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Sample records for enhancing adenosine monophosphate-activated

  1. Atorvastatin and pitavastatin enhance lipoprotein lipase production in L6 skeletal muscle cells through activation of adenosine monophosphate-activated protein kinase.

    PubMed

    Ohira, Masahiro; Endo, Kei; Saiki, Atsuhito; Miyashita, Yoh; Terai, Kensuke; Murano, Takeyoshi; Watanabe, Fusako; Tatsuno, Ichiro; Shirai, Kohji

    2012-10-01

    Pravastatin and atorvastatin increase the serum level of lipoprotein lipase (LPL) mass in vivo but do not increase LPL activity in 3T3-L1 preadipocytes in vitro. LPL is mainly produced by adipose tissue and skeletal muscle cells. Metformin enhances LPL in skeletal muscle through adenosine monophosphate-activated protein kinase (AMPK) activation but not in adipocytes. This study aimed to examine the effect of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) on LPL production and to investigate the mechanism by which statins enhance skeletal muscle cell LPL production. L6 skeletal muscle cells were incubated with pravastatin, simvastatin, atorvastatin or pitavastatin. LPL activity, protein levels and mRNA expression were measured. Atorvastatin and pitavastatin significantly increased LPL activity, protein levels and mRNA expression in L6 skeletal muscle cells at 1 μmol/L, but neither statin had an effect at 10 μmol/L. We measured AMPK to clarify the mechanism by which statins increase LPL production in skeletal muscle cells. At 1 μmol/L, both atorvastatin and pitavastatin enhanced AMPK activity, but this enhancement was abolished when AMPK signaling was blocked by compound C. The increased expressions of LPL protein and mRNA by atorvastatin and pitavastatin were reduced by compound C. In addition, mevalonic acid abolished atorvastatin- and pitavastatin-induced AMPK activation and LPL expression. These results suggest that atorvastatin and pitavastatin increase LPL activity, protein levels and LPL mRNA expression by activating AMPK in skeletal muscle cells.

  2. SIRT1/Adenosine Monophosphate-Activated Protein Kinase α Signaling Enhances Macrophage Polarization to an Anti-inflammatory Phenotype in Rheumatoid Arthritis.

    PubMed

    Park, So Youn; Lee, Sung Won; Lee, Sang Yeob; Hong, Ki Whan; Bae, Sun Sik; Kim, Koanhoi; Kim, Chi Dae

    2017-01-01

    Macrophages are crucially involved in the pathogenesis of rheumatoid arthritis (RA). Macrophages of the M1 phenotype act as pro-inflammatory mediators in synovium, whereas those of the M2 phenotype suppress inflammation and promote tissue repair. SIRT1 is a class 3 histone deacetylase with anti-inflammatory characteristics. However, the role played by SIRT1 in macrophage polarization has not been defined in RA. We investigated whether SIRT1 exerts anti-inflammatory effects by modulating M1/M2 polarization in macrophages from RA patients. In this study, SIRT1 activation promoted the phosphorylation of an adenosine monophosphate-activated protein kinase (AMPK) α/acetyl-CoA carboxylase in macrophages exposed to interleukin (IL)-4, and that this resulted in the expressions of M2 genes, including MDC, FcεRII, MrC1, and IL-10, at high levels. Furthermore, these expressions were inhibited by sirtinol (an inhibitor of SIRT1) and compound C (an inhibitor of AMPK). Moreover, SIRT1 activation downregulated LPS/interferon γ-mediated NF-κB activity by inhibiting p65 acetylation and the expression of M1 genes, such as CCL2, iNOS, IL-12 p35, and IL-12 p40. Macrophages from SIRT1 transgenic (Tg)-mice exhibited enhanced polarization of M2 phenotype macrophages and reduced polarization of M1 phenotype macrophages. In line with these observations, SIRT1-Tg mice showed less histological signs of arthritis, that is, lower TNFα and IL-1β expressions and less severe arthritis in the knee joints, compared to wild-type mice. Taken together, the study shows activation of SIRT1/AMPKα signaling exerts anti-inflammatory activities by regulating M1/M2 polarization, and thereby reduces inflammatory responses in RA. Furthermore, it suggests that SIRT1 signaling be viewed as a therapeutic target in RA.

  3. Enhanced Production of Adenosine Triphosphate by Pharmacological Activation of Adenosine Monophosphate-Activated Protein Kinase Ameliorates Acetaminophen-Induced Liver Injury.

    PubMed

    Hwang, Jung Hwan; Kim, Yong-Hoon; Noh, Jung-Ran; Choi, Dong-Hee; Kim, Kyoung-Shim; Lee, Chul-Ho

    2015-10-01

    The hepatic cell death induced by acetaminophen (APAP) is closely related to cellular adenosine triphosphate (ATP) depletion, which is mainly caused by mitochondrial dysfunction. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a key sensor of low energy status. AMPK regulates metabolic homeostasis by stimulating catabolic metabolism and suppressing anabolic pathways to increase cellular energy levels. We found that the decrease in active phosphorylation of AMPK in response to APAP correlates with decreased ATP levels, in vivo. Therefore, we hypothesized that the enhanced production of ATP via AMPK stimulation can lead to amelioration of APAP-induced liver failure. A769662, an allosteric activator of AMPK, produced a strong synergistic effect on AMPK Thr172 phosphorylation with APAP in primary hepatocytes and liver tissue. Interestingly, activation of AMPK by A769662 ameliorated the APAP-induced hepatotoxicity in C57BL/6N mice treated with APAP at a dose of 400 mg/kg intraperitoneally. However, mice treated with APAP alone developed massive centrilobular necrosis, and APAP increased their serum alanine aminotransferase and aspartate aminotransferase levels. Furthermore, A769662 administration prevented the loss of intracellular ATP without interfering with the APAP-mediated reduction of mitochondrial dysfunction. In contrast, inhibition of glycolysis by 2-deoxy-glucose eliminated the beneficial effects of A769662 on APAP-mediated liver injury. In conclusion, A769662 can effectively protect mice against APAP-induced liver injury through ATP synthesis by anaerobic glycolysis. Furthermore, stimulation of AMPK may have potential therapeutic application for APAP overdose.

  4. Irisin ameliorates hepatic glucose/lipid metabolism and enhances cell survival in insulin-resistant human HepG2 cells through adenosine monophosphate-activated protein kinase signaling.

    PubMed

    So, Wing Yan; Leung, Po Sing

    2016-09-01

    Irisin is a newly identified myokine that promotes the browning of white adipose tissue, enhances glucose uptake in skeletal muscle and modulates hepatic metabolism. However, the signaling pathways involved in the effects on hepatic glucose and lipid metabolism have not been resolved. This study aimed to examine the role of irisin in the regulation of hepatic glucose/lipid metabolism and cell survival, and whether adenosine monophosphate-activated protein kinase (AMPK), a master metabolic regulator in the liver, is involved in irisin's actions. Human liver-derived HepG2 cells were cultured in normal glucose-normal insulin (NGNI) or high glucose-high insulin (HGHI/insulin-resistant) condition. Hepatic glucose and lipid metabolism was evaluated by glucose output and glycogen content or triglyceride accumulation assays, respectively. Our results showed that irisin stimulated phosphorylation of AMPK and acetyl-CoA-carboxylase (ACC) via liver kinase B1 (LKB1) rather than Ca(2+)/calmodulin-dependent protein kinase kinase β (CaMKKβ) in HepG2 cells. Irisin ameliorated hepatic insulin resistance induced by HGHI condition. Irisin reduced hepatic triglyceride content and glucose output, but increased glycogen content, with those effects reversed by dorsomorphin, an AMPK inhibitor. Furthermore, irisin also stimulated extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and promoted cell survival in an AMPK-dependent manner. In conclusion, our data indicate that irisin ameliorates dysregulation of hepatic glucose/lipid metabolism and cell death in insulin-resistant states via AMPK activation. These findings reveal a novel irisin-mediated protective mechanism in hepatic metabolism which provides a scientific basis for irisin as a potential therapeutic target for the treatment of insulin resistance and type 2 diabetes mellitus.

  5. Impaired adenosine monophosphate-activated protein kinase signalling in dorsal root ganglia neurons is linked to mitochondrial dysfunction and peripheral neuropathy in diabetes

    PubMed Central

    Smith, Darrell R.; Saleh, Ali; Schapansky, Jason; Marquez, Alexandra; Gomes, Suzanne; Akude, Eli; Morrow, Dwane; Calcutt, Nigel A.; Fernyhough, Paul

    2012-01-01

    Mitochondrial dysfunction occurs in sensory neurons and may contribute to distal axonopathy in animal models of diabetic neuropathy. The adenosine monophosphate-activated protein kinase and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) signalling axis senses the metabolic demands of cells and regulates mitochondrial function. Studies in muscle, liver and cardiac tissues have shown that the activity of adenosine monophosphate-activated protein kinase and PGC-1α is decreased under hyperglycaemia. In this study, we tested the hypothesis that deficits in adenosine monophosphate-activated protein kinase/PGC-1α signalling in sensory neurons underlie impaired axonal plasticity, suboptimal mitochondrial function and development of neuropathy in rodent models of type 1 and type 2 diabetes. Phosphorylation and expression of adenosine monophosphate-activated protein kinase/PGC-1α and mitochondrial respiratory chain complex proteins were downregulated in dorsal root ganglia of both streptozotocin-diabetic rats and db/db mice. Adenoviral-mediated manipulation of endogenous adenosine monophosphate-activated protein kinase activity using mutant proteins modulated neurotrophin-directed neurite outgrowth in cultures of sensory neurons derived from adult rats. Addition of resveratrol to cultures of sensory neurons derived from rats after 3–5 months of streptozotocin-induced diabetes, significantly elevated adenosine monophosphate-activated protein kinase levels, enhanced neurite outgrowth and normalized mitochondrial inner membrane polarization in axons. The bioenergetics profile (maximal oxygen consumption rate, coupling efficiency, respiratory control ratio and spare respiratory capacity) was aberrant in cultured sensory neurons from streptozotocin-diabetic rats and was corrected by resveratrol treatment. Finally, resveratrol treatment for the last 2 months of a 5-month period of diabetes reversed thermal hypoalgesia and attenuated foot skin

  6. Small molecule adenosine 5'-monophosphate activated protein kinase (AMPK) modulators and human diseases.

    PubMed

    Rana, Sandeep; Blowers, Elizabeth C; Natarajan, Amarnath

    2015-01-08

    Adenosine 5'-monophosphate activated protein kinase (AMPK) is a master sensor of cellular energy status that plays a key role in the regulation of whole-body energy homeostasis. AMPK is a serine/threonine kinase that is activated by upstream kinases LKB1, CaMKKβ, and Tak1, among others. AMPK exists as αβγ trimeric complexes that are allosterically regulated by AMP, ADP, and ATP. Dysregulation of AMPK has been implicated in a number of metabolic diseases including type 2 diabetes mellitus and obesity. Recent studies have associated roles of AMPK with the development of cancer and neurological disorders, making it a potential therapeutic target to treat human diseases. This review focuses on the structure and function of AMPK, its role in human diseases, and its direct substrates and provides a brief synopsis of key AMPK modulators and their relevance in human diseases.

  7. Metabolic syndrome: adenosine monophosphate-activated protein kinase and malonyl coenzyme A.

    PubMed

    Ruderman, Neil B; Saha, Asish K

    2006-02-01

    The metabolic syndrome can be defined as a state of metabolic dysregulation characterized by insulin resistance, central obesity, and a predisposition to type 2 diabetes, dyslipidemia, premature atherosclerosis, and other diseases. An increasing body of evidence has linked the metabolic syndrome to abnormalities in lipid metabolism that ultimately lead to cellular dysfunction. We review here the hypothesis that, in many instances, the cause of these lipid abnormalities could be a dysregulation of the adenosine monophosphate-activated protein kinase (AMPK)/malonyl coenzyme A (CoA) fuel-sensing and signaling mechanism. Such dysregulation could be reflected by isolated increases in malonyl CoA or by concurrent changes in malonyl CoA and AMPK, both of which would alter intracellular fatty acid partitioning. The possibility is also raised that pharmacological agents and other factors that activate AMPK and/or decrease malonyl CoA could be therapeutic targets.

  8. 5'-adenosine monophosphate-activated protein kinase and the metabolic syndrome.

    PubMed

    Mor, Vijay; Unnikrishnan, M K

    2011-09-01

    Lifestyle changes such as physical inactivity combined with calorie-rich, low-fibre diets have triggered an explosive surge in metabolic syndrome, outlined as a cluster of heart attack risk factors such as insulin resistance, raised fasting plasma glucose, abdominal obesity, high cholesterol and high blood pressure. By acting as a master-switch of energy homeostasis and associated pathophysiological phenomena, 5'-adenosine monophosphate-activated protein kinase (AMPK) appears to orchestrate the adaptive physiology of energy deficit, suggesting that the sedentary modern human could be suffering from chronic suboptimal AMPK activation. Addressing individual targets with potent ligands with high specificity may be inappropriate (it has not yielded any molecule superior to the sixty year old metformin) because this strategy cannot address a cluster of interrelated pathologies. However, spices, dietary supplements and nutraceuticals attenuate the multiple symptoms of metabolic syndrome in a collective and perhaps more holistic fashion with fewer adverse events. Natural selection could have favoured races that developed a taste for spices and dietary supplements, most of which are not only antioxidants but also activators of AMPK. The review will outline the various biochemical mechanisms and pathophysiological consequences of AMPK activation involving the cluster of symptoms that embrace metabolic syndrome and beyond. Recent advances that integrate energy homeostasis with a number of overarching metabolic pathways and physiological phenomena, including inflammatory conditions, cell growth and development, malignancy, life span, and even extending into environmental millieu, as in obesity mediated by gut microflora and others will also be outlined.

  9. Adenosine monophosphate-activated protein kinase-based classification of diabetes pharmacotherapy.

    PubMed

    Dutta, D; Kalra, S; Sharma, M

    2016-09-21

    The current classification of both diabetes and antidiabetes medication is complex, preventing a treating physician from choosing the most appropriate treatment for an individual patient, sometimes resulting in patient-drug mismatch. We propose a novel, simple systematic classification of drugs, based on their effect on adenosine monophosphate-activated protein kinase (AMPK). AMPK is the master regular of energy metabolism, an energy sensor, activated when cellular energy levels are low, resulting in activation of catabolic process, and inactivation of anabolic process, having a beneficial effect on glycemia in diabetes. This listing of drugs makes it easier for students and practitioners to analyze drug profiles and match them with patient requirements. It also facilitates choice of rational combinations, with complementary modes of action. Drugs are classified as stimulators, inhibitors, mixed action, possible action, and no action on AMPK activity. Metformin and glitazones are pure stimulators of AMPK. Incretin-based therapies have a mixed action on AMPK. Sulfonylureas either inhibit AMPK or have no effect on AMPK. Glycemic efficacy of alpha-glucosidase inhibitors, sodium glucose co-transporter-2 inhibitor, colesevelam, and bromocriptine may also involve AMPK activation, which warrants further evaluation. Berberine, salicylates, and resveratrol are newer promising agents in the management of diabetes, having well-documented evidence of AMPK stimulation medicated glycemic efficacy. Hence, AMPK-based classification of antidiabetes medications provides a holistic unifying understanding of pharmacotherapy in diabetes. This classification is flexible with a scope for inclusion of promising agents of future.

  10. Xylazine Activates Adenosine Monophosphate-Activated Protein Kinase Pathway in the Central Nervous System of Rats

    PubMed Central

    Shi, Xing-Xing; Yin, Bai-Shuang; Yang, Peng; Chen, Hao; Li, Xin; Su, Li-Xue; Fan, Hong-Gang; Wang, Hong-Bin

    2016-01-01

    Xylazine is a potent analgesic extensively used in veterinary and animal experimentation. Evidence exists that the analgesic effect can be inhibited using adenosine 5’-monophosphate activated protein kinase (AMPK) inhibitors. Considering this idea, the aim of this study was to investigate whether the AMPK signaling pathway is involved in the central analgesic mechanism of xylazine in the rat. Xylazine was administrated via the intraperitoneal route. Sprague-Dawley rats were sacrificed and the cerebral cortex, cerebellum, hippocampus, thalamus and brainstem were collected for determination of liver kinase B1 (LKB1) and AMPKα mRNA expression using quantitative real-time polymerase chain reaction (qPCR), and phosphorylated LKB1 and AMPKα levels using western blot. The results of our study showed that compared with the control group, xylazine induced significant increases in AMPK activity in the cerebral cortex, hippocampus, thalamus and cerebellum after rats received xylazine (P < 0.01). Increased AMPK activities were accompanied with increased phosphorylation levels of LKB1 in corresponding regions of rats. The protein levels of phosphorylated LKB1 and AMPKα in these regions returned or tended to return to control group levels. However, in the brainstem, phosphorylated LKB1 and AMPKα protein levels were decreased by xylazine compared with the control (P < 0.05). In conclusion, our data indicates that xylazine alters the activities of LKB1 and AMPK in the central nervous system of rats, which suggests that xylazine affects the regulatory signaling pathway of the analgesic mechanism in the rat brain. PMID:27049320

  11. Adenosine monophosphate-activated kinase and its key role in catabolism: structure, regulation, biological activity, and pharmacological activation.

