Deoxynucleoside overproduction in deoxyadenosine-resistant, adenosine deaminase-deficient human histiocytic lymphoma cells

Deoxyadenosine toxicity toward lymphocytes may produce immune dysfunction in patients with adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) deficiency. The relationship between endogenous deoxynucleoside synthesis in adenosine deaminase-deficient cells and sensitivity to adenosine and deoxyadenosine toxicity is unclear. The human histiocytic lymphoma cell line (DHL-9) naturally lacks adenosine deaminase, and has minimal levels of thymidine kinase. Dividing DHL-9 cells excrete deoxyadenosine and thymidine into the extracellular space. The present experiments have analyzed nucleoside synthesis and excretion in a mutagenized clone of DHL-9 cells, selected for increased resistance to deoxyadenosine toxicity. The deoxyadenosine-resistant cells excreted both deoxyadenosine and thymidine at a 6-7-fold higher rate than wild-type lymphoma cells. The deoxyadenosine overproduction was accompanied by a reduced ability to form dATP from exogenous deoxyadenosine, and a 2.5-fold increase in ribonucleotide reductase activity. The pace of adenosine excretion, the growth rate, and the levels of multiple other enzymes involved in deoxyadenosine and adenosine metabolism were equivalent in the two cell types. These results suggest that the excretion of deoxyadenosine and thymidine, but not adenosine, is exquisitely sensitive to alterations in the rate of endogenous deoxynucleotide synthesis. Apparently, small changes in deoxynucleotide synthesis can significantly influence cellular sensitivity to deoxyadenosine toxicity.     

Adenosine and the control of lipolysis in rat adipocytes during pregnancy and lactation.

The rate of noradrenaline-stimulated lipolysis is lower in fat-cells from lactating than from pregnant rats; this difference is eliminated by the addition of adenosine deaminase [Aitchison, Clegg & Vernon (1982) Biochem. J. 202, 243-247]. The activity of 5'-nucleotidase, and hence the capacity of the cells to synthesize adenosine, was the same in fat-cells and also stromal cells of adipose tissue from pregnant, lactating and male rats. The response and sensitivity of fat-cells to the anti-lipolytic effects of adenosine were measured by incubating cells in the presence of noradrenaline, adenosine deaminase (to remove endogenous adenosine) and various concentrations of the adenosine analogue N6-phenylisopropyladenosine (PIA). PIA caused a greater inhibition of the rate of noradrenaline-stimulated lipolysis in adipocytes from lactating than from pregnant rats. The concentration of PIA required to inhibit by 50% the rate of noradrenaline-stimulated lipolysis fell from over 100 nM for fat-cells from pregnant rats to 30 nM for fat-cells from lactating rats. The decreased rate of noradrenaline-stimulated lipolysis during lactation was not due to the smaller mean cell volume of adipocytes during this state.     

Autoradiographical Studies of in Vitro and Chronic in Vivo Effects of Propentofylline on Adenosine A1 and A2 Receptors and NBMPR-Sensitive Nucleoside Transporters

Abstract

Propentofylline is a novel xanthine that has been shown to limit the extent neuronal damage induced by cerebral ischemia in gerbils (DeLeo et al., 1987). This is in contrast to other xanthines, including, caffeine and theophylline, that increase the extent of damage (Rudolphi et al., 1987; Dux et al., 1987). Propentofylline has been demonstrated to decrease adenosine uptake into human erythrocytes (Fredholm and Lindström, 1986), and to increase extracellular concentration of adenosine in ischemic barain (Andine et al., 1990). Therefore, it was proposed that this compound provides protection in cerebral ischemia, in part, by adenosine receptor stimulation due to increased endogenous adenosine levels.     

