Low Adenosine Levels Research Articles

Low Levels of Adenosine and GDNF Are Potential Risk Factors for Parkinson's Disease with Sleep Disorders.

Sleep disturbances are the most prevalent non-motor symptoms in the preclinical stage of Parkinson's disease (PD). Adenosine, glial-derived neurotrophic factor (GDNF), and associated neurotransmitters are crucial in the control of sleep arousal. This study aimed to detect the serum levels of adenosine, GDNF, and associated neurotransmitters and explored their correlations with PD with sleep disorders. Demographic characteristics and clinical information of PD patients and healthy participants were assessed. Serum concentrations of adenosine, GDNF, and related neurotransmitters were detected by ELISA and LC-MS. The correlation between serum levels of adenosine, GDNF, and associated neurotransmitters and sleep disorders was explored using logistic regression. PD patients with sleep disorders had higher scores of HAMA, HAMD, ESS, UPDRS-III, and H-Y stage. Lower levels of adenosine, GDNF, and γ-GABA were observed in PD patients who had sleep problems. Logistic regression analysis showed adenosine and GDNF were protective factors for preventing sleep disorders. Adenosine combined with GDNF had a higher diagnostic efficiency in predicting PD with sleep disorders by ROC analysis. This study revealed low adenosine and GDNF levels may be risk factors for sleep disorders in PD. The decrease of serum adenosine and GDNF levels may contribute to the diagnosis of PD with sleep disturbances.

Adenosine, adenosine- deaminase and implantable recorder outcome in syncopal patients.

Adenosine test was proposed as atool for identification of syncopal patients who benefit from pacemaker implantation. Aim of the study was to assess the relationship between adenosine levels, the outcome of adenosine test and results of implantable loop recorder (ILR) monitoring in patients with syncope. In 29 patients (mean age 59±11 years, 15 men, 14 women) with unexplained syncope ILR was implanted. In addition, adenosine test (intravenous injection of 20mg adenosine bolus) and assays of plasmatic adenosine and adenosine-deaminase were performed. Adenosine test was positive in 15 patients and negative in 14 patients. Patients with positive adenosine test had lower adenosine levels compared to patients with negative test (8.86±2.07ng/ml vs. 15.18±2.14ng/ml, p=.04). No difference was observed in adenosine deaminase levels (16.35±2.20 IU/l vs. 13.20±2.48 IU/l, p=.40). There was a negative correlation between adenosine level and AVB duration during adenosine test (p=.04; R2 =0.22). Patients with positive adenosine test had more frequent asystole during ILR monitoring than patients with negative test (9pts vs. 1pt, p=.005). Adenosine levels were lower in patients with asystolic syncope on ILR compared to vasodepressor syncope 8.20±2.86ng/ml versus 13.27±7.26ng/ml, p=.05). Patients with positive adenosine test have decreased production of endogenous adenosine compared to patients with negative adenosine test. Positivity of adenosine test and low adenosine level in the peripheral blood were associated with more frequent asystolic episodes during ILR monitoring.

Evaluation of Plasma Adenosine as a Marker of Cardiovascular Risk: Analytical and Biological Considerations.

BackgroundAdenosine is a ubiquitous regulatory molecule known to modulate signaling in many cells and processes vital to vascular homeostasis. While studies of adenosine receptors have dominated research in the field, quantification of adenosine systemically and locally remains limited owing largely to technical restrictions. Given the potential clinical implications of adenosine biology, there is a need for adequately powered studies examining the role of plasma adenosine in vascular health. We sought to describe the analytical and biological factors that affect quantification of adenosine in humans in a large, real‐world cohort of patients undergoing evaluation for coronary artery disease.Methods and ResultsBetween November 2016 and April 2018, we assessed 1141 patients undergoing angiography for evaluation of coronary artery disease. High‐performance liquid chromatography was used for quantification of plasma adenosine concentration, yielding an analytical coefficient of variance (CVa) of 3.2%, intra‐subject variance (CVi) 35.8% and inter‐subject variance (CVg) 56.7%. Traditional cardiovascular risk factors, medications, and clinical presentation had no significant impact on adenosine levels. Conversely, increasing age (P=0.027) and the presence of obstructive coronary artery disease (P=0.026) were associated with lower adenosine levels. Adjusted multivariable analysis supported only age being inversely associated with adenosine levels (P=0.039).ConclusionsPlasma adenosine is not significantly impacted by traditional cardiovascular risk factors; however, advancing age and presence of obstructive coronary artery disease may be associated with lower adenosine levels. The degree of intra‐ and inter‐subject variance of adenosine has important implications for biomarker use as a prognosticator of cardiovascular outcomes and as an end point in clinical studies.