    PubMed

    Krishan, Sukriti; Richardson, Des R; Sahni, Sumit

    2015-01-01

    Adenosine monophosphate-activated protein kinase (AMPK) is a cellular energy sensor, which once activated, plays a role in several processes within the cell to restore energy homeostasis. The protein enhances catabolic pathways, such as β-oxidation and autophagy, to generate ATP, and inhibits anabolic processes that require energy, including fatty acid, cholesterol, and protein synthesis. Due to its key role in the regulation of critical cellular pathways, deregulation of AMPK is associated with the pathology of many diseases, including cancer, Wolff-Parkinson-White syndrome, neurodegenerative disorders, diabetes, and metabolic syndrome. In fact, AMPK is a target of some pharmacological agents implemented in the treatment of diabetes (metformin and thiazolidinediones) as well as other naturally derived products, such as berberine, which is used in traditional medicine. Due to its critical role in the cell and the pathology of several disorders, research into developing AMPK as a therapeutic target is becoming a burgeoning and exciting field of pharmacological research. A profound understanding of the regulation and activity of AMPK would enhance its development as a promising therapeutic target.

  12. Adenosine monophosphate-activated protein kinase attenuates cardiomyocyte hypertrophy through regulation of FOXO3a/MAFbx signaling pathway.

    PubMed

    Chen, Baolin; Wu, Qiang; Xiong, Zhaojun; Ma, Yuedong; Yu, Sha; Chen, Dandan; Huang, Shengwen; Dong, Yugang

    2016-09-01

    Control of cardiac muscle mass is thought to be determined by a dynamic balance of protein synthesis and degradation. Recent studies have demonstrated that atrophy-related forkhead box O 3a (FOXO3a)/muscle atrophy F-box (MAFbx) signaling pathway plays a central role in the modulation of proteolysis and exert inhibitory effect on cardiomyocyte hypertrophy. In this study, we tested the hypothesis that adenosine monophosphate-activated protein kinase (AMPK) activation attenuates cardiomyocyte hypertrophy by regulating FOXO3a/MAFbx signaling pathway and its downstream protein degradation. The results showed that activation of AMPK with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) attenuated cardiomyocyte hypertrophy induced by angiotensin II (Ang II). The antihypertrophic effects of AICAR were blunted by AMPK inhibitor Compound C. In addition, AMPK dramatically increased the activity of transcription factor FOXO3a, up-regulated the expression of its downstream ubiquitin ligase MAFbx, and enhanced cardiomyocyte proteolysis. Meanwhile, the effects of AMPK on protein degradation and cardiomyocyte hypertrophy were blocked after MAFbx was silenced by transfection of cardiomyocytes with MAFbx-siRNA. These results indicate that AMPK plays an important role in the inhibition of cardiomyocyte hypertrophy by activating protein degradation via FOXO3a/MAFbx signaling pathway.

  13. Alpha-Lipoic acid increases energy expenditure by enhancing adenosine monophosphate-activated protein kinase-peroxisome proliferator-activated receptor-gamma coactivator-1alpha signaling in the skeletal muscle of aged mice

    USDA-ARS?s Scientific Manuscript database

    Skeletal muscle mitochondrial dysfunction is associated with aging and diabetes, which decreases respiratory capacity and increases reactive oxygen species. Lipoic acid (LA) possesses antioxidative and antidiabetic properties. Metabolic action of LA is mediated by activation of adenosine monophospha...

  14. Electroacupuncture preconditioning attenuates ischemic brain injury by activation of the adenosine monophosphate-activated protein kinase signaling pathway

    PubMed Central

    Ran, Qiang-qiang; Chen, Huai-long; Liu, Yan-li; Yu, Hai-xia; Shi, Fei; Wang, Ming-shan

    2015-01-01

    Electroacupuncture has therapeutic effects on ischemic brain injury, but its mechanism is still poorly understood. In this study, mice were stimulated by electroacupuncture at the Baihui (GV20) acupoint for 30 minutes at 1 mA and 2/15 Hz for 5 consecutive days. A cerebral ischemia model was established by ligating the bilateral common carotid artery for 15 minutes. At 72 hours after injury, neuronal injury in the mouse hippocampus had lessened, and the number of terminal deoxynucleotide transferase-mediated dUTP nick-end labeling-positive cells reduced after electroacupuncture treatment. Moreover, expression of adenosine monophosphate-activated protein kinase α (AMPKα) and phosphorylated AMPKα was up-regulated. Intraperitoneal injection of the AMPK antagonist, compound C, suppressed this phenomenon. Our findings suggest that electroacupuncture preconditioning alleviates ischemic brain injury via AMPK activation. PMID:26330828

  15. Viscothionin isolated from Korean mistletoe improves nonalcoholic fatty liver disease via the activation of adenosine monophosphate-activated protein kinase.

    PubMed

    Kim, Sokho; Lee, Dongho; Kim, Jae-Kyung; Kim, Jae-Hun; Park, Jong-Heum; Lee, Ju-Woon; Kwon, Jungkee

    2014-12-10

    The present study investigated the effects of viscothionin, a compound isolated from Korean mistletoe (Viscum album coloratum), on nonalcoholic fatty liver disease (NAFLD) in both in vitro and in vivo models. A connection was discovered between viscothionin and the adenosine monophosphate-activated protein kinase (AMPK) signaling pathway, which is involved in lipid metabolism. Viscothionin was shown to significantly attenuate lipid accumulation in HepG2 cells treated with oleic acid, which induces lipid accumulation. Moreover, the phosphorylation of AMPK and acetyl-coenzyme A carboxylase in HepG2 cells was increased by viscothionin treatment. Viscothionin was orally administered to high fat diet-induced obese mice and subsequently histopathological analysis associated with AMPK signaling pathways was evaluated. A significant reduction in the extent of hepatic steatosis was revealed in viscothionin-treated obese mice. Thus, viscothionin mediates its beneficial effects on NAFLD via AMPK signaling pathways, suggesting that it may be a potential target for novel NAFLD treatments.

  16. Adenosine monophosphate-activated protein kinase activation and suppression of inflammatory response by cell stretching in rabbit synovial fibroblasts.

    PubMed

    Kunanusornchai, Wanlop; Muanprasat, Chatchai; Chatsudthipong, Varanuj

    2016-12-01

    Joint mobilization is known to be beneficial in osteoarthritis (OA) patients. This study aimed to investigate the effect of stretching on adenosine monophosphate-activated protein kinase (AMPK) activity and its role in modulating inflammation in rabbit synovial fibroblasts. Uniaxial stretching of isolated rabbit synovial fibroblasts for ten min was performed. Stretching-induced AMPK activation, its underlying mechanism, and its anti-inflammatory effect were investigated using Western blot. Static stretching at 20 % of initial length resulted in AMPK activation characterized by expression of phosphorylated AMPK and phosphorylated acetyl-Co A carboxylase. AMP-activated protein kinase phosphorylation peaked 1 h after stretching and declined toward resting activity. Using cell viability assays, static stretching did not appear to cause cellular damage. Activation of AMPK involves Ca(2+) influx via a mechanosensitive L-type Ca(2+) channel, which subsequently raises intracellular Ca(2+) and activates AMPK via Ca(2+)/calmodulin-dependent protein kinase kinase β (CaMKKβ). Interestingly, stretching suppressed TNFα-induced expression of COX-2, iNOS, and phosphorylated NF-κB. These effects were prevented by pretreatment with compound C, an AMPK inhibitor. These results suggest that mechanical stretching suppressed inflammatory responses in synovial fibroblasts via a L-type Ca(2+)-channel-CaMKKβ-AMPK-dependent pathway which may underlie joint mobilization's ability to alleviate OA symptoms.

  17. Methylene blue induces macroautophagy through 5′ adenosine monophosphate-activated protein kinase pathway to protect neurons from serum deprivation

    PubMed Central

    Xie, Luokun; Li, Wenjun; Winters, Ali; Yuan, Fang; Jin, Kunlin; Yang, Shaohua

    2013-01-01

    Methylene blue has been shown to be neuroprotective in multiple experimental neurodegenerative disease models. However, the mechanisms underlying the neuroprotective effects have not been fully elucidated. Previous studies have shown that macroautophagy has multiple beneficial roles for maintaining normal cellular homeostasis and that induction of macroautophagy after myocardial ischemia is protective. In the present study we demonstrated that methylene blue could protect HT22 hippocampal cell death induced by serum deprivation, companied by induction of macroautophagy. We also found that methylene blue-mediated neuroprotection was abolished by macroautophagy inhibition. Interestingly, 5′ adenosine monophosphate-activated protein kinase (AMPK) signaling, but not inhibition of mammalian target of rapamycin signaling, was activated at 12 and 24 h after methylene blue treatment in a dose-dependent manner. Methylene blue-induced macroautophagy was blocked by AMPK inhibitor. Consistent with in vitro data, macroautophagy was induced in the cortex and hippocampus of mouse brains treated with methylene blue. Our findings suggest that methylene blue-induced neuroprotection is mediated, at least in part, by macroautophagy though activation of AMPK signaling. PMID:23653592

  18. Methylene blue induces macroautophagy through 5' adenosine monophosphate-activated protein kinase pathway to protect neurons from serum deprivation.

    PubMed

    Xie, Luokun; Li, Wenjun; Winters, Ali; Yuan, Fang; Jin, Kunlin; Yang, Shaohua

    2013-01-01

    Methylene blue has been shown to be neuroprotective in multiple experimental neurodegenerative disease models. However, the mechanisms underlying the neuroprotective effects have not been fully elucidated. Previous studies have shown that macroautophagy has multiple beneficial roles for maintaining normal cellular homeostasis and that induction of macroautophagy after myocardial ischemia is protective. In the present study we demonstrated that methylene blue could protect HT22 hippocampal cell death induced by serum deprivation, companied by induction of macroautophagy. We also found that methylene blue-mediated neuroprotection was abolished by macroautophagy inhibition. Interestingly, 5' adenosine monophosphate-activated protein kinase (AMPK) signaling, but not inhibition of mammalian target of rapamycin signaling, was activated at 12 and 24 h after methylene blue treatment in a dose-dependent manner. Methylene blue-induced macroautophagy was blocked by AMPK inhibitor. Consistent with in vitro data, macroautophagy was induced in the cortex and hippocampus of mouse brains treated with methylene blue. Our findings suggest that methylene blue-induced neuroprotection is mediated, at least in part, by macroautophagy though activation of AMPK signaling.

  19. Enhanced activation of NAD(P)H: quinone oxidoreductase 1 attenuates spontaneous hypertension by improvement of endothelial nitric oxide synthase coupling via tumor suppressor kinase liver kinase B1/adenosine 5'-monophosphate-activated protein kinase-mediated guanosine 5'-triphosphate cyclohydrolase 1 preservation.

    PubMed

    Kim, Yong-Hoon; Hwang, Jung Hwan; Kim, Kyung-Shim; Noh, Jung-Ran; Gang, Gil-Tae; Oh, Won Keun; Jeong, Kyeong-Hoon; Kwak, Tae Hwan; Choi, Hueng-Sik; Lee, In-Kyu; Lee, Chul-Ho

    2014-02-01

    Guanosine 5'-triphosphate cyclohydrolase-1 (GTPCH-1) is a rate-limiting enzyme in de-novo synthesis of tetrahydrobiopterin (BH4), an essential cofactor for endothelial nitric oxide synthase (eNOS) coupling. Adenosine 5'-monophosphate-activated protein kinase (AMPK) is crucial for GTPCH-1 preservation, and tumor suppressor kinase liver kinase B1 (LKB1), an upstream kinase of AMPK, is activated by NAD-dependent class III histone deacetylase sirtuin 1 (SIRT1)-mediated deacetylation. β-Lapachone has been shown to increase cellular NAD/NADH ratio via quinone oxidoreductase 1 (NQO1) activation. In this study, we have evaluated whether β-lapachone-induced NQO1 activation modulates blood pressure (BP) through preservation of GTPCH-1 in a hypertensive animal model. Spontaneously hypertensive rats (SHRs), primary aortic endothelial cells, and endothelial cell line were used to investigate the hypotensive effect of β-lapachone and its action mechanism. β-Lapachone treatment dramatically lowered BP and vascular tension in SHRs and induced eNOS activation in endothelial cells. Consistent with these effects, β-lapachone treatment also elevated levels of both aortic cGMP and plasma nitric oxide in SHRs. Meanwhile, β-lapachone-treated SHRs showed significantly increased levels of aortic NAD, LKB1 deacetylation, and AMPK Thr phosphorylation followed by increased GTPCH-1 and tetrahydrobiopterin/dihydrobiopterin ratio. In-vitro study revealed that AMPK inhibition by overexpression of dominant-negative AMPK nearly abolished GTPCH-1 protein conservation. Enhanced LKB1 deacetylation and AMPK activation were also elicited by β-lapachone in endothelial cells. However, inhibition of LKB1 deacetylation by blocking of NQO1 or SIRT1 blunted AMPK activation by β-lapachone. This is the first study demonstrating that eNOS coupling can be regulated by NQO1 activation via LKB1/AMPK/GTPCH-1 modulation, which is possibly correlated with relieving hypertension. These findings provide strong

  20. Curcumin inhibits aerobic glycolysis in hepatic stellate cells associated with activation of adenosine monophosphate-activated protein kinase.

    PubMed

    Lian, Naqi; Jin, Huanhuan; Zhang, Feng; Wu, Li; Shao, Jiangjuan; Lu, Yin; Zheng, Shizhong

    2016-07-01

    Activation of hepatic stellate cells (HSCs) is characterized by expression of extracellular matrix and loss of adipogenic phenotype during liver fibrogenesis. Emerging evidence suggests that HSCs adopt aerobic glycolysis during activation. The present work aimed at investigating whether the anti-fibrogenic effects of curcumin was associated with interfering with glycolysis in HSCs. Primary rat HSCs were cultured in vitro. We demonstrated that inhibition of glycolysis by 2-deoxyglucose or galloflavin reduced the expression of α-smooth muscle actin (α-SMA) and α1(I)procollagen at both mRNA and protein levels, and increased the intracellular lipid contents and upregulated the gene and protein expression of adipogenic transcription factors C/EBPα and PPAR-γ in HSCs. Curcumin at 20 μM produced similar effects. Moreover, curcumin decreased the expression of hexokinase (HK), phosphofructokinase-2 (PFK2), and glucose transporter 4 (glut4), three key glycolytic parameters, at both mRNA and protein levels. Curcumin also reduced lactate production concentration-dependently in HSCs. Furthermore, curcumin increased the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), but AMPK inhibitor BML-275 significantly abolished the curcumin downregulation of HK, PFK2, and glut4. In addition, curcumin inhibition of α-SMA and α1(I)procollagen was rescued by BML-275, and curcumin upregulation of C/EBPα and PPAR-γ was abrogated by BML-275. These results collectively indicated that curcumin inhibited glycolysis in an AMPK activation-dependent manner in HSCs. We revealed a novel mechanism for curcumin suppression of HSC activation implicated in antifibrotic therapy. © 2016 IUBMB Life, 68(7):589-596, 2016. © 2016 International Union of Biochemistry and Molecular Biology.

  1. Neuroprotective effect of activated 5'-adenosine monophosphate-activated protein kinase on cone system function during retinal inflammation.

    PubMed

    Kamoshita, Mamoru; Fujinami, Kaoru; Toda, Eriko; Tsubota, Kazuo; Ozawa, Yoko

    2016-06-10

    Retinal inflammation can cause retinal neural disorders. In particular, functional disorder in the cone photoreceptor system influences visual acuity. However, the underlying mechanism is not yet fully understood. In this study, we evaluated cone system function and the role of 5'-adenosine monophosphate-activated protein kinase (AMPK) during retinal inflammation. Six to eight-week-old male C57BL/6 mice received an intraperitoneal injection of lipopolysaccharide (LPS) to induce retinal inflammation, and were treated with an AMPK activator, 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR; 250 mg/kg body weight) or phosphate-buffered saline as vehicle 3 h before the LPS injection. The b-wave of the photopic electroretinogram, which represents cone system function, was decreased 24 h after LPS injection and this reduction was suppressed by AICAR treatment. At this time point, there was no remarkable morphological change in the cone photoreceptor cells. At 1.5 h after LPS injection, the retina mRNA levels of an inflammatory cytokine, Tnf-α, were increased, and those of a regulator of mitochondrial biogenesis, Pgc1-α, were decreased. However, AICAR treatment suppressed these changes in mRNA expression. Immunohistochemistry showed that induction of glial fibrillary acidic protein expression was also suppressed by AICAR 24 h after LPS injection. Furthermore, the mouse cone photoreceptor-derived cell line 661W was treated with AICAR to increase the level of phosphorylated and activated AMPK. After 3 h of AICAR incubation, 661W cells showed decreased Tnf-α mRNA levels and increased Pgc1-α mRNA levels. AMPK activation has a neuroprotective effect on cone system function during inflammation, and the effect may, at least in part, involve the regulation of inflammatory cytokines and mitochondrial condition.