Modulation of catecholamine release by endogenous adenosine in the rat adrenal medulla

Adenosine was shown to inhibit norepinephrine (NE) release from sympathetic nerve endings. The purpose of this study was to examine whether endogenous adenosine restrains NE and epinephrine release from the adrenal medulla. The effects of an adenosine receptor antagonist, 1,3-dipropyl-8-(p-sulfophenyl) xanthine (DPSPX), on epinephrine and NE release induced by intravenous administration of insulin in conscious rats were examined. Plasma catecholamines were measured by HPLC with an electrochemical detector. DPSPX significantly increased plasma catecholamine in both control rats and rats treated with insulin. The effect of DPSPX on plasma catecholamine was significantly greater in rats treated with insulin. Additional experiments were performed in adrenalectomized rats to investigate the contribution of the adrenal medulla to the effect of DPSPX on plasma catecholamine. The effect of DPSPX and insulin on epinephrine in adrenalectomized rats was significantly reduced compared with that of the controls. Finally, we tested whether endogenous adenosine restrains catecholamine secretion partially through inhibiting the renin-angiotensin system. The effect of DPSPX on plasma catecholamine in rats pretreated with captopril (an angiotensin-converting enzyme inhibitor) was reduced. These results demonstrate that under basal physiological conditions, endogenous adenosine tonically inhibits catecholamine secretion from the adrenal medulla, and this effect is augmented when the sympathetic system is stimulated. The effect of endogenous adenosine on catecholamine secretion from the adrenal medulla is achieved partially through the inhibitory effect of adenosine on the renin-angiotensin system.     

Endogenous adenosine inhibits CNS terminal Ca(2+) currents and exocytosis

Bursts of action potentials (APs) are crucial for the release of neurotransmitters from dense core granules. This has been most definitively shown for neuropeptide release in the hypothalamic neurohypophysial system (HNS). Why such bursts are necessary, however, is not well understood. Thus far, biophysical characterization of channels involved in depolarization-secretion coupling cannot completely explain this phenomenon at HNS terminals, so purinergic feedback mechanisms have been proposed. We have previously shown that ATP, acting via P2X receptors, potentiates release from HNS terminals, but that its metabolite adenosine, via A(1) receptors acting on transient Ca(2+) currents, inhibit neuropeptide secretion. We now show that endogenous adenosine levels are sufficient to cause tonic inhibition of transient Ca(2+) currents and of stimulated exocytosis in HNS terminals. Initial non-detectable adenosine levels in the static bath increased to 2.9 microM after 40 min. These terminals exhibit an inhibition (39%) of their transient inward Ca(2+) current in a static bath when compared to a constant perfusion stream. CPT, an A(1) adenosine receptor antagonist, greatly reduced this tonic inhibition. An ecto-ATPase antagonist, ARL-67156, similarly reduced tonic inhibition, but CPT had no further effect, suggesting that endogenous adenosine is due to breakdown of released ATP. Finally, stimulated capacitance changes were greatly enhanced (600%) by adding CPT to the static bath. Thus, endogenous adenosine functions at terminals in a negative-feedback mechanism and, therefore, could help terminate peptide release by bursts of APs initiated in HNS cell bodies. This could be a general mechanism for controlling transmitter release in these and other CNS terminals.     

Effect of ozone treatment on reactive oxygen species and adenosine production during hepatic ischemia-reperfusion

This study investigates whether ozone could confer protection from hepatic ischemia reperfusion by modifying the accumulation of adenosine and xanthine during ischemia. A significant increase in both adenosine and xanthine accumulation was observed as a consequence of ATP degradation during hepatic ischemia. Adenosine exerts a protective effect on hepatic ischemia reperfusion injury since the elimination of endogenous adenosine accumulation with adenosine deaminase increased the hepatic injury associated with this process. On the other hand, the high xanthine levels observed after ischemia could exert deleterious effects during reperfusion due to reactive oxygen species generation from xanthine oxidase. The administration of allopurinol, an inhibitor of xanthine oxidase, attenuated the increase in reactive oxygen species and transaminase levels observed after hepatic reperfusion. Ozone treatment in liver maintained adenosine levels similar to those found after ischemia but led to a marked reduction in xanthine accumulation. In order to evaluate the role of both adenosine and xanthine, we tried to modify the protection confered by ozone, by modifying the concentrations of adenosine and xanthine. The metabolization of endogenous adenosine after ischemia abolished the protective effect conferred by ozone. When xanthine was administered previous to ozone treatment, the protection conferred by adenosine disappeared, showing both postischemic reactive oxygen species and transaminase levels similar to those found after hepatic ischemia reperfusion. Ozone would confer protection against the hepatic ischemia reperfusion injury by the accumulation of adenosine that in turns benefits the liver and by blocking the xanthine/xanthine oxidase pathway for reactive oxygen species generation.     

