenosine exerts these protective effects is unknown. Here, we identified a novel role for adenosine in regulation of the cardiac microtubule cytoskeleton that may contribute to its beneficial effects in the overloaded heart. In neonatal cardiomyocytes, phenylephrine promoted hypertrophy and reorganization of the cytoskeleton, which included accumulation of sarcomeric proteins, microtubules, and desmin. Treatment with adenosine or the stable adenosine analog 2-chloroadenosine, which decreased hypertrophy, specifically reduced accumulation of microtubules. In hypertrophied cardiomyocytes, 2-chloroadenosine or adenosine treatment preferentially targeted stabilized microtubules (containing detyrosinated alpha-tubulin). Consistent with a role for endogenous adenosine in reducing microtubule stability, levels of detyrosinated microtubules were elevated in hearts of CD73 knockout mice (deficient in extracellular adenosine production) compared with wild-type mice (195%, P < 0.05). In response to aortic banding, microtubules increased in hearts of wild-type mice; this increase was exaggerated in CD73 knockout mice, with significantly greater amounts of tubulin partitioning into the cold-stable Triton-insoluble fractions. The levels of this stable cytoskeletal fraction of tubulin correlated strongly with the degree of heart failure. In agreement with a role for microtubule stabilization in promoting cardiac dysfunction, colchicine treatment of aortic-banded mice reduced hypertrophy and improved cardiac function compared with saline-treated controls. These results indicate that microtubules contribute to cardiac dysfunction and identify, for the first time, a role for adenosine in regulating cardiomyocyte microtubule dynamics.

Vasodilator effects of adenosine on retinal arterioles in streptozotocin-induced diabetic rats

Adenosine is a potent vasodilator of retinal blood vessels and is implicated to be a major regulator of retinal blood flow during metabolic stress, but little is known about the impact of diabetes on the role of adenosine in regulation of retinal hemodynamics. Therefore, we examined how diabetes affects adenosine-induced vasodilation of retinal arterioles. Male Wistar rats were treated with streptozotocin (80 mg/kg, intraperitoneally), and experiments were performed 6-8 weeks later. Rats were treated with tetrodotoxin (50 microg/kg, intravenously [i.v.]) to eliminate any nerve activity and prevent movement of the eye and infused with methoxamine continuously to maintain adequate systemic circulation. Fundus images were captured with a digital camera that was equipped with a special objective lens, and diameters of retinal arterioles were measured. Adenosine increased diameters of retinal arterioles and decreased systemic blood pressure. These responses were significantly attenuated by the nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (30 mg/kg, i.v.) and the adenosine triphosphate-dependent K+ (K(ATP)) channel blocker glibenclamide (20 mg/kg, i.v.). The depressor responses to adenosine were reduced in diabetic rats, whereas diabetes did not alter vasodilation of retinal arterioles to adenosine. In contrast, both depressor response and vasodilation of retinal arteriole to acetylcholine were reduced in diabetic rats. The retinal vasodilator responses to adenosine and acetylcholine observed in diabetic rats were diminished by N(G)-nitro-L-arginine methyl ester. There were no differences in the responses to pinacidil, a K(ATP) channel opener, between the diabetic and nondiabetic rats. These results suggest that both the activation of nitric oxide synthase and opening of K(ATP) channels contribute to the vasodilator effects of adenosine in rats in vivo. However, diabetes has no significant impact on the vasodilation mediated by these mechanisms in the retinal circulation.

Adenosine regulation of alveolar fluid clearance

Adenosine is a purine nucleoside that regulates cell function through G protein-coupled receptors that activate or inhibit adenylyl cyclase. Based on the understanding that cAMP regulates alveolar epithelial active Na(+) transport, we hypothesized that adenosine and its receptors have the potential to regulate alveolar ion transport and airspace fluid content. Herein, we report that type 1 (A(1)R), 2a (A(2a)R), 2b (A(2b)R), and 3 (A(3)R) adenosine receptors are present in rat and mouse lungs and alveolar type 1 and 2 epithelial cells (AT1 and AT2). Rat AT2 cells generated and produced cAMP in response to adenosine, and micromolar concentrations of adenosine were measured in bronchoalveolar lavage fluid from mice. Ussing chamber studies of rat AT2 cells indicated that adenosine affects ion transport through engagement of A(1)R, A(2a)R, and/or A(3)R through a mechanism that increases CFTR and amiloride-sensitive channel function. Intratracheal instillation of low concentrations of adenosine (< or =10(-8)M) or either A(2a)R- or A(3)R-specific agonists increased alveolar fluid clearance (AFC), whereas physiologic concentrations of adenosine (> or =10(-6)M) reduced AFC in mice and rats via an A(1)R-dependent pathway. Instillation of a CFTR inhibitor (CFTR(inh-172)) attenuated adenosine-mediated down-regulation of AFC, suggesting that adenosine causes Cl(-) efflux by means of CFTR. These studies report a role for adenosine in regulation of alveolar ion transport and fluid clearance. These findings suggest that physiologic concentrations of adenosine allow the alveolar epithelium to counterbalance active Na(+) absorption with Cl(-) efflux through engagement of the A(1)R and raise the possibility that adenosine receptor ligands can be used to treat pulmonary edema.