Purinergic Signaling and Hippocampal Long-Term Potentiation

The purines ATP and adenosine are widely recognized for their neuromodulatory effects. They have been shown to have effects on neurons via various receptors and interactions with glial cells. In particular, long-term potentiation (LTP) in hippocampal slice preparations has been found to be modulated by ATP and adenosine. This review gives an overview of purinergic signaling in relation to hippocampal LTP and memory formation. The data supports the hypothesis that adenosine mediates a tonic suppression of synaptic transmission. Thus, low adenosine levels appear to increase basal synaptic activity via a decreased activation of the inhibitor A1 receptor, consequently making it more difficult to induce LTP because of lower contrast. During high stimulation, the inhibition of neighboring pathways by adenosine, in combination with an A2a receptor activation, appears to increase contrast of excited pathways against a nonexcited background. This would enable amplification of specific signaling while suppressing non-specific events. Although a clear role for purinergic signaling in LTP is evident, more studies are needed to scrutinize the modulatory role of ATP and adenosine and their receptors in synaptic plasticity and memory.

A₃ adenosine receptor allosteric modulator induces an anti-inflammatory effect: in vivo studies and molecular mechanism of action.

The A3 adenosine receptor (A3AR) is overexpressed in inflammatory cells and in the peripheral blood mononuclear cells of individuals with inflammatory conditions. Agonists to the A3AR are known to induce specific anti-inflammatory effects upon chronic treatment. LUF6000 is an allosteric compound known to modulate the A3AR and render the endogenous ligand adenosine to bind to the receptor with higher affinity. The advantage of allosteric modulators is their capability to target specifically areas where adenosine levels are increased such as inflammatory and tumor sites, whereas normal body cells and tissues are refractory to the allosteric modulators due to low adenosine levels. LUF6000 administration induced anti-inflammatory effect in 3 experimental animal models of rat adjuvant induced arthritis, monoiodoacetate induced osteoarthritis, and concanavalin A induced liver inflammation in mice. The molecular mechanism of action points to deregulation of signaling proteins including PI3K, IKK, IκB, Jak-2, and STAT-1, resulting in decreased levels of NF-κB, known to mediate inflammatory effects. Moreover, LUF6000 induced a slight stimulatory effect on the number of normal white blood cells and neutrophils. The anti-inflammatory effect of LUF6000, mechanism of action, and the differential effects on inflammatory and normal cells position this allosteric modulator as an attractive and unique drug candidate.

Detrimental effects of adenosine signaling in sickle cell disease

Hypoxia can act as an initial trigger to induce erythrocyte sickling and eventual end organ damage in sickle cell disease (SCD). Many factors and metabolites are altered in response to hypoxia and may contribute to the pathogenesis of the disease. Using metabolomic profiling, we found that the steady-state concentration of adenosine in the blood was elevated in a transgenic mouse model of SCD. Adenosine concentrations were similarly elevated in the blood of humans with SCD. Increased adenosine levels promoted sickling, hemolysis and damage to multiple tissues in SCD transgenic mice and promoted sickling of human erythrocytes. Using biochemical, genetic and pharmacological approaches, we showed that adenosine A(2B) receptor (A(2B)R)-mediated induction of 2,3-diphosphoglycerate, an erythrocyte-specific metabolite that decreases the oxygen binding affinity of hemoglobin, underlies the induction of erythrocyte sickling by excess adenosine both in cultured human red blood cells and in SCD transgenic mice. Thus, excessive adenosine signaling through the A(2B)R has a pathological role in SCD. These findings may provide new therapeutic possibilities for this disease.

Role of extracellular nucleotide phosphohydrolysis in intestinal ischemia‐reperfusion injury

Extracellular adenosine has been implicated as an innate antiinflammatory metabolite, particularly during conditions of limited oxygen availability such as ischemia. Because extracellular adenosine generation is primarily produced via phosphohydrolysis from its precursor molecule adenosine-monophosphate (AMP) through the enzyme ecto-5'-nucleotidase (CD73), we examined the contribution of CD73-dependent adenosine production in modulation of intestinal ischemia-reperfusion (IR) injury. Following transcriptional and translational profiling of intestinal tissue that revealed a prominent induction of murine CD73, we next determined the role of CD73 in protection against intestinal IR injury. Interestingly, pharmacological inhibition or targeted gene deletion of CD73 significantly enhanced not only local intestinal injury, but also secondary organ injury, following IR as measured by intestinal and lung myeloperoxidase, aspartate and alanine aminotransferase, interleukin (IL) -1, IL-6, and histological injury. To confirm the role of CD73 in intestinal adenosine production, we measured adenosine tissue levels and found that they were increased with IR injury. In contrast, CD73-deficient (cd73(-/-)) mice had lower adenosine levels at baseline and no increase with IR injury. Finally, reconstitution of cd73(-/-) mice or treatment of wild-type mice with soluble 5'-nucleotidase was associated with significantly lower levels of injury. These data reveal a previously unrecognized role of CD73 in attenuating intestinal IR-mediated injury.