Effect of ozone treatment on reactive oxygen species and adenosine production during hepatic ischemia-reperfusion

This study investigates whether ozone could confer protection from hepatic ischemia reperfusion by modifying the accumulation of adenosine and xanthine during ischemia. A significant increase in both adenosine and xanthine accumulation was observed as a consequence of ATP degradation during hepatic ischemia. Adenosine exerts a protective effect on hepatic ischemia reperfusion injury since the elimination of endogenous adenosine accumulation with adenosine deaminase increased the hepatic injury associated with this process. On the other hand, the high xanthine levels observed after ischemia could exert deleterious effects during reperfusion due to reactive oxygen species generation from xanthine oxidase. The administration of allopurinol, an inhibitor of xanthine oxidase, attenuated the increase in reactive oxygen species and transaminase levels observed after hepatic reperfusion. Ozone treatment in liver maintained adenosine levels similar to those found after ischemia but led to a marked reduction in xanthine accumulation. In order to evaluate the role of both adenosine and xanthine, we tried to modify the protection confered by ozone, by modifying the concentrations of adenosine and xanthine. The metabolization of endogenous adenosine after ischemia abolished the protective effect conferred by ozone. When xanthine was administered previous to ozone treatment, the protection conferred by adenosine disappeared, showing both postischemic reactive oxygen species and transaminase levels similar to those found after hepatic ischemia reperfusion. Ozone would confer protection against the hepatic ischemia reperfusion injury by the accumulation of adenosine that in turns benefits the liver and by blocking the xanthine/xanthine oxidase pathway for reactive oxygen species generation.     

Adenosine Inhibition of Calmodulin-Sensitive Adenylate Cyclase from Bovine Cerebral Cortex

Calmodulin (CaM)-sensitive adenylate cyclase has recently been purified extensively from bovine brain. In this study, the sensitivity of the CaM-sensitive adenylate cyclase to adenosine and adenosine analogs was examined. The highly purified enzyme preparation retained sensitivity to inhibition by adenosine and adenosine analogs with ribose ring modifications, but not to those with purine ring modifications. Adenosine inhibition of this enzyme was not dependent on GTP and was noncompetitive with respect to ATP. Enzyme that had been dissociated from functional guanine nucleotide binding protein interactions by gel filtration in the presence of the zwitterionic detergent 3-[3-(cholamidopropyl)-dimethylammonio]-propanesulfonate and Mn2+ retained sensitivity to adenosine inhibition. The Ki for adenosine inhibition of the CaM-sensitive adenylate cyclase was approximately 2.6 X 10(-4) M. 5'-Guanylylimidodiphosphate and CaM did not affect the Ki of 3'-deoxyadenosine for the enzyme, but the presence of Ca2+ in the millimolar range raised the Ki by a factor of 5. These results show that the CaM-sensitive form of adenylate cyclase from bovine brain is subject to adenosine inhibition, and strongly suggest that this inhibition is due to interaction of ligands with a purine-specific ("P") site located on the catalytic subunit of the enzyme.     

In vivo evidence against a role for adenosine in the exercise pressor reflex in humans.