  2. Role of activation of 5'-adenosine monophosphate-activated protein kinase in gastric ulcer healing in diabetic rats.

    PubMed

    Baraka, Azza M; Deif, Maha M

    2011-01-01

    The potential utility of 5'-adenosine monophosphate-activated protein kinase (AMPK)-activating agents, such as metformin, in inducing angiogenesis, could be a promising approach to promote healing of gastric ulcers complicated by diabetes mellitus. The aim of the present study was to assess the effect of a drug that activates AMPK, namely metformin, in gastric ulcer healing in streptozotocin-induced diabetic rats. Forty male Wistar albino rats were made diabetic by intraperitoneal (i.p.) streptozotocin injection and 10 rats were injected i.p. by a single dose of physiological saline. Six weeks following streptozotocin or saline injection, gastric ulcers were induced by serosal application of acetic acid. Three days after acetic acid application, rats were divided into group 1 (nondiabetic control), group 2 (streptozotocin-injected rats), groups 3-5 (streptozotocin-injected rats treated with metformin or metformin and an inhibitor of AMPK, namely compound C or pioglitazone) for 7 days following acetic acid application. Administration of metformin, but not pioglitazone, resulted in a significant decrease in the gastric ulcer area, a significant increase in epithelial regeneration assessed histologically, a significant increase in the number of microvessels in the ulcer margin, a significant increase in gastric vascular endothelial growth factor concentration and gastric von Willebrand factor as well as a significant increase in gastric phospho-AMPK. Compound C, an inhibitor of AMPK, blocked metformin-induced changes in assessed parameters suggesting that the effect of metformin was mediated mainly through activation of AMPK. Our results suggest the feasibility of a novel treatment strategy, namely drugs activating AMPK, for patients in whom impairment of ulcer healing constitutes a secondary complication of diabetes mellitus. Copyright © 2011 S. Karger AG, Basel.

  3. Estradiol regulation of hypothalamic astrocyte adenosine 5'-monophosphate-activated protein kinase activity: role of hindbrain catecholamine signaling.

    PubMed

    Tamrakar, Pratistha; Briski, Karen P

    2015-01-01

    Recent work challenges the conventional notion that metabolic monitoring in the brain is the exclusive function of neurons. This study investigated the hypothesis that hypothalamic astrocytes express the ultra-sensitive energy gauge adenosine 5'-monophosphate-activated protein kinase (AMPK), and that the ovarian hormone estradiol (E) controls activation of this sensor by insulin-induced hypoglycemia (IIH). E- or oil (O)-implanted ovariectomized (OVX) rats were pretreated by caudal fourth ventricular administration of the catecholamine neurotoxin 6-hydroxydopamine (6-OHDA) prior to sc insulin or vehicle injection. Individual astrocytes identified in situ by glial fibrillary acidic protein immunolabeling were laser-microdissected from the ventromedial (VMH), arcuate (ARH), and paraventricular (PVH) nuclei and the lateral hypothalamic area (LHA), and pooled within each site for Western blot analysis of AMPK and phosphoAMPK (pAMPK) protein expression. In the VMH, baseline astrocyte AMPK and pAMPK levels were respectively increased or decreased in OVX+E versus OVX+O; these profiles did not differ between E and O rats in other hypothalamic loci. In E animals, astrocyte AMPK protein was reduced [VMH] or augmented [PVH; LHA] in response to either 6-OHDA or IIH. IIH increased astrocyte pAMPK expression in each structure in vehicle-, but not 6-OHDA-pretreated E rats. Results provide novel evidence for hypothalamic astrocyte AMPK expression and hindbrain catecholamine-dependent activation of this cell-specific sensor by hypoglycemia in the presence of estrogen. Further research is needed to determine the role of astrocyte AMPK in reactivity of these glia to metabolic imbalance and contribution to restoration of neuro-metabolic stability.

  4. Adenosine monophosphate-activated protein kinase (AMPK) activators for the prevention, treatment and potential reversal of pathological pain

    PubMed Central

    Price, Theodore J.; Das, Vaskar; Dussor, Gregory

    2015-01-01

    Pathological pain is an enormous medical problem that places a significant burden on patients and can result from an injury that has long since healed or be due to an unidentifiable cause. Although treatments exist, they often either lack efficacy or have intolerable side effects. More importantly, they do not reverse the changes in the nervous system mediating pathological pain, and thus symptoms often return when therapies are discontinued. Consequently, novel therapies are urgently needed that have both improved efficacy and disease-modifying properties. Here we highlight an emerging target for novel pain therapies, adenosine monophosphate-activated protein kinase (AMPK). AMPK is capable of regulating a variety of cellular processes including protein translation, activity of other kinases, and mitochondrial metabolism, many of which are thought to contribute to pathological pain. Consistent with these properties, preclinical studies show positive, and in some cases disease-modifying effects of either pharmacological activation or genetic regulation of AMPK in models of nerve injury, chemotherapy-induced peripheral neuropathy (CIPN), postsurgical pain, inflammatory pain, and diabetic neuropathy. Given the AMPK-activating ability of metformin, a widely prescribed and well-tolerated drug, these preclinical studies provide a strong rationale for both retrospective and prospective human pain trials with this drug. They also argue for the development of novel AMPK activators, whether orthosteric, allosteric, or modulators of events upstream of the kinase. Together, this review will present the case for AMPK as a novel therapeutic target for pain and will discuss future challenges in the path toward development of AMPK-based pain therapeutics. PMID:26521775

  5. Donor pretreatment with adenosine monophosphate-activated protein kinase activator protects cardiac grafts from cold ischaemia/reperfusion injury.

    PubMed

    Yang, Chao; Xu, Honglai; Cai, Lanjun; Du, Xiaoxiao; Jiang, Yinan; Zhang, Yong; Zhou, Hongmin; Chen, Zhonghua Klaus

    2016-05-01

    Adenosine monophosphate-activated protein kinase (AMPK) is a master regulator of energy metabolism and has been shown to be protective in ischaemia/reperfusion injury (IRI). We hypothesized that preactivation of AMPK with an activator before donor heart procurement could protect heart grafts from cold IRI. Donor Sprague-Dawley rats were injected intravenously with AMPK activator 5-amino-imidazole-4-carboxamide ribonucleotide (AICAR) or vehicle 30 min before heart procurement. Heart grafts were then preserved in histidine-tryptophan-ketoglutarate (HTK) solution at 4°C for 8 h. After preservation, grafts were immediately mounted on the Langendorff perfusion system and perfused with Krebs-Henseleit buffer at 37°C for 1 h. Adenosine triphosphate (ATP) and malondialdehyde (MDA) content in graft tissue were quantified post-preservation and post-reperfusion. After reperfusion, isolated heart function was assessed using a pressure transducer; cumulative release of creatine kinase (CK) and lactate dehydrogenase (LDH) into the perfusate was measured to assess cardiomyocyte necrosis; ultrastructural changes in the mitochondria of the grafts were examined using transmission electron microscopy (TEM). After preservation, myocardial ATP content in the pretreated hearts was significantly higher than in the control hearts (3.247 ± 0.3034 vs 1.817 ± 0.2533 µmol/g protein; P < 0.05). AICAR-pretreated heart grafts exhibited significantly higher coronary flow (9.667 ± 0.3159 vs 8.033 ± 0.2459 ml/min; P < 0.05) and left ventricular developing pressure (58.67 ± 2.894 vs 42.67 ± 3.333 mmHg; P < 0.05) than the vehicle treated after reperfusion. Cumulative release of CK (300.0 ± 25.30 vs 431.7 ± 42.39 U/l; P < 0.05) and LDH (228.0 ± 16.68 vs 366.8 ± 57.41 U/l; P < 0.05) in the perfusate was significantly lower in the AICAR-pretreated group than that in the control group. Myocardial MDA content was also reduced in the pretreated group (0.5167 ± 0.1046 vs 0.9333 ± 0.1333 nmol

  6. 5-Aminoimidazole-4-carboxamide ribonucleoside-mediated adenosine monophosphate-activated protein kinase activation induces protective innate responses in bacterial endophthalmitis.

    PubMed

    Kumar, Ajay; Giri, Shailendra; Kumar, Ashok

    2016-12-01

    The retina is considered to be the most metabolically active tissue in the body. However, the link between energy metabolism and retinal inflammation, as incited by microbial infection such as endophthalmitis, remains unexplored. In this study, using a mouse model of Staphylococcus aureus (SA) endophthalmitis, we demonstrate that the activity (phosphorylation) of 5' adenosine monophosphate-activated protein kinase alpha (AMPKα), a cellular energy sensor and its endogenous substrate; acetyl-CoA carboxylase is down-regulated in the SA-infected retina. Intravitreal administration of an AMPK activator, 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), restored AMPKα and acetyl-CoA carboxylase phosphorylation. AICAR treatment reduced both the bacterial burden and intraocular inflammation in SA-infected eyes by inhibiting NF-kB and MAP kinases (p38 and JNK) signalling. The anti-inflammatory effects of AICAR were diminished in eyes pretreated with AMPK inhibitor, Compound C. The bioenergetics (Seahorse) analysis of SA-infected microglia and bone marrow-derived macrophages revealed an increase in glycolysis, which was reinstated by AICAR treatment. AICAR also reduced the expression of SA-induced glycolytic genes, including hexokinase 2 and glucose transporter 1 in microglia, bone marrow-derived macrophages and the mouse retina. Interestingly, AICAR treatment enhanced the bacterial phagocytic and intracellular killing activities of cultured microglia, macrophages and neutrophils. Furthermore, AMPKα1 global knockout mice exhibited increased susceptibility towards SA endophthalmitis, as evidenced by increased inflammatory mediators and bacterial burden and reduced retinal function. Together, these findings provide the first evidence that AMPK activation promotes retinal innate defence in endophthalmitis by modulating energy metabolism and that it can be targeted therapeutically to treat ocular infections.

  7. Effect of dietary nonstructural carbohydrate content on activation of 5'-adenosine monophosphate-activated protein kinase in liver, skeletal muscle, and digital laminae of lean and obese ponies.

    PubMed

    Burns, T A; Watts, M R; Weber, P S; McCutcheon, L J; Geor, R J; Belknap, J K

    2014-01-01

    In EMS-associated laminitis, laminar failure may occur in response to energy failure related to insulin resistance (IR) or to the effect of hyperinsulinemia on laminar tissue. 5'-Adenosine-monophosphate-activated protein kinase (AMPK) is a marker of tissue energy deprivation, which may occur in IR. To characterize tissue AMPK regulation in ponies subjected to a dietary carbohydrate (CHO) challenge. Twenty-two mixed-breed ponies. Immunohistochemistry and immunoblotting for total AMPK and phospho(P)-AMPK and RT-qPCR for AMPK-responsive genes were performed on laminar, liver, and skeletal muscle samples collected after a 7-day feeding protocol in which ponies stratified on body condition score (BCS; obese or lean) were fed either a low-CHO diet (ESC + starch, approximately 7% DM; n = 5 obese, 5 lean) or a high-CHO diet (ESC + starch, approximately 42% DM; n = 6 obese, 6 lean). 5'-Adenosine-monophosphate-activated protein kinase was immunolocalized to laminar keratinocytes, dermal constituents, and hepatocytes. A high-CHO diet resulted in significantly decreased laminar [P-AMPK] in lean ponies (P = .03), but no changes in skeletal muscle (lean, P = .33; obese, P = .43) or liver (lean, P = .84; obese, P = .13) [P-AMPK]. An inverse correlation existed between [blood glucose] and laminar [P-AMPK] in obese ponies on a high-CHO diet. Laminar tissue exhibited a normal response to a high-CHO diet (decreased [P-AMPK]), whereas this response was not observed in liver and skeletal muscle in both lean (skeletal muscle, P = .33; liver, P = .84) and obese (skeletal muscle, P = .43; liver, P = .13) ponies. Copyright © 2014 by the American College of Veterinary Internal Medicine.

  8. GROWTH AND DEVELOPMENT SYMPOSIUM: Adenosine monophosphate-activated protein kinase and mitochondria in Rendement Napole pig growth.

    PubMed

    Scheffler, T L; Gerrard, D E

    2016-09-01

    The Rendement Napole mutation (RN-), which is well known to influence pork quality, also has a profound impact on metabolic characteristics of muscle. Pigs with RN- possess a SNP in the γ3 subunit of adenosine monophosphate (AMP)-activated protein kinase (AMPK); AMPK, a key energy sensor in skeletal muscle, modulates energy producing and energy consuming pathways to maintain cellular homeostasis. Importantly, AMPK regulates not only acute response to energy stress but also facilitates long-term adaptation via changes in gene and protein expression. The RN- allele increases AMPK activity, which alters the metabolic phenotype of skeletal muscle by increasing mitochondrial content and oxidative capacity. Fibers with greater oxidative capacity typically exhibit increased protein turnover and smaller fiber size, which indicates that RN- pigs may exhibit decreased efficiency and growth potential. However, whole body and muscle growth of RN- pigs appear similar to that of wild-type pigs and despite increased oxidative capacity, fibers maintain the capacity for hypertrophic growth. This indicates that compensatory mechanisms may allow RN- pigs to achieve rates of muscle growth similar to those of wild-type pigs. Intriguingly, lipid oxidation and mitochondria function are enhanced in RN- pig muscle. Thus far, characteristics of RN- muscle are largely based on animals near market weight. To better understand interaction between energy signaling and protein accretion in muscle, further work is needed to define age-dependent relationships between AMPK signaling, metabolism, and muscle growth.

  9. Caffeine acutely activates 5'adenosine monophosphate-activated protein kinase and increases insulin-independent glucose transport in rat skeletal muscles.

    PubMed

    Egawa, Tatsuro; Hamada, Taku; Kameda, Naoko; Karaike, Kouhei; Ma, Xiao; Masuda, Shinya; Iwanaka, Nobumasa; Hayashi, Tatsuya

    2009-11-01

    Caffeine (1,3,7-trimethylxanthine) has been implicated in the regulation of glucose and lipid metabolism including actions such as insulin-independent glucose transport, glucose transporter 4 expression, and fatty acid utilization in skeletal muscle. These effects are similar to the exercise-induced and 5'adenosine monophosphate-activated protein kinase (AMPK)-mediated metabolic changes in skeletal muscle, suggesting that caffeine is involved in the regulation of muscle metabolism through AMPK activation. We explored whether caffeine acts on skeletal muscle to stimulate AMPK. Incubation of rat epitrochlearis and soleus muscles with Krebs buffer containing caffeine (> or =3 mmol/L, > or =15 minutes) increased the phosphorylation of AMPKalpha Thr(172), an essential step for full kinase activation, and acetyl-coenzyme A carboxylase Ser(79), a downstream target of AMPK, in dose- and time-dependent manners. Analysis of isoform-specific AMPK activity revealed that both AMPKalpha1 and alpha2 activities increased significantly. This enzyme activation was associated with a reduction in phosphocreatine content and an increased rate of 3-O-methyl-d-glucose transport activity in the absence of insulin. These results suggest that caffeine has similar actions to exercise by acutely stimulating skeletal muscle AMPK activity and insulin-independent glucose transport with a reduction of the intracellular energy status.

  10. Estrogen regulates energy metabolic pathway and upstream adenosine 5'-monophosphate-activated protein kinase and phosphatase enzyme expression in dorsal vagal complex metabolosensory neurons during glucostasis and hypoglycemia.

    PubMed

    Tamrakar, Pratistha; Ibrahim, Baher A; Gujar, Amit D; Briski, Karen P

    2015-02-01

    The ability of estrogen to shield the brain from the bioenergetic insult hypoglycemia is unclear. Estradiol (E) prevents hypoglycemic activation of the energy deficit sensor adenosine 5'-monophosphate-activated protein kinase (AMPK) in hindbrain metabolosensory A2 noradrenergic neurons. This study investigates the hypothesis that estrogen regulates A2 AMPK through control of fuel metabolism and/or upstream protein kinase/phosphatase enzyme expression. A2 cells were harvested by laser microdissection after insulin or vehicle (V) injection of E- or oil (O)-implanted ovariectomized female rats. Cell lysates were evaluated by immunoblot for glycolytic, tricarboxylic acid cycle, respiratory chain, and acetyl-CoA-malonyl-CoA pathway enzymes. A2 phosphofructokinase (PFKL), isocitrate dehydrogenase, pyruvate dehydrogenase, and ATP synthase subunit profiles were elevated in E/V vs. O/V; hypoglycemia augmented PFKL and α-ketoglutarate dehydrogenase expression in E only. Hypoglycemia increased A2 Ca(2+) /calmodulin-dependent protein kinase-β in O and reduced protein phosphatase in both groups. A2 phospho-AMPK levels were equivalent in O/V vs. E/V but elevated during hypoglycemia in O only. These results implicate E in compensatory upregulation of substrate catabolism and corresponding maintenance of energy stability of A2 metabolosensory neurons during hypoglycemia, outcomes that support the potential viability of molecular substrates for hormone action as targets for therapies alleviating hypoglycemic brain injury.