Integration of vascular, contractile and metabolic responses to hypoxia: effects of maturation and adenosine.

We examined myocardial responses to reduced arterial PO2 and the role of endogenous adenosine in constant-pressure perfused hearts from immature (Imm) and mature (Mat) rabbits. During normoxia, coronary flow and myocardial O2 consumption were similar in both groups. With moderate hypoxia, coronary perfusate flow increased by 125 +/- 16% in Imm but by only 68 +/- 12% in Mat hearts. Imm hearts displayed better maintenance of contractile function (87 vs. 67% in Mat hearts) and metabolic state. Blockade of cardiac adenosine receptors with 25 microM 8-p-sulfophenyltheophylline attenuated vasodilation during hypoxia, reduced contractile function, and abolished age-related differences in one response to hypoxia. Myocardial purine release and extracellular purine levels were threefold higher in Mat compared with Imm hearts and was associated with higher cytosolic 5'-AMP concentration (and lower [ATP]/[ADP].[P(i)], where [ATP], [ADP], and [P(i)] are concentrations of ATP, ADP and P(i), respectively) in Mat hearts. In summary, 1) Imm hearts are functionally and metabolically more tolerant of hypoxic perfusion, largely because of improved hypoxia-induced coronary vasodilation; 2) endogenous adenosine mediates this beneficial vasodilation, enhancing hypoxic tolerance; and 3) improved vascular sensitivity to adenosine allows for enhanced vasodilatory responses in the face of lower adenosine levels and higher energy state in immature hearts.

1015-71 Lower Adenosine Levels During Early Ischemia: Cause of Increased Injury in the Aging Heart?

Myocardial injury following ischemia and reperfusion is increased in the aging heart, including in the aging 24 mo Fischer 344 rat (mean survival 29 mo) compared to the Fischer 344 adult (6 mo). Adenosine (ADO) has been increasingly appreciated as a cardioprotective agent in myocardial ischemia. We hypothesized that a potential mechanism of the increased injury in the aging heart is decreased production of ADO in response to ischemia. Isolated buffer perfused (glucose 5 mM — insulin 5 μ /1) hearts from aging and adult Fischer 344 rats were subjected to stop-flow ischemia for either 2, 5, 10, 15 or 25 min after a 15 min equilibration phase. ADO and hypoxanthine (HX) were measured by HPLC. Tissue total adenylates, ATP, glycogen and lactate were measured. Ischemia Pre-ischemia 2 min 5 min 10 min 15 min 25 min (n = 5) (n = 5) (n = 5) (n = 5) (n = 5) (n = 5) ADO 6 mo 0.2 ± 0.1 0.7 ± 0.1 0.7 ± 0.1 2.2 ± 0.2 37 ± 0.6 4.5 ± 0.3 24 mo 0.2 ± 0.1 05 ± 0.1 0.3 ± 0.1 * 1.1 ± 0.3 * 2.1 ± 0.3 * 3.5 ± 0.4 HX 6 mo 2.0 ± 0.2 1.6 ± 0.2 2.3 ± 0.3 2.2 ± 0.1 25 ± 0.2 2.8 ± 0.2 24 mo 1.9 ± 0.2 1.4 ± 0.1 1.4 ± 0.2 * 2.1 ± 0.2 2.2 ± 0.8 3.5 ± 0.3 (Mean ± SE, nmol/mg protein) * p < 0.05 vs 6 mo ADO levels were 50% lower in the aging heart at 5 and 10 min, remained depressed at 15 min and had not fully equalized to adult levels by 25 min of ischemia. HX was decreased at 5 min, consistent with decreased ADO production at that time. The change in total adenylate pool was similar in both groups. The decrease in tissue glycogen and increase in lactate were similar during ischemia in both groups, suggesting comparable glycolytic activity. ATP levels were decreased in aging hearts at 5 min (11.1 ± 2.3 vs 18.7 ± 1.9 nmol/mg protein), but reached levels similar to adult hearts as ischemia progressed. Thus, decreased ADO levels during early ischemia may reflect reduced production of ADO in the aging heart, and increase the susceptibility of the aging heart to damage during ischemia and reperfusion.