Adenosine potentiates stimulatory effects on granulocyte-macrophage hematopoietic progenitor cells in vitro of IL-3 and SCF, but not those of G-CSF, GM-CSF and IL-11

The aim of the studies was to ascertain if adenosine is able to co-operate with selected hematopoietic growth factors and cytokines, namely with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), interleukin-3 (IL-3), and interleukin-11 (IL-11), in inducing the growth of colonies from hematopoietic progenitor cells for granulocytes and macrophages (GM-CFC) from normal bone marrow cells in vitro. Adenosine was found not to produce any colonies when present in the cultures as the only potential stimulator. All the tested cytokines and growth factors were observed to induce the growth of distinct numbers of GM-CFC colonies, with the exception of IL-11. When suboptimal concentrations of the evaluated cytokines and growth factors were tested in the cultures in which various concentrations of adenosine were concomitantly present, mutually potentiating effects were found in the case of IL-3 and SCF. These results confirm the role of adenosine in regulation of granulopoiesis and predict IL-3 and SCF as candidates for further in vivo studies of their combined administration with adenosine.

Adenosine

Adenosine has been recognized as a potentially important signaling molecule in the heart for nearly a half century. The original adenosine hypothesis proposed that production of adenosine by cardiac myocytes reflects the metabolic state of the myocardium and serves to regulate vasomotor tone in the coronary resistance vessels, thereby coupling blood flow to the energetic needs of the heart.1 There are two major pathways for adenosine production in the cardiac myocyte. The transmethylation pathway involves the hydrolysis of S -adenosylhomocysteine (SAH) by SAH hydrolase to l-homocysteine and adenosine. A second pathway, the hydrolysis of AMP by 5′-nucleotidase, predominates during ischemic or hypoxic conditions.2 According to the adenosine hypothesis, when the energy requirements of the heart increase, the increased rate of ATP hydrolysis would cause cytosolic free ADP levels to rise. In this situation, the myokinase reaction can utilize two molecules of ADP to produce one molecule each of ATP and AMP, the later being the substrate for 5′-nucleotidase to produce adenosine, which can enter the interstitial space to cause coronary vasodilation. Although this hypothesis for preserving the oxygen supply/demand relationship is attractive, it does not appear to operate during physiological conditions in the normal heart, since adenosine receptor inhibition does not decrease coronary blood flow or impair the increase of coronary flow during exercise.3 Implicit in the adenosine hypothesis is the assumption that during increased cardiac work, increases of cytosolic free [ADP] would be required to drive mitochondrial respiration and would serve to increase adenosine production. In fact, over a wide range of workloads, cytosolic free [ADP] remains at a stable low level; only at extreme workloads does ADP rise significantly in the normal heart.4 Although it has been difficult to demonstrate adenosine effects during normal physiological conditions, adenosine is clearly able to exert important effects …

Role of adenosine in regulation of uterine blood flow during nonpreeclamptic twin gestation.

We evaluated the possible role of increased adenosine in the regulation of uteroplacental circulation during nonpreeclamptic twin gestations. We performed uterine artery Doppler velocimetry and maternal arterial blood sampling for measurement of plasma adenosine levels in 16 women with singleton gestation and 14 women with dichorionic twin gestation without preeclampsia at 11-12, 25-26 and 34-36 weeks. In singleton gestations, a negative correlation between maternal plasma adenosine levels and maternal uterine artery pulsatility index was observed. In twin gestations, there were no noticeable association between maternal plasma adenosine levels and maternal uterine artery pulsatility index. Our results suggest that the decrease in vascular resistance in twin gestations may be through the mechanisms that are independent of adenosine.