The pressor response to exercise is of great importance in both physiology and pathophysiology. Whether endogenous adenosine is a trigger for this reflex remains controversial. Muscle interstitial adenosine concentration can be determined by microdialysis. However, there are indications that local muscle cell damage by the microdialysis probe confounds these measurements in exercising muscle. Therefore, we used the nucleoside uptake inhibitor dipyridamole as pharmacological tool to bypass this confounding. We used microdialysis probes to measure endogenous adenosine in forearm skeletal muscle of healthy volunteers during two cycles of 15 min of intermittent isometric handgripping. During the second contraction, dipyridamole (12 microg.min(-1).dl forearm(-1)) was administered into the brachial artery. Dipyridamole potentiated the exercise-induced increase in dialysate adenosine from 0.30 +/- 0.08 to 0.48 +/- 0.10 micromol/l (n = 9, P < 0.05), but it did not potentiate the exercise-induced increase in blood pressure. A time-control study without dipyridamole revealed no difference in exercise-induced increase in adenosine between both contractions (n = 8). To exclude the possibility that the dipyridamole-induced increase in dialysate adenosine originates from extravasation of increased circulating adenosine, we simultaneously measured adenosine with microdialysis probes in forearm muscle and antecubital vein. In a separate group of nine volunteers, simultaneous intrabrachial infusion of 100 microg.min(-1).dl(-1) dipyridamole and 5 microg.min(-1).dl(-1) adenosine increased dialysate adenosine from the intravenous but not the interstitial probe, indicating preserved endothelial barrier function for adenosine. We conclude that dipyridamole significantly inhibits uptake of interstitial adenosine without affecting the pressor response to exercise, suggesting that interstitial adenosine is not involved in the pressor response to rhythmic isometric exercise.     

Rat adipocyte lipoprotein lipase activity is inhibited by theophylline and stimulated by inosine through adenosine-independent mechanisms

Incubation of rat adipose tissue or isolated rat adipocytes with high (50 mM) but not with low concentrations (0.5 mM) of theophylline results in a decrease of lipoprotein lipase (LPL) activity. This effect is not altered by the addition of adenosine deaminase, indicating that the decrease of adipose LPL activity by theophylline is not due to the competition of theophylline with adenosine. On the contrary, incubation of isolated fat cells with adenosine (0.1 – 100 μM) results in an increase of the intracellular form of LPL activity. As this effect is also observed in cells incubated with adenosine deaminase (40 mU/ml) or with inosine (0.1 – 100 μM) but not in cells incubated with the adenosine analog N⁶-phenylisopropyladenosine, it is concluded that the increase in the intracellular form of LPL found after incubation with adenosine is not due to adenosine $\text{per se}$ but to inosine generated from the breakdown of endogenous adenosine by adenosine deaminase.     

Interactions between adenosine and oscillatory cAMP signaling regulate size and pattern in Dictyostelium

We present evidence for the hypothesis that in multicellular structures of Dictyostelium, production of adenosine by hydrolysis of cAMP near the tip region prevents both generation of competing tips and differentiation of prespore cells near the tip, and thus establishes a "prestalk" region. We demonstrate that adenosine affects the immunological prespore specific staining pattern in slugs in a manner opposite to cAMP:cAMP induces an increase of prespore antigen; adenosine induces a decrease. When endogenous adenosine is removed from slugs, prespore vacuoles are synthesized throughout the prestalk region. Adenosine was found to inhibit the induction of prespore differentiation by cAMP in an apparently competitive manner. It was also found that adenosine specifically increased the amount of tissue controlled by one tip, probably by inhibiting generation of competing oscillators. Removing endogenous adenosine from slugs resulted in a decrease of tip dominance.     