Subtraction method for the high-performance liquid chromatographic measurement of plasma adenosine

The measurement of plasma adenosine with traditional high-performance liquid chromatographic techniques is difficult because of its nanomolar concentration, its short half-life in blood, and because of the difficulty in isolating adenosine from interfering peaks in the chromatogram. To prevent loss of adenosine in the blood sample, a “stop solution” is used to prevent enzymatic degradation and cellular uptake. Peak-shifting techniques on fractionated samples to measure adenosine derivatives have been used in the past to avoid interfering peaks in the chromatogram. A new method has been developed by which nanomolar levels of plasma adenosine can be accurately measured despite co-eluting peaks in the chromatogram. In this method, plasma samples are collected with a stop solution, processed, and divided. Adenosine deaminase is added to part of the sample to form a blank. A computer program subtracts the blank chromatogram from the paired unknown, and the result is compared to adenosine standards prepared from the blank and subtracted in a similar fashion. With this subtraction method, the overall recovery of physiological concentrations of adenosine was 89% from dog blood, and the average coefficient of variation was 12%. In summary, the subtraction method of plasma adenosine measurement offers good recovery, reproducibility, and the ability to quantify low levels of adenosine despite interfering peaks in the chromatogram.

Improved high-pressure liquid chromatographic-fluorometric assay for measurement of adenosine in plasma

A high-pressure liquid chromatographic system using fluorescence detection was characterized for the determination of adenosine levels in plasma from anesthetized cat, rat, dog, mouse, rabbit, and guinea pig. The optimal concentration of chloroacetaldehyde necessary to convert physiological levels of adenosine to its fluorescent derivative 1,N6-ethenoadenosine (ethenoadenosine) was in excess of 220 mM. As little as 0.2 pmol of ethenoadenosine could be measured, and detection was linear up to 200 pmol. Derivatization of adenosine into ethenoadenosine was highly dependent on temperature and time. However, ethenoadenosine showed thermal instability in that levels dropped sharply after 30 min at 100 degrees C, 2 h at 80 degrees C, and 24 h at 55 degrees C. Adenosine nucleotides were extracted from plasma samples with an efficiency of greater than 91% to prevent adenosine formation from ATP and AMP that would have otherwise occurred during the derivatization procedure. Plasma levels (microM) of adenosine in venous blood were 0.31 in dog, 0.54 in cat, 0.71 in guinea pig, 1.03 in mouse, 1.04 in rat, and 1.68 in rabbit. Plasma levels of adenosine in arterial blood were not significantly different from levels in venous blood. This method can be used to measure even very low levels of adenosine without interference from nucleotide breakdown.

Circadian variations of adenosine level in blood and liver and its possible physiological significance

The role of adenosine as a possible physiological modulator was explored by measuring its concentration in different tissues during a 24-hour period. Initially the circadian variations of adenosine and other purine compounds such as inosine, hypoxanthine, uric acid and adenine nucleotides were studied in the rat blood. A daily cyclic response was observed, with low levels of adenosine from 08.00 – 20.00 h, followed by an increase from this time on. Inosine and hypoxanthine levels were elevated during the day and low at night. The uric acid changes observed indicate that the decrease in purine catabolism coincides with a decrease in inosine and hypoxanthine levels and an increase in adenosine. The blood adenine nucleotides, energy charge and phosphorylation potential remained constant during the day and showed oscillatory changes during the night. Similar studies were made in the liver, a primary source of circulating purines. Liver adenosine was high during the night while inosine and hypoxanthine remained low along the 24 hours. The results suggest that liver purine metabolism might participate in the maintenance and renewal of the blood purine pool and in the energy state of erythrocytes in vivo .

Cyclic AMP-generating systems in cell-free preparations from guinea pig cerebral cortex: loss of adenosine and amine responsiveness due to low levels of endogenous adenosine.

The accumulations of radioactive cyclic AMP elicited by adenosine, norepinephrine, and histamine in adenine-labeled vesicular entities of a particulate fraction from guinea pig cerebral cortex are greatly reduced as a result of prolonged preincubation. The presence of adenosine deaminase during preincubations largely prevents the loss of adenosine, norepinephrine and histamine responses. Adenosine deaminase was inactivated by deoxycoformycin prior to stimulation of cyclic AMP accumulation by adenosine or amines. If adenosine deaminase is not inactivated, responses to norepinephrine are not significant and histamine responses are reduced by 50%. Adenosine deaminase cannot restore responsiveness of the cyclic AMP-generating systems. It is proposed that, in particulate fractions of guinea pig cerebral cortex, low levels of adenosine cause a slow loss of receptors and/or coupling of receptors to cyclic AMP-generating systems.