The role of adenosine in regulation of cerebral blood flow during hypoxia in the near-term fetal sheep.

The aim of this study was to determine in the near-term ovine fetus the role of adenosine in the basal regulation of cerebral blood flow and in the increases in cerebral blood flow in response to acute hypoxic insult. We measured cerebral blood flow in chronically instrumented fetal sheep (127-135 days gestation, term approximately 145 days) using laser Doppler flowmetry probes implanted in the parietal cortices. Hypoxia was administered for 30 min by lowering the ewe's inspired oxygen to 10-12 % during an infusion of either saline or theophylline, a non-specific adenosine receptor antagonist. The theophylline infusion was begun 30 min prior to and ended 30 min after the completion of the hypoxic insult. The administration of theophylline had no significant effect on cerebral blood flow during the baseline period. During control hypoxic periods, cerebral blood flow increased by approximately 45 %. During theophylline experiments, however, there was no significant increase in cerebral blood flow during hypoxia. In the control experiments, cerebral blood flow returned to baseline levels during the recovery period, while in the theophylline experiments cerebral blood flow fell below baseline levels. We conclude that, in the near-term ovine fetus, adenosine plays a minimal role in the regulation of basal cerebral blood flow. However, these data are strong evidence for the involvement of adenosine in increased fetal cerebral blood flow during an acute hypoxic insult. Finally, adenosine may also play an important role in the maintenance of fetal cerebral blood flow immediately following hypoxic insult.

Time of day determines modulation of synaptic transmission by adenosine in the rat hippocampal slices.

Adenosine, an endogenous modulator of synaptic transmission, has been implicated in regulation of sleep and arousal. The effect of adenosine on neuronal excitability depends on its concentration in the extracellular space. The present study shows that the state of activity of laboratory rats determines the level of tonic inhibition by adenosine in hippocampal slices prepared from these animals. Thus, slices taken at the end of the active period showed significantly more inhibition by adenosine, as determined by the effects of the A1 receptor blocker 8-CPT, in comparison to slices taken in the inactive state. The results support the proposed role of adenosine in regulation of sleep and arousal and point to the importance of the time of day at which slices for electrophysiological experiments are prepared.

Role of adenosine in regulation of brain temperature in fetal sheep

Objective: The purpose of this study was to test adenosine’s possible suppression of heat production by the ovine fetal brain during acute hypoxia. Study Design: Hypoxia was induced by occluding the umbilical cord for 5 minutes in 8 fetal sheep with and without an adenosine receptor blocker, theophylline, and fetal brain and core temperatures were recorded. Results: In 8 untreated fetuses, cord occlusion induced severe but reversible hypoxemia (decrease in carotid arterial P O 2 from 23 ± 1 to 5 ± 1 mm Hg ( P < .01), a 2.3-fold increase in plasma adenosine, and an increase in body core temperature of 0.19°C ± 0.03°C ( P < .01), yet brain temperature remained constant. However, after cord occlusion with prior and continuing administration of theophylline, brain temperature did not increase as hypothesized but rather decreased, suggesting marked reductions in cerebral metabolic rate. This response occurred despite similar degrees of hypoxemia and similar increases in plasma adenosine and body core temperature. Conclusion: We conclude that the temperature of the fetal ovine brain remains constant or decreases during severe reversible hypoxemia by mechanisms other than those dependent on adenosine. (Am J Obstet Gynecol 1999;181:681-7.)

Role of adenosine in regulation of carbohydrate metabolism in contracting muscle.

Adenosine production from AMP in the sarcoplasm and interstitial space of muscle is markedly enhanced during contractions. The produced adenosine may act as a 'local hormone' by binding to various types of adenosine receptors present in the membrane of adjacent cells, including skeletal muscle, vascular smooth muscle and neurons. Thus, interstitial adenosine may significantly contribute to regulation of muscle carbohydrate metabolism, both by adjusting metabolism and local blood flow to the energy needs imposed by a given degree of contratile activity on the muscle cell. The studies presented here demonstrate that endogenous adenosine via A1-adenosine receptors is able to directly stimulate insulin-mediated glucose transport in oxidative muscle cells during contractions. In addition, adenosine may further contribute to stimulation of muscle glucose uptake during contractions by increasing blood flow and thereby targetting glucose and insulin delivery to active muscle fibres. Furthermore, our findings demonstrate that adenosine via A1- and A2-receptors may inhibit glycogen breakdown in oxidative muscle tissue which during contractions is simultaneously exposed to insulin and beta-adrenergic stimulation. It is concluded that adenosine importantly contributes to regulation of carbohydrate metabolism in oxidative muscle fibers during contractions.