Adenosine receptor down-regulation and insulin resistance following prolonged incubation of adipocytes with an A1 adenosine receptor agonist

Adenosine, via interaction with A1 adenosine receptors, increases insulin sensitivity and inhibits lipolysis in adipocytes. To investigate regulation of this system, adipocytes were incubated for up to 72 h with the nonmetabolizable adenosine receptor agonist, N6-phenylisopropyl adenosine (PIA). Adenosine receptors were measured by the binding of 125I-hydroxyphenylisopropyl adenosine to membranes. PIA down-regulated adenosine receptors, decreasing the number of binding sites with no change in affinity. Adipocytes were incubated for 48 h without or with 100 nM PIA to down-regulate the A1 receptors by approximately 60%. The cells were washed, and lipolysis and glucose transport were assessed. The ability of PIA to inhibit lipolysis was markedly attenuated in the down-regulated cells. Furthermore, the EC50 of insulin was increased approximately 3-fold in the PIA-treated cells. 125I-Insulin binding to the PIA-treated cells was unchanged, demonstrating that the decreased insulin sensitivity is not due to decreased insulin receptor binding. Pertussis toxin catalyzed ADP-ribosylation of a 41-kDa protein thought to be the alpha-subunit of Gi. This 41-kDa protein was decreased in membranes from cells treated with PIA, with a maximal 50% loss. This suggests that Gi is down-regulated and that loss of both the A1 adenosine receptor and Gi are involved in the metabolic changes observed after PIA treatment.     

IL-4 amplifies the pro-inflammatory effect of adenosine in human mast cells by changing expression levels of adenosine receptors

Adenosine inhalation produces immediate bronchoconstriction in asthmatics but not in normal subjects. The bronchospastic effect of adenosine is largely mediated through adenosine-induced mast cell activation, the mechanism of which is poorly understood due to limitations in culturing human primary mast cells. Here, we show that human umbilical cord blood -derived mast cells incubated with the Th2 cytokine IL-4 develop increased sensitivity to adenosine. Potentiation of anti-IgE- induced and calcium ionophore/PMA-induced degranulation was augmented in mast cells cultured with IL-4, and this effect was reduced or abolished by pre-treatment with A(2B)siRNA and selective A(2B) receptor antagonists, respectively. IL-4 incubation resulted in the increased expression of A(2B) and reduced expression of A(2A) adenosine receptors on human mast cells. These results suggest that Th2 cytokines in the asthmatic lung may alter adenosine receptor expression on airway mast cells to promote increased responsiveness to adenosine.     

Contribution of adenosine to changes in coronary flow in metabolically stimulated rat heart

The extent to which endogenous, extracellular adenosine mediates increased coronary flow in crystalloid-perfused, isovolumic rat hearts stimulated with either norepinephrine or isoproterenol was examined. When infused into the coronary circulation, norepinephrine (1 x 10(-7) M) rapidly increased left ventricular developed pressure (LVDP) from 81 +/- 6 to 235 +/- 13 mmHg (1 mmHg = 133.3 Pa) and coronary flow from 12.7 +/- 0.8 to 18.4 +/- 0.7 mL.min-1.g-1. The presence of either adenosine deaminase (2 U.mL-1) or the adenosine receptor antagonist, 8-phenyltheophylline (5 x 10(-6) M) in the perfusate of norepinephrine-stimulated hearts augmented the increase in LVDP and +/- dP/dtmax by 10-20% but reduced the increase in coronary flow by 34%. Doubling the rate of adenosine deaminase infusion, or infusing the enzyme and 8-phenyltheophylline together did not alter their inhibitory effectiveness. Similar results were observed with hearts stimulated with isoproterenol (5 x 10(-8) M). These data show that about a third of the vasodilation that results from the metabolic stimulation of rat heart by catecholamines is due to the receptor-mediated action of extracellular adenosine.     