  11. Adenosine monophosphate-activated protein kinase involved in variations of muscle glycogen and breast meat quality between lean and fat chickens.

    PubMed

    Sibut, V; Le Bihan-Duval, E; Tesseraud, S; Godet, E; Bordeau, T; Cailleau-Audouin, E; Chartrin, P; Duclos, M J; Berri, C

    2008-11-01

    The present study was aimed at evaluating the molecular mechanisms associated with the differences in muscle glycogen content and breast meat quality between 2 experimental lines of chicken divergently selected on abdominal fatness. The glycogen at death (estimated through the glycolytic potential) of the pectoralis major muscle and the quality of the resulting meat were estimated in the 2 lines. The fat chickens exhibited greater glycolytic potential, and in turn lower ultimate pH than the lean chickens. Consequently, the breast meat of fat birds was paler and less colored (i.e., less red and yellow), and exhibited greater drip loss compared with that of lean birds. In relation to these variations, transcription and activation levels of adenosine monophosphate-activated protein kinase (AMPK) were investigated. The main difference observed between lines was a 3-fold greater level of AMPK activation, evaluated through phosphorylation of AMPKalpha-(Thr(172)), in the muscle of lean birds. At the transcriptional level, data indicated concomitant down- and upregulation for the gamma1 and gamma2 AMPK subunit isoforms, respectively, in the muscle of lean chickens. Transcriptional levels of enzymes directly involved in glycogen turnover were also investigated. Data showed greater gene expression for glycogen synthase, glycogen phosphorylase, and the gamma subunit of phosphorylase kinase in lean birds. Together, these data indicate that selection on body fatness in chicken alters the muscle glycogen turnover and content and consequently the quality traits of the resulting meat. Alterations of AMPK activity could play a key role in these changes.

  12. Adenosine Monophosphate-Activated Protein Kinase (AMPK) as a New Target for Antidiabetic Drugs: A Review on Metabolic, Pharmacological and Chemical Considerations

    PubMed Central

    Gruzman, Arie; Babai, Gali; Sasson, Shlomo

    2009-01-01

    In view of the epidemic nature of type 2 diabetes and the substantial rate of failure of current oral antidiabetic drugs the quest for new therapeutics is intensive. The adenosine monophosphate-activated protein kinase (AMPK) is an important regulatory protein for cellular energy balance and is considered a master switch of glucose and lipid metabolism in various organs, especially in skeletal muscle and liver. In skeletal muscles, AMPK stimulates glucose transport and fatty acid oxidation. In the liver, it augments fatty acid oxidation and decreases glucose output, cholesterol and triglyceride synthesis. These metabolic effects induced by AMPK are associated with lowering blood glucose levels in hyperglycemic individuals. Two classes of oral antihyperglycemic drugs (biguanidines and thiazolidinediones) have been shown to exert some of their therapeutic effects by directly or indirectly activating AMPK. However, side effects and an acquired resistance to these drugs emphasize the need for the development of novel and efficacious AMPK activators. We have recently discovered a new class of hydrophobic D-xylose derivatives that activates AMPK in skeletal muscles in a non insulin-dependent manner. One of these derivatives (2,4;3,5-dibenzylidene-D-xylose-diethyl-dithioacetal) stimulates the rate of hexose transport in skeletal muscle cells by increasing the abundance of glucose transporter-4 (GLUT-4) in the plasma membrane through activation of AMPK. This compound reduces blood glucose levels in diabetic mice and therefore offers a novel strategy of therapeutic intervention strategy in type 2 diabetes. The present review describes various classes of chemically-related compounds that activate AMPK by direct or indirect interactions and discusses their potential for candidate antihyperglycemic drug development. PMID:19557293

  13. A high isoflavone diet decreases 5' adenosine monophosphate-activated protein kinase activation and does not correct selenium-induced elevations in fasting blood glucose in mice.

    PubMed

    Stallings, Michael T; Cardon, Brandon R; Hardman, Jeremy M; Bliss, Tyler A; Brunson, Scott E; Hart, Chris M; Swiss, Maria D; Hepworth, Squire D; Christensen, Merrill J; Hancock, Chad R

    2014-04-01

    Selenium (Se) has been implicated as a micronutrient that decreases adenosine monophosphate-activated protein kinase (AMPK) signaling and may increase diabetes risk by reducing insulin sensitivity. Soy isoflavones (IF) are estrogen-like compounds that have been shown to attenuate insulin resistance, hyperglycemia, adiposity, and increased AMPK activation. We hypothesized that a high IF (HIF) diet would prevent the poor metabolic profile associated with high Se intake. The purpose of this study was to examine changes in basal glucose metabolism and AMPK signaling in response to an HIF diet and/or supplemental Se in a mouse model. Male FVB mice were divided into groups receiving either a control diet with minimal IF (low IF) or an HIF diet. Each dietary group was further subdivided into groups receiving either water or Se at a dose of 3 mg Se/kg body weight daily, as Se-methylselenocysteine (SMSC). After 5 months, mice receiving SMSC had elevated fasting glucose (P < .05) and a tendency for glucose intolerance (P = .08). The increase in dietary IF did not result in improved fasting blood glucose. Interestingly, after 6 months, HIF-fed mice had decreased basal AMPK activation in liver and skeletal muscle tissue (P < .05). Basal glucose metabolism was changed by SMSC supplementation as evidenced by increased fasting blood glucose and glucose intolerance. High dietary IF levels did not protect against aberrant blood glucose. In FVB mice, decreased basal AMPK activation is not the mechanism through which Se exerts its effect. These results suggest that more research must be done to elucidate the role of Se and IF in glucose metabolism.

  14. Metformin inhibits growth of human non-small cell lung cancer cells via liver kinase B-1-independent activation of adenosine monophosphate-activated protein kinase

    PubMed Central

    GUO, QIANQIAN; LIU, ZHIYAN; JIANG, LILI; LIU, MENGJIE; MA, JIEQUN; YANG, CHENGCHENG; HAN, LILI; NAN, KEJUN; LIANG, XUAN

    2016-01-01

    Metformin, the most widely administered oral anti-diabetic therapeutic agent, exerts its glucose-lowering effect predominantly via liver kinase B1 (LKB1)-dependent activation of adenosine monophosphate-activated protein kinase (AMPK). Accumulating evidence has demonstrated that metformin possesses potential antitumor effects. However, whether the antitumor effect of metformin is via the LKB1/AMPK signaling pathway remains to be determined. In the current study, the effects of metformin on proliferation, cell cycle progression, and apoptosis of human non-small cell lung cancer (NSCLC) H460 (LKB1-null) and H1299 (LKB1-positive) cells were assessed, and the role of LKB1/AMPK signaling in the anti-growth effects of metformin were investigated. Cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, cell cycle distribution and apoptosis were assessed by flow cytometry, and protein expression levels were measured by western blotting. Metformin inhibited proliferation, induced significant cell cycle arrest at the G0–G1 phase and increased apoptosis in NSCLC cells in a time- and concentration-dependent manner, regardless of the level of LKB1 protein expression. Furthermore, knockdown of LKB1 with short hairpin RNA (shRNA) did not affect the antiproliferative effect of metformin in the H1299 cells. Metformin stimulated AMPK phosphorylation and subsequently suppressed the phosphorylation of mammalian target of rapamycin and its downstream effector, 70-kDa ribosomal protein S6 kinase in the two cell lines. These effects were abrogated by silencing AMPK with small interfering RNA (siRNA). In addition, knockdown of AMPK with siRNA inhibited the effect of metformin on cell proliferation in the two cell lines. These results provide evidence that the growth inhibition of metformin in NSCLC cells is mediated by LKB1-independent activation of AMPK, indicating that metformin may be a potential therapeutic agent for the treatment of

  15. Activation of 5' adenosine monophosphate-activated protein kinase blocks cumulus cell expansion through inhibition of protein synthesis during in vitro maturation in Swine.

    PubMed

    Santiquet, Nicolas; Sasseville, Maxime; Laforest, Martin; Guillemette, Christine; Gilchrist, Robert B; Richard, François J

    2014-08-01

    The serine/threonine kinase 5' adenosine monophosphate-activated protein kinase (AMPK), a heterotrimeric protein known as a metabolic switch, is involved in oocyte nuclear maturation in mice, cattle, and swine. The present study analyzed AMPK activation in cumulus cell expansion during in vitro maturation (IVM) of porcine cumulus-oocyte complexes (COC). 5-Aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR) is a well-known activator of AMPK. It inhibited oocyte meiotic resumption in COC. Moreover, cumulus cell expansion did not occur in the presence of AICAR, demonstrating its marked impact on cumulus cells. Activation of AMPK was supported by AICAR-mediated phosphorylation of alpha AMPK subunits. Furthermore, the presence of AICAR increased glucose uptake, a classical response to activation of this metabolic switch in response to depleted cellular energy levels. Neither nuclear maturation nor cumulus expansion was reversed by glucosamine, an alternative substrate in hyaluronic acid synthesis, through the hexosamine biosynthetic pathway, which ruled out possible depletion of substrates. Both increased gap junction communication and phosphodiesterase activity in COC are dependent on protein synthesis during the initial hours of IVM; however, both were inhibited in the presence of AICAR, which supports the finding that activation of AMPK by AICAR mediated inhibition of protein synthesis. Moreover, this protein synthesis inhibition was equivalent to that of the well-known protein synthesis inhibitor cycloheximide, as observed on cumulus expansion and protein concentration. Finally, the phosphorylation level of selected kinases was investigated. The pattern of raptor phosphorylation is supportive of activation of AMPK-mediated inhibition of protein synthesis. In conclusion, AICAR-mediated AMPK activation in porcine COC inhibited cumulus cell expansion and protein synthesis. These results bring new considerations to the importance of this kinase in ovarian

  16. Metformin decreases GnRH- and activin-induced gonadotropin secretion in rat pituitary cells: potential involvement of adenosine 5' monophosphate-activated protein kinase (PRKA).

    PubMed

    Tosca, Lucie; Froment, Pascal; Rame, Christelle; McNeilly, J R; McNeilly, A S; Maillard, Virginie; Dupont, Joelle

    2011-02-01

    Metformin is an insulin sensitizer molecule used for the treatment of infertility in women with polycystic ovary syndrome and insulin resistance. It modulates the reproductive axis, affecting the release of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH). However, metformin's mechanism of action in pituitary gonadotropin-secreting cells remains unclear. Adenosine 5' monophosphate-activated protein kinase (PRKA) is involved in metformin action in various cell types. Here, we investigated the effects of metformin on gonadotropin secretion in response to activin and GnRH in primary rat pituitary cells (PRP), and studied PRKA in rat pituitary. In PRP, metformin (10 mM) reduced LH and follicle-stimulating hormone (FSH) secretion induced by GnRH (10(-8) M, 3 h), FSH secretion, and mRNA FSHbeta subunit expression induced by activin (10(-8) M, 12 or 24 h). The different subunits of PRKA are expressed in pituitary. In particular, PRKAA1 is detected mainly in gonadotrophs and thyrotrophs, is less abundant in lactotrophs and somatotrophs, and is undetectable in corticotrophs. In PRP, metformin increased phosphorylation of both PRKA and acetyl-CoA carboxylase. Metformin decreased activin-induced SMAD2 phosphorylation and GnRH-induced mitogen-activated protein kinase (MAPK) 3/1 (ERK1/2) phosphorylation. The PRKA inhibitor compound C abolished the effects of metformin on gonadotropin release induced by GnRH and on FSH secretion and Fshb mRNA induced by activin. The adenovirus-mediated production of dominant negative PRKA abolished the effects of metformin on the FSHbeta subunit mRNA and SMAD2 phosphorylation induced by activin and on the MAPK3/1 phosphorylation induced by GnRH. Thus, in rat pituitary cells, metformin decreases gonadotropin secretion and MAPK3/1 phosphorylation induced by GnRH and FSH release, FSHbeta subunit expression, and SMAD2 phosphorylation induced by activin through PRKA activation.

  17. Involvement of adenosine monophosphate-activated protein kinase in the influence of timed high-fat evening diet on the hepatic clock and lipogenic gene expression in mice.

    PubMed

    Huang, Yan; Zhu, Zengyan; Xie, Meilin; Xue, Jie

    2015-09-01

    A high-fat diet may result in changes in hepatic clock gene expression, but potential mechanisms are not yet elucidated. Adenosine monophosphate-activated protein kinase (AMPK) is a serine/threonine protein kinase that is recognized as a key regulator of energy metabolism and certain clock genes. Therefore, we hypothesized that AMPK may be involved in the alteration of hepatic clock gene expression under a high-fat environment. This study aimed to examine the effects of timed high-fat evening diet on the activity of hepatic AMPK, clock genes, and lipogenic genes. Mice with hyperlipidemic fatty livers were induced by orally administering high-fat milk via gavage every evening (19:00-20:00) for 6 weeks. Results showed that timed high-fat diet in the evening not only decreased the hepatic AMPK protein expression and activity but also disturbed its circadian rhythm. Accordingly, the hepatic clock genes, including clock, brain-muscle-Arnt-like 1, cryptochrome 2, and period 2, exhibited prominent changes in their expression rhythms and/or amplitudes. The diurnal rhythms of the messenger RNA expression of peroxisome proliferator-activated receptorα, acetyl-CoA carboxylase 1α, and carnitine palmitoyltransferase 1 were also disrupted; the amplitude of peroxisome proliferator-activated receptorγcoactivator 1α was significantly decreased at 3 time points, and fatty liver was observed. These findings demonstrate that timed high-fat diet at night can change hepatic AMPK protein levels, activity, and circadian rhythm, which may subsequently alter the circadian expression of several hepatic clock genes and finally result in the disorder of hepatic lipogenic gene expression and the formation of fatty liver. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Effect of the growth stage and cultivar on policosanol profiles of barley sprouts and their adenosine 5'-monophosphate-activated protein kinase activation.

    PubMed

    Seo, Woo Duck; Yuk, Heung Joo; Curtis-Long, Marcus J; Jang, Ki Chang; Lee, Jin Hwan; Han, Sang-Ik; Kang, Hang Won; Nam, Min Hee; Lee, Sung-Joon; Lee, Ji Hae; Park, Ki Hun

    2013-02-06

    Adenosine 5'-monophosphate-activated protein kinase (AMPK) is an intracellular sensor that can regulate glucose levels within the cell. For this reason, it is well-known to be a target for drugs against diabetes and obesity. AMPK was activated significantly by the hexane extract of barley sprouts. This AMPK activation emerges across the growth stages of the sprout, becoming most significant (3 times above the initial stages) 10 days after sprouting. After this time, the activation decreased between 13 and 20 days post-sprouting. Analysis of the hexane extracts by gas chromatography-mass spectrometry showed that the amounts of policosanols (PCs, which are linear, primary aliphatic alcohols with 20-30 carbons) in the plant dramatically increased between 5 days (109.7 mg/100 g) and 10 days (343.7 mg/100 g) post-sprouting and then levels fell back down, reaching 76.4 mg/100 g at 20 days post-sprouting. This trend is consistent with PCs being the active ingredient in the barley plants. We validate this by showing that hexacosanol is an activator of AMPK. The richest cultivar for PCs was found to be the Daejin cultivar. Cultivars had a significant effect on the total PC content (113.2-183.5 mg/100 g) within the plant up to 5 days post-sprouting. However this dependence upon the cultivar was not so apparent at peak stages of PC production (10 days post-sprouting). The most abundant PC in barley sprout, hexacosanol, contributed 62-80% of the total PC content at every stage. These results are valuable to determine the optimal times of harvest to obtain the highest yield of PCs.

  19. [Alteration of metabolic characteristics on the masseter muscle fiber of unilateral chewing rats and its adenosine monophosphate activated protein kinase regulatory mechanism].

    PubMed

    Andi, Shi; Lin, Zeng; Jing, Liu

    2017-06-01

    This study aims to determine the influence of unilateral chewing on metabolic characteristics of masseter muscle fibers in rats and the regulatory effect of an adenosine monophosphate activated protein kinase (AMPK) signal pathway on metabolism. Rats were submitted to exodontia of all the right maxillary molars and divided into 2, 4, 6, and 8 weeks groups, and corresponding control groups were set as well. Sections were stained by nicotine adenine dinucleotide tetrazolim reductase(NADH-TRase) to demonstrate the types, proportion, and density of masseter muscle fibers. AMPKα1 and p-AMPK(Thr172) levels in bilateral masseter muscles were detected by Western blot. In the 2-week group, the percentage of dark fibers augmented in the ipsilateral side, whereas the percentage of intermediary fibers in the contralateral side was increased accompanied by a decrease of light fibers, compared with the control group (P<0.05). The percentage of dark fibers was increased in the bilateral sides, whereas the percentage of dark fiber in the ipsilateral sides surpassed that of the contralateral sides in the 4, 6, and 8-week groups. The percentage of intermediary fibers was decreased in the bilateral sides in the 6 and 8-week groups (P<0.05). The percentage of light fibers was reduced in the ipsilateral sides in the 8-week group, whereas no alteration was observed in contralateral sides (P>0.05). In the ipsilateral sides, p-AMPK (Thr172)/AMPKα1 levels were increased in the 2 and 4-week groups (P<0.05), whereas no change was observed in the contralateral sides in either group (P>0.05). Unilateral chewing increases the oxidative metabolic ability in bilateral masseter muscle fibers especially in the non-working side accompanied with change of muscle fiber types. The improvement of aerobic metabolism ability is related to the AMPK signal pathway.
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  20. Muscarinic Acetylcholine Receptors Potentiate 5'-Adenosine Monophosphate-Activated Protein Kinase Stimulation and Glucose Uptake Triggered by Thapsigargin-Induced Store-Operated Ca(2+) Entry in Human Neuroblastoma Cells.