Role of adenosine in regulation of coronary flow in dogs with inhibited synthesis of endothelium-derived nitric oxide.

Endothelium-derived nitric oxide (NO) regulates coronary blood flow, but it is unclear how NO synthesis inhibition affects myocardial metabolism. In pentobarbital sodium-anesthetized dogs, myocardial oxygen metabolism, adenosine release, lactate extraction rate (LER), and systolic ventricular wall thickening (SWT) at baseline and during atrial pacing were estimated before and after intracoronary NG-nitro-L-arginine methyl ester (L-NAME) infusion. Coronary blood flow and PO2 in the anterior interventricular vein at baseline were both significantly decreased by L-NAME (3 x 10(-4) M in the coronary blood). Coronary flow was increased during pacing, which was not affected by L-NAME. Myocardial adenosine release remained unchanged during pacing before L-NAME, but it was significantly increased after L-NAME infusion. Neither LER nor SWT changed during pacing performed before and after L-NAME. The experiment was also performed in dogs pretreated with 8-phenyltheophyl-line. After L-NAME, pacing-induced increase in coronary flow was suppressed, and both LER and SWT were significantly decreased during pacing. In conclusion, when NO synthesis is inhibited, adenosine release is increased in response to the increase in myocardial oxygen demand. With this compensatory adenosine release, coronary flow is increased and ventricular function is unaffected.

P-919 - Antioliguric action of FK352, a novel adenosine A1 receptor antagonist, in dogs - role of adenosine in regulation of renal function in the endotoxin shock

P-919 - Antioliguric action of FK352, a novel adenosine A1 receptor antagonist, in dogs - role of adenosine in regulation of renal function in the endotoxin shock

A role for adenosine in coronary vasoregulation in man. Effects of theophylline and enprofylline

Adenosine has been suggested to have a role in regulation of the tone of the cardiac resistance vessels. To elucidate the coronary vasoregulatory role of endogenous adenosine in man, we studied the effects of adenosine receptor antagonism by theophylline on coronary blood flow at rest and during light exercise. However, theophylline may also exert pharmacological effects not related to adenosine antagonism. To clarify the contribution of endogenous adenosine in coronary hyperaemia, the effect of theophylline was compared to that of enprofylline, a xanthine which exerts similar pharmacological effects as theophylline while lacking antagonistic action at adenosine receptors. Twenty healthy subjects (10 males) aged 22-39 years were examined. Coronary sinus (CS) blood flow and blood oxygen content were determined at rest and during supine bicycle exercise, at a load of 50 watts, for 10 min. Thereafter, stepwise infusion of adenosine (30 to 60 micrograms/kg/min into the subclavian vein) was performed. Theophylline or enprofylline treatment was instituted randomly and double-blind (10 in each group), and the procedures (i.e. determinations at rest, during exercise and during infusion of adenosine) were repeated. In all 20 subjects, basal CS flow was 70 +/- 6 ml/min and the cardiac oxygen extraction ((A-CS)O2D) was 123 +/- 3 ml/l. During exercise, CS flow and (A-CS)O2D increased to 135 +/- 17 ml/min and 132 +/- 3 ml/l, respectively. Adenosine increased CS flow dose dependently to 161 +/- 27 ml/min, while (A-CS)O2D decreased to 66 +/- 7 ml/l. The vasodilatory effect of adenosine was readily counteracted by theophylline, the increase in CS flow being 33% vs. 133% in the control situation. Enprofylline, on the other hand, enhanced the response to exogenous adenosine. Theophylline, at a dose lacking effect on heart rate and blood pressure, decreased CS flow at rest by 14% (P < 0.05) and during exercise by 18% (P < 0.05). ((A-CS)O2D increased by 14% at rest and during exercise (P < 0.001). Enprofylline, on the other hand, was without effect. The differences in responses between theophylline and enprofylline with respect to coronary flow and oxygen extraction were significant both at rest and during exercise. It is concluded that theophylline increases coronary vascular resistance, while enprofylline, lacking adenosine antagonistic properties, was without such effect. This indicates a physiological role of adenosine in regulation of coronary flow.