Modulation of [3H]Acetylcholine Release from Cultured Amacrine-Like Neurons by Adenosine A1 Receptors

Abstract: We have investigated the effect of endogenous adenosine on the release of [³H]acetylcholine ([³H]ACh) in cultured chick amacrine-like neurons. The release of [³H]ACh evoked by 50 m M KCl was mostly Ca²⁺ dependent, and it was increased in the presence of adenosine deaminase and in the presence of 1,3-dipropyl-8-cyclopentylxanthine (DPCPX), an adenosine A₁ receptor antagonist. The effect of adenosine on [³H]ACh release was sensitive to pertussis toxin (PTX) and was due to a selective inhibition of N-type Ca²⁺ channels. Ligand binding studies using [³H]DPCPX confirmed the presence of adenosine A₁ receptors in the preparation. Using specific inhibitors of the plasma membrane adenosine carriers and of the ectonucleotidases, we found that the extracellular accumulation of adenosine in response to KCl depolarization was due to the release of endogenous adenosine per se and to the extracellular conversion of released nucleotides into adenosine. Activation of adenosine A₁ receptors was without effect on the intracellular levels of cyclic AMP under depolarizing conditions, but it inhibited the accumulation of inositol phosphates. Our results indicate that in cultured amacrine-like neurons, the Ca²⁺-dependent release of [³H]ACh evoked by KCl is under tonic inhibition by adenosine, which activates A₁ receptors. The effect of adenosine on the [³H]ACh release may be due to a direct inhibition of N-type Ca²⁺ channels and/or secondary to the inhibition of phospholipase C and involves the activation of PTX-sensitive G proteins.     

Effect of Acute Ethanol on Release of Endogenous Adenosine from Rat Cerebellar Synaptosomes

The effects of pharmacologically relevant concentrations of ethanol on the release of endogenous adenosine from rat cerebellar synaptosomes were investigated. Release was conducted for 5, 10, 30, or 60 s after which time the incubation medium (containing the released adenosine) was rapidly separated from the synaptosomal membranes by vacuum filtration. The adenosine content of the filtrate was measured by HPLC-fluorescence detection. Both basal and KCl-stimulated adenosine release consisted of an initial rapid phase, for the first 10 s, that was followed by a relatively slower phase. Basal endogenous adenosine release was estimated as 199 +/- 14 pmol/mg protein/5 s. Potassium (chloride) increased adenosine release from the basal level to 433 +/- 83 pmol/mg protein/5 s. Ethanol caused a dose-dependent increase of adenosine release. The interaction between dilazep and ethanol indicates that ethanol-stimulated release does not involve the dilazep-sensitive transport system. The results support previous findings that indicate that cerebellar adenosine is involved in the mediation of ethanol-induced motor disturbances in the rat.     

5'-Adenosine monophosphate and adenosine metabolism, and adenosine responses in mouse, rat and guinea pig heart.

We examined myocardial 5'-adenosine monophosphate (5'-AMP) catabolism, adenosine salvage and adenosine responses in perfused guinea pig, rat and mouse heart. MVO(2) increased from 71+/-8 microl O(2)/min per g in guinea pig to 138+/-17 and 221+/-15 microl O(2)/min per g in rat and mouse. VO(2)/beat was 0.42+/-0.03, 0.50+/-0.03 and 0.55+/-0.04 microl O(2)/g in guinea pig, rat and mouse, respectively. Resting and peak coronary flows were highest in mouse vs. rat and guinea pig, and peak ventricular pressures and Ca(2+) sensitivity declined as heart mass increased. Net myocardial 5'-AMP dephosphorylation increased significantly as mass declined (3.8+/-0.5, 9.0+/-1.4 and 11.0+/-1.6 nmol/min per g in guinea pig, rat and mouse, respectively). Despite increased 5'-AMP catabolism, coronary venous [adenosine] was similar in guinea pig, rat and mouse (45+/-8, 69+/-10 and 57+/-14 nM, respectively). Comparable venous [adenosine] was achieved by increased salvage vs. deamination: 64%, 41% and 39% of adenosine formed was rephosphorylated while 23%, 46%, and 50% was deaminated in mouse, rat and guinea pig, respectively. Moreover, only 35-45% of inosine and its catabolites derive from 5'-AMP (vs. IMP) dephosphorylation in all species. Although post-ischemic purine loss was low in mouse (due to these adaptations), functional tolerance to ischemia decreased with heart mass. Cardiovascular sensitivity to adenosine also differed between species, with A(1) receptor sensitivity being greatest in mouse while A(2) sensitivity was greatest in guinea pig. In summary: (i) cardiac 5'-AMP dephosphorylation, VO(2), contractility and Ca(2+) sensitivity all increase as heart mass falls; (ii) adaptations in adenosine salvage vs. deamination limit purine loss and yield similar adenosine levels across species; (iii) ischemic tolerance declines with heart mass; and (iv) cardiovascular sensitivity to adenosine varies, with increasing A(2) sensitivity relative to A(1) sensitivity in larger hearts.     