    PubMed

    Olianas, Maria C; Dedoni, Simona; Onali, Pierluigi

    2017-10-09

    The 5'-adenosine monophosphate-activated protein kinase (AMPK) is a key regulator of the cellular energy metabolism and may induce either cell survival or death. We previously reported that in SH-SY5Y human neuroblastoma cells stimulation of muscarinic acetylcholine receptors (mAChRs) activate AMPK by triggering store-operated Ca(2+) entry (SOCE). However, whether mAChRs may control AMPK activity by regulating additional mechanisms beyond SOCE remains to be investigated. In the present study we examined the effects of mAChRs on AMPK when SOCE was induced by the sarco-endoplasmic reticulum Ca(2+)-ATPase inhibitor thapsigargin. We found that in SH-SY5Y cells depleted of Ca(2+) by thapsigargin, the re-addition Ca(2+) to the medium stimulated AMPK phosphorylation at Thr172, which is required for full kinase activity. This response occurred through SOCE, as it was blocked by either the SOCE modulator 2-aminoethoxydiphephenyl borate, knockdown of the SOCE molecular component STIM1, or inhibition of Ca(2+)/calmodulin (CaM)-dependent protein kinase kinase β (CaMKKβ). In thapsigargin-pretreated cells, stimulation of pharmacologically defined M3 mAChRs potentiated SOCE-induced AMPK activation. This potentiation did not involve an increased Ca(2+) influx, but was associated with CaM mobilization from membrane to cytosol, increased CaM/CaMKKβ interaction, and enhanced CaMKK stimulation by thapsigargin-induced SOCE. In thapsigargin-pretreated cells Ca(2+) re-addition stimulated glucose uptake and increased the membrane expression of the glucose transporter GLUT1. Both responses were significantly potentiated by mAChRs. These data indicate that in human neuroblastoma cells mAChRs up-regulate AMPK and the downstream glucose uptake by triggering not only SOCE but also CaM translocation and enhanced formation of active CaM/CaMKKβ complexes.

  1. Post-meal responses of elongation factor 2 (eEF2) and adenosine monophosphate-activated protein kinase (AMPK) to leucine and carbohydrate supplements for regulating protein synthesis duration and energy homeostasis in rat skeletal muscle.

    PubMed

    Wilson, Gabriel J; Moulton, Christopher J; Garlick, Peter J; Anthony, Tracy G; Layman, Donald K

    2012-11-13

    Previous research demonstrates that the anabolic response of muscle protein synthesis (MPS) to a meal is regulated at the level of translation initiation with signals derived from leucine (Leu) and insulin to activate mTORC1 signaling. Recent evidence suggests that the duration of the meal response is limited by energy status of the cell and inhibition of translation elongation factor 2 (eEF2). This study evaluates the potential to extend the anabolic meal response with post-meal supplements of Leu or carbohydrates. Adult (~256 g) male Sprague-Dawley rats were food deprived for 12 h, then either euthanized before a standard meal (time 0) or at 90 or 180 min post-meal. At 135 min post-meal, rats received one of five oral supplements: 270 mg leucine (Leu270), 80:40:40 mg leucine, isoleucine, and valine (Leu80), 2.63 g carbohydrates (CHO2.6), 1 g carbohydrates (CHO1.0), or water (Sham control). Following the standard meal, MPS increased at 90 min then declined to pre-meal baseline at 180 min. Rats administered Leu270, Leu80, CHO2.6, or CHO1.0 maintained elevated rates of MPS at 180 min, while Sham controls declined from peak values. Leu80 and CHO1.0 treatments maintained MPS, but with values intermediate between Sham controls and Leu270 and CHO2.6 supplements. Consistent with MPS findings, the supplements maintained elongation activity and cellular energy status by preventing increases in AMP/ATP and phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), acetyl-CoA carboxylase ACC and eEF2. The impact of the supplements on MPS and cellular energy status was in proportion to the energy content within the individual treatments (i.e., Leu270 > Leu80; CHO2.6 > CHO1.0), but the Leu supplements produced a disproportionate anabolic stimulation of MPS, eEF2 and energy status with significantly lower energy content. In summary, the incongruity between MPS and translation initiation at 180 min reflects a block in translation elongation due to reduced

  2. Adenosine Monophosphate-Activated Protein Kinase Abates Hyperglycaemia-Induced Neuronal Injury in Experimental Models of Diabetic Neuropathy: Effects on Mitochondrial Biogenesis, Autophagy and Neuroinflammation.

    PubMed

    Yerra, Veera Ganesh; Kumar, Ashutosh

    2017-04-01

    Impaired adenosine monophosphate kinase (AMPK) signalling under hyperglycaemic conditions is known to cause mitochondrial dysfunction in diabetic sensory neurons. Facilitation of AMPK signalling is previously reported to ameliorate inflammation and induce autophagic response in various complications related to diabetes. The present study assesses the role of AMPK activation on mitochondrial biogenesis, autophagy and neuroinflammation in experimental diabetic neuropathy (DN) using an AMPK activator (A769662). A769662 (15 and 30 mg/kg, i.p) was administered to Sprague-Dawley rats (250-270 g) for 2 weeks after 6 weeks of streptozotocin (STZ) injection (55 mg/kg, i.p.). Behavioural parameters (mechanical/thermal hyperalgesia) and functional characteristics (motor/sensory nerve conduction velocities (MNCV and SNCV) and sciatic nerve blood flow (NBF)) were assessed. For in vitro studies, Neuro2a (N2A) cells were incubated with 25 mM glucose to simulate high glucose condition and then studied for mitochondrial dysfunction and protein expression changes. STZ administration resulted in significant hyperglycaemia (>250 mg/dl) in rats. A769662 treatment significantly improved mechanical/thermal hyperalgesia threshold and enhanced MNCV, SNCV and NBF in diabetic animals. A769662 exposure normalised the mitochondrial superoxide production, membrane depolarisation and markedly increased neurite outgrowth of N2A cells. Further, AMPK activation also abolished the NF-κB-mediated neuroinflammation. A769662 treatment increased Thr-172 phosphorylation of AMPK results in stimulated PGC-1α-directed mitochondrial biogenesis and autophagy induction. Our study supports that compromised AMPK signalling in hyperglycaemic conditions causes defective mitochondrial biogenesis ultimately leading to neuronal dysfunction and associated deficits in DN and activation of AMPK can be developed as an attractive therapeutic strategy for the management of DN.

  3. The fruit of Acanthopanax senticosus (Rupr. et Maxim.) Harms improves insulin resistance and hepatic lipid accumulation by modulation of liver adenosine monophosphate-activated protein kinase activity and lipogenic gene expression in high-fat diet-fed obese mice.

    PubMed

    Saito, Tetsuo; Nishida, Miyako; Saito, Masafumi; Tanabe, Akari; Eitsuka, Takahiro; Yuan, Shi-Hua; Ikekawa, Nobuo; Nishida, Hiroshi

    2016-10-01

    Obesity-associated insulin resistance is a major risk factor for most metabolic diseases, including dyslipidemia and type 2 diabetes. Acanthopanax senticosus (Rupr. et Maxim.) Harms (Goka) root has been used in traditional Chinese medicine for treatment of diabetes and other conditions; however, little is known about the effects of Goka fruit (GF). Goka fruit is rich in anthocyanin, which has beneficial effects on obesity and insulin resistance via activation of adenosine monophosphate-activated protein kinase (AMPK). We hypothesized that GF can improve obesity-associated insulin resistance. The aim of the present study was to investigate whether GF improves insulin resistance in high-fat diet (HFD)-induced obese mice. High-fat diet mice treated with GF (500 and 1000 mg/kg) for 12 weeks showed an improved glucose tolerance and insulin sensitivity, as well as reduced plasma insulin and liver lipid accumulation. Moreover, GF administration to HFD mice resulted in down-regulation of fatty acid synthase expression and up-regulation of cholesterol 7-alpha-hydroxylase expression in the liver. Notably, AMPK phosphorylation in the liver increased after GF administration. In summary, GF supplementation improved obesity-associated insulin resistance and hepatic lipid accumulation through modulation of AMPK activity and lipid metabolism-associated gene expression.

  4. Genistein promotes insulin action through adenosine monophosphate-activated protein kinase activation and p70 ribosomal protein S6 kinase 1 inhibition in the skeletal muscle of mice fed a high energy diet.

    PubMed

    Arunkumar, Elumalai; Anuradha, Carani Venkatraman

    2012-08-01

    Genistein (GEN), a soy isoflavone, exerts insulin-sensitizing actions in animals; however, the underlying mechanisms have not been determined. Because GEN is a known activator of adenosine monophosphate-activated protein kinase (AMPK), we hypothesize that GEN activates insulin signaling through AMPK activation. To test this hypothesis, a high fat-high fructose diet (HFFD)-fed mice model of insulin resistance was administered GEN, and the insulin signaling pathway proteins in the skeletal muscle were examined. Hyperglycemia and hyperinsulinemia observed in HFFD-fed mice were significantly lowered by GEN. GEN increased insulin-stimulated tyrosine phosphorylation of insulin receptor-β and insulin receptor substrate (IRS) 1 but down-regulated IRS-1 serine phosphorylation in the skeletal muscle of HFFD-fed mice. Furthermore, GEN treatment improved muscle IRS-1-associated phospatidylinositol-3 kinase expression, phosphorylation of Akt at Ser(473), and translocation of glucose transporter subtype 4. Phosphorylation of AMPK at Thr(172) and acetyl coenzyme A carboxylase (ACC) at Ser(79) was augmented, whereas phosphorylation of p70 ribosomal protein S6 kinase 1 at Thr(389) was significantly decreased after GEN treatment in the skeletal muscle of HFFD-fed mice. These results suggest that GEN might improve insulin action in the skeletal muscle by targeting AMPK.

  5. Advanced glycation end-products impair Na⁺/K⁺-ATPase activity in diabetic cardiomyopathy: role of the adenosine monophosphate-activated protein kinase/sirtuin 1 pathway.

    PubMed

    Yuan, Qiong; Zhou, Qian-Yi; Liu, Du; Yu, Lun; Zhan, Lin; Li, Xiao-Jing; Peng, Hong-Yan; Zhang, Xiu-Ling; Yuan, Xin-Chu

    2014-02-01

    Decreased Na(+) /K(+) -ATPase activity, and both sirtuin 1 (SIRT1) and adenosine monophosphate-activated protein kinase (AMPK) have been reported to be involved in the development of diabetic cardiomyopathy (DCM). The present study aimed to investigate the advanced glycation end-products (AGE) that impair Na(+) /K(+) -ATPase stability by regulating the AMPK/SIRT1 pathway during progression of DCM. To study type 1 diabetic mellitus (T1DM), a disease model in rats was established by a single intraperitoneal injection of streptozotocin (STZ; 65 mg/kg), and neonatal rat cardiomyocytes were also cultured. Heart function was detected by Doppler, and SIRT1 and AMPK protein expression were detected by immunohistochemistry and western blotting. Na(+) /K(+) -ATPase activity was also monitored. Using in vivo rat models of DCM, we showed that Na(+) /K(+) -ATPase activity decreased when both AMPK and SIRT1 expression were downregulated. In vitro, AGE impaired Na(+) /K(+) -ATPase activity and decreased the AMPK and SIRT1 expression. Sirtuin 1 overexpression increased Na(+) /K(+) -ATPase activity. 5-aminoimidazole-4-carboxamide-3-ribonucleoside (AICAR) upregulated SIRT1 expression and increased Na(+) /K(+) -ATPase activity, which could be partially abolished by splitomicin. Our results suggest that the dysfunction of DCM is related to AGE-induced Na(+) /K(+) -ATPase activity impairment through a mechanism involving the AMPK/SIRT1 pathway.

  6. Diabetic complications within the context of aging: Nicotinamide adenine dinucleotide redox, insulin C-peptide, sirtuin 1-liver kinase B1-adenosine monophosphate-activated protein kinase positive feedback and forkhead box O3.

    PubMed

    Ido, Yasuo

    2016-07-01

    Recent research in nutritional control of aging suggests that cytosolic increases in the reduced form of nicotinamide adenine dinucleotide and decreasing nicotinamide adenine dinucleotide metabolism plays a central role in controlling the longevity gene products sirtuin 1 (SIRT1), adenosine monophosphate-activated protein kinase (AMPK) and forkhead box O3 (FOXO3). High nutrition conditions, such as the diabetic milieu, increase the ratio of reduced to oxidized forms of cytosolic nicotinamide adenine dinucleotide through cascades including the polyol pathway. This redox change is associated with insulin resistance and the development of diabetic complications, and might be counteracted by insulin C-peptide. My research and others' suggest that the SIRT1-liver kinase B1-AMPK cascade creates positive feedback through nicotinamide adenine dinucleotide synthesis to help cells cope with metabolic stress. SIRT1 and AMPK can upregulate liver kinase B1 and FOXO3, key factors that help residential stem cells cope with oxidative stress. FOXO3 directly changes epigenetics around transcription start sites, maintaining the health of stem cells. 'Diabetic memory' is likely a result of epigenetic changes caused by high nutritional conditions, which disturb the quiescent state of residential stem cells and impair tissue repair. This could be prevented by restoring SIRT1-AMPK positive feedback through activating FOXO3.

  7. Expression of adenosine 5'-monophosphate-Activated protein kinase (AMPK) in ovine testis (Ovis aries): In vivo regulation by nutritional state.

    PubMed

    Taibi, N; Dupont, J; Bouguermouh, Z; Froment, P; Ramé, C; Anane, A; Amirat, Z; Khammar, F

    2017-03-01

    In the present study, we identified AMPK and investigated its potential role in steroidogenesis in vivo in the ovine testis in response to variation in nutritional status (fed control vs. restricted). We performed immunoblotting to show that both active and non-active forms of AMPK exist in ovine testis and liver. In testis, we confirmed these results by immunohistochemistry. We found a correlation between ATP (Adenosine-Triphosphate) levels and the expression of AMPK in liver. Also, low and high caloric diets induce isoform-dependent AMPK expression, with an increase in α2, ß1ß2 and γ1 activity levels. Although the restricted group exhibited an increase in lipid balance, only the triglyceride and HC-VLDL (Cholesterol-Very low density lipoprotein) fractions showed significant differences between groups, suggesting an adaptive mechanism. Moreover, the relatively low rate of non-esterified fatty acid released into the circulation implies re-esterification to compensate for the physiological need. In the fed control group, AMPK activates the production of testosterone in Leydig cells; this is, in turn, associated with an increase in the expression of 3ß-HSD (3 beta hydroxy steroid deshydrogenase), p450scc (Cholesterol side-chain cleavage enzyme) and StAR (Steroidogenic acute regulatory protein) proteins induced by decreased MAPK ERK½ (Extracellular signal-regulated kinase -Mitogen-activated protein kinase) phosphorylation. In contrast, in the restricted group, testosterone secretion was reduced but intracellular cholesterol concentration was not. Furthermore, the combination of high levels of lipoproteins and emergence of the p38 MAP kinase pathway suggest the involvement of pro-inflammatory cytokines, as confirmed by transcriptional repression of the StAR protein. Taken together, these results suggest that AMPK expression is tissue dependent. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Berberine treatment prevents cardiac dysfunction and remodeling through activation of 5'-adenosine monophosphate-activated protein kinase in type 2 diabetic rats and in palmitate-induced hypertrophic H9c2 cells.

    PubMed

    Chang, Wenguang; Zhang, Ming; Meng, Zhaojie; Yu, Yang; Yao, Fan; Hatch, Grant M; Chen, Li

    2015-12-15

    Diabetic cardiomyopathy is the major cause of death in type 2 diabetic patients. Berberine is an isoquinoline alkaloid extract from traditional chinese herbs and its hypoglycemic and hypolipidemic effects make it a promising drug for treatment of type 2 diabetes. We examined if berberine improved cardiac function and attenuated cardiac hypertrophy and fibrosis in high fat diet and streptozotocin induced-type 2 diabetic rats in vivo and reduced expression of hypertrophy markers in palmitate-induced hypertrophic H9c2 cells in vitro. Treatment of diabetic animals with berberine partially improved cardiac function and restored fasting blood insulin, fasting blood glucose, total cholesterol, and triglyceride levels to that of control. In addition, berberine treatment of diabetic animals increased cardiac 5'-adenosine monophosphate-activated protein kinase (AMPK) and protein kinase B (AKT) activation and reduced glycogen synthase kinase 3 beta (GSK3β) activation compared to control. Palmitate incubation of H9c2 cells resulted in cellular hypertrophy and decreased expression of alpha-myosin heavy chain (α-MHC) and increased expression of beta-myosin heavy chain (β-MHC) compared to controls. Berberine treatment of palmitate-incubated H9c2 cells reduced hypertrophy, increased α-MHC expression and decreased β-MHC expression. In addition, berberine treatment of palmitate-incubated H9c2 cells increased AMPK and AKT activation and reduced GSK3β activation. The presence of the AMPK inhibitor Compound C attenuated the effects of berberine. The results strongly indicate that berberine treatment may be protective against the development of diabetic cardiomyopathy.