Role of adenosine in regulation of regional cerebral blood flow in sensory cortex

We have previously demonstrated that rat pial arterioles located on the somatosensory cortex dilated in response to contralateral sciatic nerve stimulation (SNS). We hypothesized that the vasodilation was mediated by adenosine, released as a result of somatosensory cortex activation. To test this hypothesis, we examined the effects of SNS (0.15-0.2 V, 5 ms, 5 Hz for 20 s) on pial arterioles under conditions of altered adenosine availability. Cerebrospinal fluid (CSF) adenosine was altered by perfusing mock CSF, under a cranial window in anesthetized rats, containing either an adenosine uptake competitor (dipyridamole or inosine) or an adenosine receptor blocker (theophylline). With CSF only, SNS caused pial arterioles (resting diam, 29 +/- 1 micron) to dilate by 38 +/- 10% (peak magnitude) for 32 +/- 2 s. Dipyridamole (10(-6) M) significantly (P less than 0.02) enhanced both the magnitude (to 62 +/- 12%) and duration (to 68 +/- 10 s) of the response. Similarly, inosine (10(-3) M) significantly (P less than 0.02) potentiated the vasodilative response from resting values of 27 +/- 5% and 34.8 +/- 4.1 s to 37 +/- 6% and 89.6 +/- 14.1 s. In contrast, theophylline (5 x 10(-5) M) significantly (P less than 0.001) attenuated arteriolar vasodilation from resting values of 38 +/- 5% and 29.3 +/- 1.2 s to 18 +/- 3% and 22.0 +/- 0.9 s. Neither dipyridamole nor theophylline had a significant effect on neuronal response (sensory-evoked response) recorded from the somatosensory cortex. These results suggest that adenosine is involved in the regulation of pial vasodilation during cerebral cortical activation.

S-40 - Role of adenosine in regulation of mesenteric arterial blood flow

S-40 - Role of adenosine in regulation of mesenteric arterial blood flow

Ecto-5'-nucleotidase: localization in rat kidney by light microscopic histochemical and immunohistochemical methods.

We demonstrated the distribution pattern of ecto-5'-nucleotidase (5'-Nu) in rat kidney by enzymatic activity (lead salt precipitation) and by immunohistochemistry with a polyclonal antibody raised in rabbits. Enzyme activity was found in the brush border of the proximal tubule, highest in the P1 segments with decreasing intensity in the P2 segments and weakest in P3 segments in the medullary rays of the cortex. The P3 segments of the outer stripe showed slightly higher activity. Activity was also apparent in the intercalated cells in the connecting tubule and collecting duct, whereas all other tubular and glomerular structures were negative. Activity in peritubular and perivascular connective tissue was highest in the cortical labyrinth, weak or absent in the medullary rays of the cortex, and entirely absent in the medulla. The distribution of the antigen was fully congruent with that of the enzyme activity. With respect to the role of adenosine in regulation of renal blood flow and glomerular filtration rate, the distribution of 5'-Nu in the cortical interstitium may be particularly significant. The possibility of nucleotide cleavage at the brush-border membranes may be important for salvage of nucleotides from the tubular lumen.

Role of adenosine in regulation of cerebral blood flow: effects of theophylline during normoxia and hypoxia.

We studied the effects of the methylxanthine theophylline, an adenosine receptor blocker, on cerebral circulation. Cerebral blood flow (CBF) was measured by the retroglenoid outflow and microsphere techniques, and pial circulation changes were observed through a closed cranial window. Intraperitoneal administration of theophylline in normoxic animals resulted in a biphasic response of pial vessels and CBF. At low concentrations (0.05 mumol/g) of theophylline, pial vessel diameter and CBF decreased, whereas vasodilatation and hyperemia were observed at higher levels. After intraperitoneal administration of either 0.05 or 0.2 mumol/g, hypoxic hyperemia was attenuated both during short (c. 30 s) and sustained (c. 2-3 min) hypoxia, as was hypoxic pial arteriolar vasodilatation. These actions of theophylline appear to be due to adenosine receptor blockade, since micromolar concentrations were achieved in cerebrospinal fluid (CSF), and no increases in adenosine 3',5'-cyclic monophosphate concentrations in brain were noted. Moreover, theophylline (either intraperitoneal or topical) blocked pial vasodilatation caused by topically applied adenosine, but had little effect on hypercarbic hyperemia or pial vasodilatation induced by topically applied acetylcholine. The results of these studies suggest that adenosine is involved in the maintenance of resting cerebral vascular tone and has a paramount role in the regulation of CBF during hypoxia.