Effect of nitrogen starvation on the level of adenosine 3′,5′-monophosphate in Anabaena variabilis

Low levels of adenosine 3′,5′-monophosphate (cyclic AMP) were detected in the cyanobacterium Anabaena variabilis using a protein binding assay and two radioisotopic labelling methods. The basal concentration of intracellular cyclic AMP ranged from 0.27 pmol/mg protein in A. variabilis Kutz grown under heterotrophic conditions to 1.0–2.7 pmol/mg protein in A. variabilis strain 377 grown autotrophically. Extracellular cyclic AMP was found to comprise as much as 90% of the total cyclic AMP in rapidly growing cultures. When A. variabilis strain 377 was starved of nitrogen, a 3–4-fold increase in intracellular cyclic AMP was observed during the 24 h period coincident with early heterocyst development.

Glucose transport in isolated prosthecae of Asticcacaulis biprosthecum.

Active transport of glucose in prosthecae isolated from cells of Asticcacaulis biprosthecum was stimulated by the non-physiological electron donor N, N, N', N'-tetramethyl-p-phenylenediamine dihydrochloride. Glucose uptake was mediated by two transport systems; the apparent Km of the high-affinity system was 1.8 muM and that of the low-affinity system was 34 muM. Free glucose accumulated within prosthecae at a concentration 60 to 200 times above that present externally, depending on the Km of the system being observed. The glucose transport system in prosthecae was stereospecific for D-glucose, and neither methyl alpha-D-glucopyranoside nor 2-deoxyglucose was transported. Uptake of glucose was inhibited by N-ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB), and the inhibition by PCMB but not by NEM was reversed by dithiothreitol. Glucose uptake was also inhibited by the uncoupling agents 5-chloro-3-t-butyl-2'-nitrosalicylanilide (S-13), 5-chloro-3-(p-chlorophenyl)-4'-chlorosalicylanilide (S-6), and carbonyl-cyanide m-chlorophenylhydrazone (CCCP) and by the respiratory inhibitor KCN. Efflux of glucose from preloaded prosthecae was induced by PCMB and KCN, but not by S-13 or CCCP. Glucose uptake was not affected by arsenate or an inhibitor of membrane-bound adenosine triphosphatases, N, N'-dicyclohexylcarbodiimide. The lack of inhibition by these two compounds, combined with the extremely low levels of adenosine 5'-triphosphate present in prosthecae, indicates that adenosine 5'-triphosphate is not involved in the transport of glucose by prosthecae.

Comparison of the adenine nucleotide metabolism of dog atrial and ventricular myocardium

Measurements of atrial and ventricular adenine nucleotides, nucleosides and bases in dog heart indicated that normal atrial ATP, ADP and AMP levels are about one-half those of normal ventricular tissue, but atrial and ventricular tissue levels of adenosine, inosine, and hypoxanthine were about equal. With ischemia, production of adenosine and inosine was less in atrial than in ventricular muscle. In the ventricle, 5′-nucleotidase is associated with external membranes, including those of the T-tubular system and the lack of a T-tubular network in atrium suggested that there might be fewer AMP hydrolyzing sites and hence a slower rate of adenosine production in response to ischemia. This possibility was ruled out since measurements of 5′-nucleotidase on crude tissue extracts indicated that atrial tissue has 2 to 3 times the activity of ventricular tissue. Furthermore, measurements of adenosine kinase activity indicate that atrial and ventricular activities of this enzyme are about equal; hence the lower adenosine levels in atria are also not due to a higher rate of adenosine rephosphorylation. Inhibition of 5′-nucleotidase by ATP and ADP can also be eliminated as a cause of the differences in atrial and ventricular adenosine formation since the concentrations of these compounds are lower in atrial than in ventricular muscle. Finally, the lower levels of IMP and inosine in atrial tissue indicate that deamination by adenylate deaminase is not the major pathway for AMP degradation in atrial tissue. It is possible that the lower substrate (AMP) concentration for 5′-nucleotidase is responsible for the lower adenosine production by atrial tissue in response to ischemia. However, to what extent this contributed to the lower atrial adenosine levels cannot be determined from these data.