  9. Adenosine monophosphate activated protein kinase (AMPK), a mediator of estradiol-induced apoptosis in long-term estrogen deprived breast cancer cells.

    PubMed

    Chen, Haiyan; Wang, Ji-Ping; Santen, Richard J; Yue, Wei

    2015-06-01

    Estrogens stimulate growth of hormone-dependent breast cancer but paradoxically induce tumor regress under certain circumstances. We have shown that long-term estrogen deprivation (LTED) enhances the sensitivity of hormone dependent breast cancer cells to estradiol (E2) so that physiological concentrations of estradiol induce apoptosis in these cells. E2-induced apoptosis involve both intrinsic and extrinsic pathways but precise mechanisms remain unclear. We found that exposure of LTED MCF-7 cells to E2 activated AMP activated protein kinase (AMPK). In contrast, E2 inhibited AMPK activation in wild type MCF-7 cells where E2 prevents apoptosis. As a result of AMPK activation, the transcriptional activity of FoxO3, a downstream factor of AMPK, was up-regulated in E2 treatment of LTED. Increased activity of FoxO3 was demonstrated by up-regulation of three FoxO3 target genes, Bim, Fas ligand (FasL), and Gadd45α. Among them, Bim and FasL mediate intrinsic and extrinsic apoptosis respectively and Gadd45α causes cell cycle arrest at the G2/M phase. To further confirm the role of AMPK in apoptosis, we used AMPK activator AICAR in wild type MCF-7 cells and examined apoptosis, proliferation and expression of Bim, FasL, and Gadd45α. The effects of AICAR on these parameters recapitulated those observed in E2-treated LTED cells. Activation of AMPK by AICAR also increased expression of Bax in MCF-7 cells and its localization to mitochondria, which is a required process for apoptosis. These results reveal that AMPK is an important factor mediating E2-induced apoptosis in LTED cells, which is implicative of therapeutic potential for relapsing breast cancer after hormone therapy.

  10. Discovery and Preclinical Characterization of 6-Chloro-5-[4-(1-hydroxycyclobutyl)phenyl]-1H-indole-3-carboxylic Acid (PF-06409577), a Direct Activator of Adenosine Monophosphate-activated Protein Kinase (AMPK), for the Potential Treatment of Diabetic Nephropathy.

    PubMed

    Cameron, Kimberly O; Kung, Daniel W; Kalgutkar, Amit S; Kurumbail, Ravi G; Miller, Russell; Salatto, Christopher T; Ward, Jessica; Withka, Jane M; Bhattacharya, Samit K; Boehm, Markus; Borzilleri, Kris A; Brown, Janice A; Calabrese, Matthew; Caspers, Nicole L; Cokorinos, Emily; Conn, Edward L; Dowling, Matthew S; Edmonds, David J; Eng, Heather; Fernando, Dilinie P; Frisbie, Richard; Hepworth, David; Landro, James; Mao, Yuxia; Rajamohan, Francis; Reyes, Allan R; Rose, Colin R; Ryder, Tim; Shavnya, Andre; Smith, Aaron C; Tu, Meihua; Wolford, Angela C; Xiao, Jun

    2016-09-08

    Adenosine monophosphate-activated protein kinase (AMPK) is a protein kinase involved in maintaining energy homeostasis within cells. On the basis of human genetic association data, AMPK activators were pursued for the treatment of diabetic nephropathy. Identification of an indazole amide high throughput screening (HTS) hit followed by truncation to its minimal pharmacophore provided an indazole acid lead compound. Optimization of the core and aryl appendage improved oral absorption and culminated in the identification of indole acid, PF-06409577 (7). Compound 7 was advanced to first-in-human trials for the treatment of diabetic nephropathy.

  11. Hindbrain A2 noradrenergic neuron adenosine 5'-monophosphate-activated protein kinase activation, upstream kinase/phosphorylase protein expression, and receptivity to hormone and fuel reporters of short-term food deprivation are regulated by estradiol.

    PubMed

    Briski, Karen P; Alenazi, Fahaad S H; Shakya, Manita; Sylvester, Paul W

    2016-09-12

    Estradiol (E) mitigates acute and postacute adverse effects of 12 hr-food deprivation (FD) on energy balance. Hindbrain 5'-monophosphate-activated protein kinase (AMPK) regulates hyperphagic and hypothalamic metabolic neuropeptide and norepinephrine responses to FD in an E-dependent manner. Energy-state information from AMPK-expressing hindbrain A2 noradrenergic neurons shapes neural responses to metabolic imbalance. Here we investigate the hypothesis that FD causes divergent changes in A2 AMPK activity in E- vs. oil (O)-implanted ovariectomized female rats, alongside dissimilar adjustments in circulating metabolic fuel (glucose, free fatty acids [FFA]) and energy deficit-sensitive hormone (corticosterone, glucagon, leptin) levels. FD decreased blood glucose in oil (O)- but not E-implanted ovariectomized female rats and elevated and reduced glucagon levels in O and E, respectively. FD decreased circulating leptin in O and E, but increased corticosterone and FFA concentrations in E only. Western blot analysis of laser-microdissected A2 neurons showed that glucocorticoid receptor type II and very-long-chain acyl-CoA synthetase 3 protein profiles were amplified in FD/E vs. FD/O. A2 total AMPK protein was elevated without change in activity in FD/O, whereas FD/E exhibited increased AMPK activation along with decreased upstream phosphatase expression. The catecholamine biosynthetic enzyme dopamine-β-hydroxylase (DβH) was increased in FD/O but not FD/E A2 cells. The data show discordance between A2 AMPK activation and glycemic responses to FD; sensor activity was refractory to glucose decrements in FD/O but augmented in FD/E despite stabilized glucose and elevated FFA levels. E-dependent amplification of AMPK activity may reflect adaptive conversion to fatty acid oxidation and/or glucocorticoid stimulation. FD augmentation of A2 DβH protein profiles in FD/O but not FD/E animals suggests that FD may correspondingly regulate NE synthesis vs. metabolism/release in the

  12. Adenosine conjugated lipidic nanoparticles for enhanced tumor targeting.

    PubMed

    Swami, Rajan; Singh, Indu; Jeengar, Manish Kumar; Naidu, V G M; Khan, Wahid; Sistla, Ramakrishna

    2015-01-01

    Delivering chemotherapeutics by nanoparticles into tumor is impeded majorly by two factors: nonspecific targeting and inefficient penetration. Targeted delivery of anti-cancer agents solely to tumor cells introduces a smart strategy because it enhances the therapeutic index compared with untargeted drugs. The present study was performed to investigate the efficiency of adenosine (ADN) to target solid lipid nanoparticles (SLN) to over expressing adenosine receptor cell lines such as human breast cancer and prostate cancer (MCF-7 and DU-145 cells), respectively. SLN were prepared by emulsification and solvent evaporation process using docetaxel (DTX) as drug and were characterized by various techniques like dynamic light scattering, differential scanning calorimeter and transmission electron microscopy. DTX loaded SLNs were surface modified with ADN, an adenosine receptors ligand using carbodiimide coupling. Conjugation was confirmed using infrared spectroscopy and quantified using phenol-sulfuric acid method. Conjugated SLN were shown to have sustained drug release as compared to unconjugated nanoparticles and drug suspension. Compared with free DTX and unconjugated SLN, ADN conjugated SLN showed significantly higher cytotoxicity of loaded DTX, as evidenced by in vitro cell experiments. The IC50 was 0.41 μg/ml for native DTX, 0.30 μg/ml for unconjugated SLN formulation, and 0.09 μg/ml for ADN conjugated SLN formulation in MCF-7 cell lines. Whereas, in DU-145, there was 2 fold change in IC50 of ADN-SLN as compared to DTX. IC50 was found to be 0.44 μg/ml for free DTX, 0.39 μg/ml for unconjugated SLN and 0.22 μg/ml for ADN-SLN. Annexin assay and cell cycle analysis assay further substantiated the cell cytotoxicity. Fluorescent cell uptake and competitive ligand-receptor binding assay corroborated the receptor mediated endocytosis pathway indicated role of adenosine receptors in internalization of conjugated particles. Pharmacokinetic studies of lipidic

  13. Adenine arabinoside inhibition of adenovirus replication enhanced by an adenosine deaminase inhibitor.

    PubMed

    Wigand, R

    1979-01-01

    The inhibition of adenovirus multiplication by adenine arabinoside was determined by yield reduction in one-step multiplication cycle. Inhibition was greatly enhanced by an adenosine deaminase inhibitor (2-deoxycoformycin) in concentrations down to 10 ng/ml. Adenovirus types from four subgroups showed similar results. However, the enhancing effect of adenosine deaminase inhibitor was great in HeLa cells, moderate in human fibroblasts, and negligible in Vero cells. This difference could be explained by different concentrations of adenosine deaminase found in cell homogenates.

  14. The ABCD's of 5'-adenosine monophosphate-activated protein kinase and adrenoleukodystrophy.

    PubMed

    Weidling, Ian; Swerdlow, Russell H

    2016-07-01

    This Editorial highlights a study by Singh and coworkers in the current issue of Journal of Neurochemistry, in which the authors present additional evidence that AMPKα1 is reduced in X-linked adrenoleukodystrophy (X-ALD). They make a case for increasing AMPKα1 activity for therapeutic purposes in this disease, and indicate how this goal may be achieved. Read the highlighted article 'Metformin-induced mitochondrial function and ABCD2 up regulation in X-linked adrenoleukodystrophy involves AMP activated protein kinase' on page 86. © 2016 International Society for Neurochemistry.

  15. Adenosine enhances sweet taste through A2B receptors in the taste bud.

    PubMed

    Dando, Robin; Dvoryanchikov, Gennady; Pereira, Elizabeth; Chaudhari, Nirupa; Roper, Stephen D

    2012-01-04

    Mammalian taste buds use ATP as a neurotransmitter. Taste Receptor (type II) cells secrete ATP via gap junction hemichannels into the narrow extracellular spaces within a taste bud. This ATP excites primary sensory afferent fibers and also stimulates neighboring taste bud cells. Here we show that extracellular ATP is enzymatically degraded to adenosine within mouse vallate taste buds and that this nucleoside acts as an autocrine neuromodulator to selectively enhance sweet taste. In Receptor cells in a lingual slice preparation, Ca(2+) mobilization evoked by focally applied artificial sweeteners was significantly enhanced by adenosine (50 μM). Adenosine had no effect on bitter or umami taste responses, and the nucleoside did not affect Presynaptic (type III) taste cells. We also used biosensor cells to measure transmitter release from isolated taste buds. Adenosine (5 μM) enhanced ATP release evoked by sweet but not bitter taste stimuli. Using single-cell reverse transcriptase (RT)-PCR on isolated vallate taste cells, we show that many Receptor cells express the adenosine receptor, Adora2b, while Presynaptic (type III) and Glial-like (type I) cells seldom do. Furthermore, Adora2b receptors are significantly associated with expression of the sweet taste receptor subunit, Tas1r2. Adenosine is generated during taste stimulation mainly by the action of the ecto-5'-nucleotidase, NT5E, and to a lesser extent, prostatic acid phosphatase. Both these ecto-nucleotidases are expressed by Presynaptic cells, as shown by single-cell RT-PCR, enzyme histochemistry, and immunofluorescence. Our findings suggest that ATP released during taste reception is degraded to adenosine to exert positive modulation particularly on sweet taste.

  16. Adenosine enhances sweet taste through A2B receptors in the taste bud

    PubMed Central

    Dando, Robin; Dvoryanchikov, Gennady; Pereira, Elizabeth; Chaudhari, Nirupa; Roper, Stephen D.

    2012-01-01

    Mammalian taste buds use ATP as a neurotransmitter. Taste Receptor (Type II) cells secrete ATP via gap junction hemichannels into the narrow extracellular spaces within a taste bud. This ATP excites primary sensory afferent fibers and also stimulates neighboring taste bud cells. Here we show that extracellular ATP is enzymatically degraded to adenosine within mouse vallate taste buds and that this nucleoside acts as an autocrine neuromodulator to selectively enhance sweet taste. In Receptor cells in a lingual slice preparation, Ca2+ mobilization evoked by focally applied artificial sweeteners was significantly enhanced by adenosine (50 µM). Adenosine had no effect on bitter or umami taste responses, and the nucleoside did not affect Presynaptic (Type III) taste cells. We also used biosensor cells to measure transmitter release from isolated taste buds. Adenosine (5 µM) enhanced ATP release evoked by sweet but not bitter taste stimuli. Using single-cell RT-PCR on isolated vallate taste cells, we show that many Receptor cells express adenosine receptors, Adora2b, while Presynaptic (Type III) and Glial-like (Type I) cells seldom do. Furthermore, Adora2b receptors are significantly associated with expression of the sweet taste receptor subunit, Tas1r2. Adenosine is generated during taste stimulation mainly by the action of the ecto-5′-nucleotidase, NT5E, and to a lesser extent, prostatic acid phosphatase (ACPP). Both these ecto-nucleotidases are expressed by Presynaptic cells, as shown by single-cell RT-PCR, enzyme histochemistry and immunofluorescence. Our findings suggest that ATP released during taste reception is degraded to adenosine to exert positive modulation particularly on sweet taste. PMID:22219293

  17. Methyl xanthines enhance taste: evidence for modulation of taste by adenosine receptor.

    PubMed

    Schiffman, S S; Gill, J M; Diaz, C

    1985-02-01

    The methyl xanthines (MX), theophylline, caffeine, and theobromine, are potent antagonists of adenosine receptors. Adaptation of the human tongue to methyl xanthines at concentrations ranging from 10(-5) M to 10(-2) M was found to potentiate taste. The artificial sweetener acesulfam-K, which has a bitter component, was potentiated the most by MX, i.e., approximately 100%. This increase in perceived intensity for acesulfam-K occurred at 10(-5) M MX, a concentration known to inhibit adenosine receptors but below that required to inhibit phosphodiesterase. Increasing the concentration of MX as high as 10(-2) M did not increase the degree of enhancement appreciably. Taste enhancement was found for NaCl and quinine hydrochloride as well. When 10(-5) M adenosine was added to the MX, the potentiation was reversed. The human results were confirmed by animal studies in which single unit extracellular recordings were made from the nucleus of the solitary tract. These results suggest that the inhibitory A1 adenosine receptor plays an important local role in taste perception.

  18. Targeting the adenosine 2A receptor enhances chimeric antigen receptor T cell efficacy

    PubMed Central

    Beavis, Paul A.; Henderson, Melissa A.; Giuffrida, Lauren; Mills, Jane K.; Sek, Kevin; Cross, Ryan S.; Davenport, Alexander J.; John, Liza B.; Mardiana, Sherly; Slaney, Clare Y.; Johnstone, Ricky W.; Trapani, Joseph A.; Stagg, John; Loi, Sherene; Kats, Lev; Gyorki, David; Kershaw, Michael H.; Darcy, Phillip K.