Adenosine production and its interaction with protection of ischemic and reperfusion injury of the myocardium

Adenosine exerts cardioprotective effects on the ischemic myocardium. A flexibly mounted microdialysis probe was used to measure the concentration of interstitial adenosine and to assess the activity of ecto-5'-nucleotidase (a key enzyme responsible for adenosine production) in in vivo rat hearts. The level of adenosine during perfusion of adenosine 5'-adenosine monophosphate (AMP) was given as an index of the activity of ecto-5'-nucleotidase in the tissue. Endogenous norepinephrine (NE) activates both alpha(1)-adrenoceptors and protein kinase C (PKC), which, in turn, activates ecto-5'-nucleotidase via phosphorylation thereby enhancing the production of interstitial adenosine. Histamine-release NE activates PKC, which increased ecto-5'-nucleotidase activity and augmented release of adenosine. Opening of cardiac ATP sensitive K(+) (K(ATP)) channels may cause hydroxyl radical (.OH) generation through NE release. Lysophosphatidylcholine (LPC), an endogenous amphiphiphilic lipid metabolite, also increases the concentration of interstitial adenosine in rat hearts, through the PKC-mediated activation of endogenous ecto-5'-nucleotidase. Nitric oxide (NO) facilitates the production of interstitial adenosine, via guanosine 3',5'-cyclic monophosphate (cGMP)-mediated activation of ecto-5'-nucleotidase as another pathway. These mechanisms play an important role in high sensitivity of the cardiac adenosine system. Adenosine plays an important role as a modulator of ischemic reperfusion injury, and that the production and mechanism of action of adenosine are linked with NE release.     

Evaluation of the tonic and phasic effects of endogenous adenosine on the transmitter secretion in the neuromuscular junction

Exogenous adenosine reduced the amplitude of multiquantal end-plate currents due to a depressant action on transmitter release. Theophylline did not change the amplitude of end-plate currents under low-rate motor nerve stimulation. The findings suggest a possibility of both tonic and phasic inhibitory actions of endogenous adenosine on transmitter release when utilization of this purine in synaptic cleft is inactivated.     

Adenosine metabolism in dog whole blood: Effects of dipyridamole

The effects of dipyridamole on the metabolism of adenosine added to dog whole blood were studied in vitro at 37°C. The half-lives for 8.8μM and 100μM adenosine were 3.42 and 6.89 min, respectively. Dipyridamole, in concentrations of 10^?7 to 10^?4M increased the half-life of 8.8μM adenosine 2 to 5-fold. The disappearance rate of adenosine in the presence of 10^?4 dipyridamole was similar to the disappearance rate of adenosine in plasma. Inosine formation was enhanced by dipyridamole. Control blood samples not receiving exogenous adenosine showed an increase in endogenous adenosine and inosine during 30 min of incubation. Endogenous production of nucleosides was unaffected by dipyridamole. Analysis showed that endogenous adenosine approached a steady-state concentration of 1μM. The results indicate that the half-life for adenosine in dog whole blood is significantly greater than previously reported and that dipyridamole is effective in inhibiting adenosine disappearance in concentrations as low as 10^?7M. The implications of endogenous production of adenosine are discussed.