    2017-01-01

    Chimeric antigen receptor (CAR) T cells have been highly successful in treating hematological malignancies, including acute and chronic lymphoblastic leukemia. However, treatment of solid tumors using CAR T cells has been largely unsuccessful to date, partly because of tumor-induced immunosuppressive mechanisms, including adenosine production. Previous studies have shown that adenosine generated by tumor cells potently inhibits endogenous antitumor T cell responses through activation of adenosine 2A receptors (A2ARs). Herein, we have observed that CAR activation resulted in increased A2AR expression and suppression of both murine and human CAR T cells. This was reversible using either A2AR antagonists or genetic targeting of A2AR using shRNA. In 2 syngeneic HER2+ self-antigen tumor models, we found that either genetic or pharmacological targeting of the A2AR profoundly increased CAR T cell efficacy, particularly when combined with PD-1 blockade. Mechanistically, this was associated with increased cytokine production of CD8+ CAR T cells and increased activation of both CD8+ and CD4+ CAR T cells. Given the known clinical relevance of the CD73/adenosine pathway in several solid tumor types, and the initiation of phase I trials for A2AR antagonists in oncology, this approach has high translational potential to enhance CAR T cell efficacy in several cancer types. PMID:28165340

  19. The Second Extracellular Loop of the Adenosine A1 Receptor Mediates Activity of Allosteric Enhancers

    PubMed Central

    Kennedy, Dylan P.; McRobb, Fiona M.; Leonhardt, Susan A.; Purdy, Michael; Figler, Heidi; Marshall, Melissa A.; Chordia, Mahendra; Figler, Robert; Linden, Joel

    2014-01-01

    Allosteric enhancers of the adenosine A1 receptor amplify signaling by orthosteric agonists. Allosteric enhancers are appealing drug candidates because their activity requires that the orthosteric site be occupied by an agonist, thereby conferring specificity to stressed or injured tissues that produce adenosine. To explore the mechanism of allosteric enhancer activity, we examined their action on several A1 receptor constructs, including (1) species variants, (2) species chimeras, (3) alanine scanning mutants, and (4) site-specific mutants. These findings were combined with homology modeling of the A1 receptor and in silico screening of an allosteric enhancer library. The binding modes of known docked allosteric enhancers correlated with the known structure-activity relationship, suggesting that these allosteric enhancers bind to a pocket formed by the second extracellular loop, flanked by residues S150 and M162. We propose a model in which this vestibule controls the entry and efflux of agonists from the orthosteric site and agonist binding elicits a conformational change that enables allosteric enhancer binding. This model provides a mechanism for the observations that allosteric enhancers slow the dissociation of orthosteric agonists but not antagonists. PMID:24217444

  20. Enhanced Diffusion of Molecular Motors in the Presence of Adenosine Triphosphate and External Force

    NASA Astrophysics Data System (ADS)

    Shinagawa, Ryota; Sasaki, Kazuo

    2016-06-01

    The diffusion of a molecular motor in the presence of a constant external force is considered on the basis of a simple theoretical model. The motor is represented by a Brownian particle moving in a series of parabolic potentials placed periodically on a line, and the potential is switched stochastically from one parabola to another by a chemical reaction, which corresponds to the hydrolysis or synthesis of adenosine triphosphate (ATP) in motor proteins. It is found that the diffusion coefficient as a function of the force exhibits peaks. The mechanism of this diffusion enhancement and the possibility of observing it in F1-ATPase, a biological rotary motor, are discussed.

  1. Desensitization of adenosine receptor-mediated inhibition of lipolysis. The mechanism involves the development of enhanced cyclic adenosine monophosphate accumulation in tolerant adipocytes.

    PubMed Central

    Hoffman, B B; Chang, H; Dall'Aglio, E; Reaven, G M

    1986-01-01

    Adipocytes contain adenosine receptors, termed A1 receptors, which inhibit lipolysis by decreasing adenylate cyclase activity. The inhibition of lipolysis by adenosine agonists in vivo acutely suppresses the plasma concentrations of free fatty acids (FFA) and triglycerides. We have found that infusions of the adenosine receptor agonist phenylisopropyladenosine (PIA) initially decreases plasma FFA concentrations; however, with prolonged exposure (6 d), rats become very tolerant to the effects of the drug. Adipocytes isolated from epididymal fat pads from PIA-infused rats have altered lipolytic responses. When lipolysis is stimulated with a relatively high concentration of isoproterenol (10(-7) M), PIA does not inhibit lipolysis in adipocytes from the infused animals. However, PIA inhibits isoproterenol-stimulated cyclic AMP (cAMP) accumulation in adipocytes from the infused rats although with decreased sensitivity compared with controls. The explanation for the impaired antilipolytic effect appears to be due to the fact that isoproterenol-stimulated cAMP accumulation is markedly increased in cells from infused rats. Indeed, basal lipolysis and lipolysis stimulated with lower concentrations of isoproterenol (10(-9), 10(-8) M) are effectively inhibited by PIA. cAMP accumulation is greatly increased in adipocytes from infused rats when stimulated by isoproterenol, ACTH, and forskolin. The results have some striking analogies to changes induced in nerve cells by prolonged exposure to narcotics. These data suggest that tolerance to PIA develops in adipocytes as a consequence of enhanced cAMP accumulation. PMID:3013937

  2. Genetically controlled upregulation of adenosine A(1) receptor expression enhances the survival of primary cortical neurons.

    PubMed

    Serchov, Tsvetan; Atas, Hasan-Cem; Normann, Claus; van Calker, Dietrich; Biber, Knut

    2012-10-01

    Adenosine has a key endogenous neuroprotective role in the brain, predominantly mediated by the adenosine A(1) receptor (A(1)R). This has been mainly explored using pharmacological tools and/or receptor knockout mice strains. It has long been suggested that the neuroprotective effects of A(1)R are increased following receptor upregulation, thus attenuating neuronal damage in pathological conditions. We have previously shown that the neuroprotective and neuromodulatory actions of the cytokines IL-6 and oncostatin M are mediated by induction of neuronal A(1)R expression. In order to investigate the direct effects of A(1)R upregulation in neurons, we have generated a tetracycline-regulated expression system with a bidirectional promoter, directing the simultaneous expression of the mouse A(1)R and GFP/mCherry reporter genes. In a first step, we tested the efficacy of the system in transiently transfected human embryonic kidney 293 cells. In addition, we confirmed the functional integrity of the expressed A(1)R by whole-cell patch clamp recordings. We demonstrated that A(1)R-transfected primary neurons show enhanced survival against N-methyl-D-aspartate-induced excitotoxicity. Pretreatment with an A(1)R-selective agonist additionally strongly decreased neuronal cell death, while an A(1)R antagonist completely abolished the neuroprotective effects of A(1)R upregulation. The presented data provide for the first time direct evidence that the upregulation of A(1)R enhances neuronal survival.

  3. Targeting myeloid-derived suppressor cells using a novel adenosine monophosphate-activated protein kinase (AMPK) activator.

    PubMed

    Trikha, Prashant; Plews, Robert L; Stiff, Andrew; Gautam, Shalini; Hsu, Vincent; Abood, David; Wesolowski, Robert; Landi, Ian; Mo, Xiaokui; Phay, John; Chen, Ching-Shih; Byrd, John; Caligiuri, Michael; Tridandapani, Susheela; Carson, William

    2016-01-01

    Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of early myeloid cells that accumulate in the blood and tumors of patients with cancer. MDSC play a critical role during tumor evasion and promote immune suppression through variety of mechanisms, such as the generation of reactive oxygen and nitrogen species (ROS and RNS) and cytokines. AMPactivated protein kinase (AMPK) is an evolutionarily conserved serine/threonine kinase that regulates energy homeostasis and metabolic stress. However, the role of AMPK in the regulation of MDSC function remains largely unexplored. This study was designed to investigate whether treatment of MDSC with OSU-53, a PPAR-inactive derivative that stimulates AMPK kinase, can modulate MDSC function. Our results demonstrate that OSU-53 treatment increases the phosphorylation of AMPK, significantly reduces nitric oxide production, inhibits MDSC migration, and reduces the levels of IL-6 in murine MDSC cell line (MSC2 cells). OSU53 treatment mitigated the immune suppressive functions of murine MDSC, promoting T-cell proliferation. Although OSU-53 had a modest effect on tumor growth in mice inoculated with EMT-6 cells, importantly, administration of OSU53 significantly (p < 0.05) reduced the levels of MDSC in the spleens and tumors. Furthermore, mouse MDSC from EMT-6 tumor-bearing mice and human MDSC isolated from melanoma patients treated with OSU-53 showed a significant reduction in the expression of immune suppressive genes iNOS and arginase. In summary, these results demonstrate a novel role of AMPK in the regulation of MDSC functions and provide a rationale of combining OSU-53 with immune checkpoint inhibitors to augment their response in cancer patients.

  4. Digoxin and Adenosine Triphosphate Enhance the Functional Properties of Tissue-Engineered Cartilage

    PubMed Central

    Makris, Eleftherios A.; Huang, Brian J.; Hu, Jerry C.; Chen-Izu, Ye

    2015-01-01

    Toward developing engineered cartilage for the treatment of cartilage defects, achieving relevant functional properties before implantation remains a significant challenge. Various chemical and mechanical stimuli have been used to enhance the functional properties of engineered musculoskeletal tissues. Recently, Ca2+-modulating agents have been used to enhance matrix synthesis and biomechanical properties of engineered cartilage. The objective of this study was to determine whether other known Ca2+ modulators, digoxin and adenosine triphosphate (ATP), can be employed as novel stimuli to increase collagen synthesis and functional properties of engineered cartilage. Neocartilage constructs were formed by scaffold-free self-assembling of primary bovine articular chondrocytes. Digoxin, ATP, or both agents were added to the culture medium for 1 h/day on days 10–14. After 4 weeks of culture, neocartilage properties were assessed for gross morphology, biochemical composition, and biomechanical properties. Digoxin and ATP were found to increase neocartilage collagen content by 52–110% over untreated controls, while maintaining proteoglycan content near native tissue values. Furthermore, digoxin and ATP increased the tensile modulus by 280% and 180%, respectively, while the application of both agents increased the modulus by 380%. The trends in tensile properties were found to correlate with the amount of collagen cross-linking. Live Ca2+ imaging experiments revealed that both digoxin and ATP were able to increase Ca2+ oscillations in monolayer-cultured chondrocytes. This study provides a novel approach toward directing neocartilage maturation and enhancing its functional properties using novel Ca2+ modulators. PMID:25473799

  5. Digoxin and adenosine triphosphate enhance the functional properties of tissue-engineered cartilage.

    PubMed

    Makris, Eleftherios A; Huang, Brian J; Hu, Jerry C; Chen-Izu, Ye; Athanasiou, Kyriacos A

    2015-03-01

    Toward developing engineered cartilage for the treatment of cartilage defects, achieving relevant functional properties before implantation remains a significant challenge. Various chemical and mechanical stimuli have been used to enhance the functional properties of engineered musculoskeletal tissues. Recently, Ca(2+)-modulating agents have been used to enhance matrix synthesis and biomechanical properties of engineered cartilage. The objective of this study was to determine whether other known Ca(2+) modulators, digoxin and adenosine triphosphate (ATP), can be employed as novel stimuli to increase collagen synthesis and functional properties of engineered cartilage. Neocartilage constructs were formed by scaffold-free self-assembling of primary bovine articular chondrocytes. Digoxin, ATP, or both agents were added to the culture medium for 1 h/day on days 10-14. After 4 weeks of culture, neocartilage properties were assessed for gross morphology, biochemical composition, and biomechanical properties. Digoxin and ATP were found to increase neocartilage collagen content by 52-110% over untreated controls, while maintaining proteoglycan content near native tissue values. Furthermore, digoxin and ATP increased the tensile modulus by 280% and 180%, respectively, while the application of both agents increased the modulus by 380%. The trends in tensile properties were found to correlate with the amount of collagen cross-linking. Live Ca(2+) imaging experiments revealed that both digoxin and ATP were able to increase Ca(2+) oscillations in monolayer-cultured chondrocytes. This study provides a novel approach toward directing neocartilage maturation and enhancing its functional properties using novel Ca(2+) modulators.

  6. Adenosine preferentially suppresses serotonin2A receptor-enhanced excitatory postsynaptic currents in layer V neurons of the rat medial prefrontal cortex.

    PubMed

    Stutzmann, G E; Marek, G J; Aghajanian, G K

    2001-01-01

    Serotonin induces 'spontaneous' (non-electrically evoked) excitatory postsynaptic currents in layer V pyramidal neurons in the prefrontal cortex. This is likely due to a serotonin2A receptor-mediated focal release of glutamate onto apical dendrites. In addition, activation of the serotonin2A receptor selectively enhances late components of electrically evoked excitatory postsynaptic currents. In this study, using in vitro intracellular and whole-cell recording in rat brain slices, we examined the role of adenosine in modulating serotonin2A-enhanced 'spontaneous' and electrically evoked excitatory postsynaptic currents in layer V pyramidal neurons in the medial prefrontal cortex. Adenosine and N6-cyclopentyladenosine, an A1 adenosine agonist, markedly suppressed the serotonin2A-induced ('spontaneous') excitatory postsynaptic currents. However, adenosine had no effect on spontaneous miniature (tetrodotoxin-insensitive) postsynaptic potentials. Adenosine also blocked the late excitatory postsynaptic currents induced by the serotonin2A/2C agonist R(-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride. Surprisingly, in contrast to other regions, adenosine had a relatively small effect on electrically evoked fast excitatory postsynaptic currents. These findings represent a novel demonstration of adenosine's ability to preferentially modulate serotonin2A-mediated synaptic events in the medial prefrontal cortex. As the serotonin2A receptor is closely linked with the effects of atypical antipsychotics and hallucinogens, further understanding of the modulators of this receptor such as adenosine may provide useful therapeutic applications.

  7. Extracellular adenosine triphosphate and adenosine in cancer.

    PubMed

    Stagg, J; Smyth, M J

    2010-09-30

    Adenosine triphosphate (ATP) is actively released in the extracellular environment in response to tissue damage and cellular stress. Through the activation of P2X and P2Y receptors, extracellular ATP enhances tissue repair, promotes the recruitment of immune phagocytes and dendritic cells, and acts as a co-activator of NLR family, pyrin domain-containing 3 (NLRP3) inflammasomes. The conversion of extracellular ATP to adenosine, in contrast, essentially through the enzymatic activity of the ecto-nucleotidases CD39 and CD73, acts as a negative-feedback mechanism to prevent excessive immune responses. Here we review the effects of extracellular ATP and adenosine on tumorigenesis. First, we summarize the functions of extracellular ATP and adenosine in the context of tumor immunity. Second, we present an overview of the immunosuppressive and pro-angiogenic effects of extracellular adenosine. Third, we present experimental evidence that extracellular ATP and adenosine receptors are expressed by tumor cells and enhance tumor growth. Finally, we discuss recent studies, including our own work, which suggest that therapeutic approaches that promote ATP-mediated activation of inflammasomes, or inhibit the accumulation of tumor-derived extracellular adenosine, may constitute effective new means to induce anticancer activity.

  8. Graphene/Cu nanoparticle hybrids fabricated by chemical vapor deposition as surface-enhanced Raman scattering substrate for label-free detection of adenosine.

    PubMed

    Xu, Shicai; Man, Baoyuan; Jiang, Shouzhen; Wang, Jihua; Wei, Jie; Xu, Shida; Liu, Hanping; Gao, Shoubao; Liu, Huilan; Li, Zhenhua; Li, Hongsheng; Qiu, Hengwei

    2015-05-27

    We present a graphene/Cu nanoparticle hybrids (G/CuNPs) system as a surface-enhanced Raman scattering (SERS) substrate for adenosine detection. The Cu nanoparticles wrapped by a monolayer graphene shell were directly synthesized on flat quartz by chemical vapor deposition in a mixture of methane and hydrogen. The G/CuNPs showed an excellent SERS enhancement activity for adenosine. The minimum detected concentration of the adenosine in serum was demonstrated as low as 5 nM, and the calibration curve showed a good linear response from 5 to 500 nM. The capability of SERS detection of adenosine in real normal human urine samples based on G/CuNPs was also investigated and the characteristic peaks of adenosine were still recognizable. The reproducible and the ultrasensitive enhanced Raman signals could be due to the presence of an ultrathin graphene layer. The graphene shell was able to enrich and fix the adenosine molecules, which could also efficiently maintain chemical and optical stability of G/CuNPs. Based on the G/CuNPs system, the ultrasensitive SERS detection of adenosine in varied matrices was expected for the practical applications in medicine and biotechnology.

  9. Selected C8 two-chain linkers enhance the adenosine A1/A2A receptor affinity and selectivity of caffeine.

    PubMed

    van der Walt, M M; Terre'Blanche, G

    2017-01-05

    Recent research exploring C8 substitution on the caffeine core identified 8-(2-phenylethyl)-1,3,7-trimethylxanthine as a non-selective adenosine receptor antagonist. To elaborate further, we included various C8 two-chain-length linkers to enhance adenosine receptor affinity. The results indicated that the unsubstituted benzyloxy linker (1e A1Ki = 1.52 μM) displayed the highest affinity for the A1 adenosine receptor and the para-chloro-substituted phenoxymethyl (1d A2AKi = 1.33 μM) linker the best A2A adenosine receptor affinity. The position of the oxygen revealed that the phenoxymethyl linker favoured A1 adenosine receptor selectivity over the benzyloxy linker and, by introducing a para-chloro substituent, A2A adenosine receptor selectivity was obtained. Selected compounds (1c, 1e) behaved as A1 adenosine receptor antagonists in GTP shift assays and therefore represent selective and non-selective A1 and A2A adenosine receptor antagonists that may have potential for treating neurological disorders.

  10. Extracellular Adenosine Production by ecto-5'-Nucleotidase (CD73) Enhances Radiation-Induced Lung Fibrosis.

    PubMed

    Wirsdörfer, Florian; de Leve, Simone; Cappuccini, Federica; Eldh, Therese; Meyer, Alina V; Gau, Eva; Thompson, Linda F; Chen, Ning-Yuan; Karmouty-Quintana, Harry; Fischer, Ute; Kasper, Michael; Klein, Diana; Ritchey, Jerry W; Blackburn, Michael R; Westendorf, Astrid M; Stuschke, Martin; Jendrossek, Verena

    2016-05-15

    Radiation-induced pulmonary fibrosis is a severe side effect of thoracic irradiation, but its pathogenesis remains poorly understood and no effective treatment is available. In this study, we investigated the role of the extracellular adenosine as generated by the ecto-5'-nucleotidase CD73 in fibrosis development after thoracic irradiation. Exposure of wild-type C57BL/6 mice to a single dose (15 Gray) of whole thorax irradiation triggered a progressive increase in CD73 activity in the lung between 3 and 30 weeks postirradiation. In parallel, adenosine levels in bronchoalveolar lavage fluid (BALF) were increased by approximately 3-fold. Histologic evidence of lung fibrosis was observed by 25 weeks after irradiation. Conversely, CD73-deficient mice failed to accumulate adenosine in BALF and exhibited significantly less radiation-induced lung fibrosis (P < 0.010). Furthermore, treatment of wild-type mice with pegylated adenosine deaminase or CD73 antibodies also significantly reduced radiation-induced lung fibrosis. Taken together, our findings demonstrate that CD73 potentiates radiation-induced lung fibrosis, suggesting that existing pharmacologic strategies for modulating adenosine may be effective in limiting lung toxicities associated with the treatment of thoracic malignancies. Cancer Res; 76(10); 3045-56. ©2016 AACR. ©2016 American Association for Cancer Research.

  11. Hypocretin/orexin antagonism enhances sleep-related adenosine and GABA neurotransmission in rat basal forebrain.

    PubMed

    Vazquez-DeRose, Jacqueline; Schwartz, Michael D; Nguyen, Alexander T; Warrier, Deepti R; Gulati, Srishti; Mathew, Thomas K; Neylan, Thomas C; Kilduff, Thomas S

    2016-03-01

    Hypocretin/orexin (HCRT) neurons provide excitatory input to wake-promoting brain regions including the basal forebrain (BF). The dual HCRT receptor antagonist almorexant (ALM) decreases waking and increases sleep. We hypothesized that HCRT antagonists induce sleep, in part, through disfacilitation of BF neurons; consequently, ALM should have reduced efficacy in BF-lesioned (BFx) animals. To test this hypothesis, rats were given bilateral IgG-192-saporin injections, which predominantly targets cholinergic BF neurons. BFx and intact rats were then given oral ALM, the benzodiazepine agonist zolpidem (ZOL) or vehicle (VEH) at lights-out. ALM was less effective than ZOL at inducing sleep in BFx rats compared to controls. BF adenosine (ADO), γ-amino-butyric acid (GABA), and glutamate levels were then determined via microdialysis from intact, freely behaving rats following oral ALM, ZOL or VEH. ALM increased BF ADO and GABA levels during waking and mixed vigilance states, and preserved sleep-associated increases in GABA under low and high sleep pressure conditions. ALM infusion into the BF also enhanced cortical ADO release, demonstrating that HCRT input is critical for ADO signaling in the BF. In contrast, oral ZOL and BF-infused ZOL had no effect on ADO levels in either BF or cortex. ALM increased BF ADO (an endogenous sleep-promoting substance) and GABA (which is increased during normal sleep), and required an intact BF for maximal efficacy, whereas ZOL blocked sleep-associated BF GABA release, and required no functional contribution from the BF to induce sleep. ALM thus induces sleep by facilitating the neural mechanisms underlying the normal transition to sleep.

  12. Scaffold Repurposing of Nucleosides (Adenosine Receptor Agonists): Enhanced Activity at the Human Dopamine and Norepinephrine Sodium Symporters.

    PubMed

    Tosh, Dilip K; Janowsky, Aaron; Eshleman, Amy J; Warnick, Eugene; Gao, Zhan-Guo; Chen, Zhoumou; Gizewski, Elizabeth; Auchampach, John A; Salvemini, Daniela; Jacobson, Kenneth A

    2017-04-13

    We have repurposed (N)-methanocarba adenosine derivatives (A3 adenosine receptor (AR) agonists) to enhance radioligand binding allosterically at the human dopamine (DA) transporter (DAT) and inhibit DA uptake. We extended the structure-activity relationship of this series with small N(6)-alkyl substitution, 5'-esters, deaza modifications of adenine, and ribose restored in place of methanocarba. C2-(5-Halothien-2-yl)-ethynyl 5'-methyl 9 (MRS7292) and 5'-ethyl 10 (MRS7232) esters enhanced binding at DAT (EC50 ∼ 35 nM) and at the norepinephrine transporter (NET). 9 and 10 were selective for DAT compared to A3AR in the mouse but not in humans. At DAT, the binding of two structurally dissimilar radioligands was enhanced; NET binding of only one radioligand was enhanced; SERT radioligand binding was minimally affected. 10 was more potent than cocaine at inhibiting DA uptake (IC50 = 107 nM). Ribose analogues were weaker in DAT interaction than the corresponding bicyclics. Thus, we enhanced the neurotransmitter transporter activity of rigid nucleosides while reducing A3AR affinity.

  13. Selective Allosteric Enhancement of Agonist Binding and Function at Human A3 Adenosine Receptors by a Series of Imidazoquinoline Derivatives

    PubMed Central

    Gao, Zhan-Guo; Kim, Seong Gon; Soltysiak, Kelly A.; Melman, Neli; Ijzerman, Adriaan P.; Jacobson, Kenneth A.

    2014-01-01

    We have identified a series of 1H-imidazo-[4,5-c]quinolines as selective allosteric enhancers of human A3 adenosine receptors. Several of these compounds potentiated both the potency and maximal efficacy of agonist-induced responses and selectively decreased the dissociation of the agonist N6-(4-amino-3-[125I]iodobenzyl)-5′-N-methylcarboxamidoadenosine from human A3 adenosine receptors. There was no effect on the dissociation of the antagonist [3H]8-ethyl-4-methyl-2-phenyl-(8R)-4,5,7,8-tetrahydro-1H-imidazo[2.1-i]purin-5-one (PSB-11) from the A3 receptors, as well as [3H]N6-[(R)-phenylisopropy-l]adenosine from rat brain A1 receptors and [3H]2-[p-(2-carboxyethyl)phenyl-ethylamino]-5′-N-ethylcarboxamidoad-enosine from rat striatal A2A receptors, suggesting the selective enhancement of agonist binding at A3 receptors. The analogs were tested as antagonists of competitive binding at human A3 receptors, and Ki values ranging from 120 nM to 101 μM were observed; as for many allosteric modulators of G protein-coupled receptors, an orthosteric effect was also present. The most promising leads from the present set of analogs seem to be the 2-cyclopentyl-1H-imidazo[4,5-c]quinoline derivatives, of which the 4-phenylamino analog DU124183 had the most favorable degree of allosteric modulation versus receptor antagonism. The inhibition of forskolin-stimulated cyclic AMP accumulation in intact cells that express human A3 receptors was employed as a functional index of A3 receptor activation. The enhancer DU124183 caused a marked leftward shift of the concentration-response curve of the A3 receptor agonists in the presence of antagonist and, surprisingly, a potentiation of the maximum agonist efficacy by approximately 30%. Thus, we have identified a novel structural lead for developing allosteric enhancers of A3 adenosine receptors; such enhancers may be useful for treating brain ischemia and other hypoxic conditions. PMID:12065758

  14. Enhanced neurodegeneration after a high dose of methamphetamine in Adenosine A3 receptor Null mutant mice

    PubMed Central

    Shen, Hui; Luo, Yu; Yu, Seong-Jin; Wang, Yun

    2011-01-01

    Previous reports have indicated that adenosine A3 receptor (A3R) knockout mice are more sensitive to ischemic or hypoxic brain injury. The purpose of this study was to examine if suppression of A3R expression is associated with increase in sensitivity to injury induced by a high dose of methamphetamine (Meth). Adult male A3R null mutant (−/−) mice and their controls (+/+) were injected with 4 doses (2 hours apart) of Meth (10 mg/kg) or saline. Animals were placed in a behavioral activity chamber, equipped with food and water, for 52 hours starting from one day after injections. The first 4 hours were used for studying exploratory behaviors and the next 48 hours were used to measure locomotor activity. High doses of Meth equally reduced the 4-hour exploratory behavior in −/− and +/+ mice. Meth suppressed locomotor activity between 4 and 52 hours in both groups, with a greater reduction being found in the −/− mice. Brain tissues were collected at 3 days after the Meth or saline injections. Meth treatment reduced striatal dopamine (DA) levels in both +/+ and −/− mice, examined by HPLC, with an increase in DOPAC/DA ratio being found only in −/− animals. Meth also significantly increased ionized calcium-binding adaptor molecule 1 (Iba-1) and cleaved caspase-3 level in striatum as well as Iba-1 and TNFα mRNA expression in nigra in −/−, compared to +/+, mice. Previous studies have shown that pharmacological suppression of VMAT2 by reserpine enhanced Meth toxicity by increasing cytosolic DA and inflammation. A significant reduction in striatal VMAT2 expression was found in −/− mice, compared to +/+ mice, suggesting that increase in sensitivity to Meth injury in −/− mice may be related to a reduction in VMAT2 expression in these mice. In conclusion, our data suggest that A3R −/− mice are more sensitive to high doses of Meth. PMID:21867746

  15. Direct or indirect stimulation of adenosine A2A receptors enhances bone regeneration as well as bone morphogenetic protein-2.

    PubMed

    Mediero, Aránzazu; Wilder, Tuere; Perez-Aso, Miguel; Cronstein, Bruce N

    2015-04-01

    Promoting bone regeneration and repair of bone defects is a need that has not been well met to date. We have previously found that adenosine, acting via A2A receptors (A2AR) promotes wound healing and inhibits inflammatory osteolysis and hypothesized that A2AR might be a novel target to promote bone regeneration. Therefore, we determined whether direct A2AR stimulation or increasing endogenous adenosine concentrations via purine transport blockade with dipyridamole regulates bone formation. We determined whether coverage of a 3 mm trephine defect in a mouse skull with a collagen scaffold soaked in saline, bone morphogenetic protein-2 (BMP-2; 200 ng), 1 μM CGS21680 (A2AR agonist, EC50 = 160 nM), or 1 μM dipyridamole (EC50 = 32 nM) promoted bone regeneration. Microcomputed tomography examination demonstrated that CGS21680 and dipyridamole markedly enhanced bone regeneration as well as BMP-2 8 wk after surgery (60 ± 2%, 79 ± 2%, and 75 ± 1% bone regeneration, respectively, vs. 32 ± 2% in control, P < 0.001). Blockade by a selective A2AR antagonist (ZM241385, 1 μM) or deletion of A2AR abrogated the effect of CGS21680 and dipyridamole on bone regeneration. Both CGS21680 and dipyridamole treatment increased alkaline phosphatase-positive osteoblasts and diminished tartrate resistance acid phosphatase-positive osteoclasts in the defects. In vivo imaging with a fluorescent dye for new bone formation revealed a strong fluorescent signal in treated animals that was equivalent to BMP-2. In conclusion, stimulation of A2AR by specific agonists or by increasing endogenous adenosine levels stimulates new bone formation as well as BMP-2 and represents a novel approach to stimulating bone regeneration. © FASEB.

  16. A2A Adenosine Receptor Antagonism Enhances Synaptic and Motor Effects of Cocaine via CB1 Cannabinoid Receptor Activation

    PubMed Central

    Tozzi, Alessandro; de Iure, Antonio; Marsili, Valentina; Romano, Rosaria; Tantucci, Michela; Di Filippo, Massimiliano; Costa, Cinzia; Napolitano, Francesco; Mercuri, Nicola Biagio; Borsini, Franco; Giampà, Carmen; Fusco, Francesca Romana; Picconi, Barbara; Usiello, Alessandro; Calabresi, Paolo

    2012-01-01

    Background Cocaine increases the level of endogenous dopamine (DA) in the striatum by blocking the DA transporter. Endogenous DA modulates glutamatergic inputs to striatal neurons and this modulation influences motor activity. Since D2 DA and A2A-adenosine receptors (A2A-Rs) have antagonistic effects on striatal neurons, drugs targeting adenosine receptors such as caffeine-like compounds, could enhance psychomotor stimulant effects of cocaine. In this study, we analyzed the electrophysiological effects of cocaine and A2A-Rs antagonists in striatal slices and the motor effects produced by this pharmacological modulation in rodents. Principal Findings Concomitant administration of cocaine and A2A-Rs antagonists reduced glutamatergic synaptic transmission in striatal spiny neurons while these drugs failed to produce this effect when given in isolation. This inhibitory effect was dependent on the activation of D2-like receptors and the release of endocannabinoids since it was prevented by L-sulpiride and reduced by a CB1 receptor antagonist. Combined application of cocaine and A2A-R antagonists also reduced the firing frequency of striatal cholinergic interneurons suggesting that changes in cholinergic tone might contribute to this synaptic modulation. Finally, A2A-Rs antagonists, in the presence of a sub-threshold dose of cocaine, enhanced locomotion and, in line with the electrophysiological experiments, this enhanced activity required activation of D2-like and CB1 receptors. Conclusions The present study provides a possible synaptic mechanism explaining how caffeine-like compounds could enhance psychomotor stimulant effects of cocaine. PMID:22715379

  17. A2B adenosine receptor blockade enhances macrophage-mediated bacterial phagocytosis and improves polymicrobial sepsis survival in mice.

    PubMed

    Belikoff, Bryan G; Hatfield, Stephen; Georgiev, Peter; Ohta, Akio; Lukashev, Dmitriy; Buras, Jon A; Remick, Daniel G; Sitkovsky, Michail

    2011-02-15

    Antimicrobial treatment strategies must improve to reduce the high mortality rates in septic patients. In noninfectious models of acute inflammation, activation of A2B adenosine receptors (A2BR) in extracellular adenosine-rich microenvironments causes immunosuppression. We examined A2BR in antibacterial responses in the cecal ligation and puncture (CLP) model of sepsis. Antagonism of A2BR significantly increased survival, enhanced bacterial phagocytosis, and decreased IL-6 and MIP-2 (a CXC chemokine) levels after CLP in outbred (ICR/CD-1) mice. During the CLP-induced septic response in A2BR knockout mice, hemodynamic parameters were improved compared with wild-type mice in addition to better survival and decreased plasma IL-6 levels. A2BR deficiency resulted in a dramatic 4-log reduction in peritoneal bacteria. The mechanism of these improvements was due to enhanced macrophage phagocytic activity without augmenting neutrophil phagocytosis of bacteria. Following ex vivo LPS stimulation, septic macrophages from A2BR knockout mice had increased IL-6 and TNF-α secretion compared with wild-type mice. A therapeutic intervention with A2BR blockade was studied by using a plasma biomarker to direct therapy to those mice predicted to die. Pharmacological blockade of A2BR even 32 h after the onset of sepsis increased survival by 65% in those mice predicted to die. Thus, even the late treatment with an A2BR antagonist significantly improved survival of mice (ICR/CD-1) that were otherwise determined to die according to plasma IL-6 levels. Our findings of enhanced bacterial clearance and host survival suggest that antagonism of A2BRs offers a therapeutic target to improve macrophage function in a late treatment protocol that improves sepsis survival.

  18. Cooperation of Adenosine with Macrophage Toll-4 Receptor Agonists Leads to Increased Glycolytic Flux through the Enhanced Expression of PFKFB3 Gene*

    PubMed Central

    Ruiz-García, Almudena; Monsalve, Eva; Novellasdemunt, Laura; Navarro-Sabaté, Àurea; Manzano, Anna; Rivero, Samuel; Castrillo, Antonio; Casado, Marta; Laborda, Jorge; Bartrons, Ramón; Díaz-Guerra, María José M.

    2011-01-01

    Macrophages activated through Toll receptor triggering increase the expression of the A2A and A2B adenosine receptors. In this study, we show that adenosine receptor activation enhances LPS-induced pfkfb3 expression, resulting in an increase of the key glycolytic allosteric regulator fructose 2,6-bisphosphate and the glycolytic flux. Using shRNA and differential expression of A2A and A2B receptors, we demonstrate that the A2A receptor mediates, in part, the induction of pfkfb3 by LPS, whereas the A2B receptor, with lower adenosine affinity, cooperates when high adenosine levels are present. pfkfb3 promoter sequence deletion analysis, site-directed mutagenesis, and inhibition by shRNAs demonstrated that HIF1α is a key transcription factor driving pfkfb3 expression following macrophage activation by LPS, whereas synergic induction of pfkfb3 expression observed with the A2 receptor agonists seems to depend on Sp1 activity. Furthermore, levels of phospho-AMP kinase also increase, arguing for increased PFKFB3 activity by phosphorylation in long term LPS-activated macrophages. Taken together, our results show that, in macrophages, endogenously generated adenosine cooperates with bacterial components to increase PFKFB3 isozyme activity, resulting in greater fructose 2,6-bisphosphate accumulation. This process enhances the glycolytic flux and favors ATP generation helping to develop and maintain the long term defensive and reparative functions of the macrophages. PMID:21464136

  19. GABAB and adenosine receptors mediate enhancement of the K+ current, IAHP, by reducing adenylyl cyclase activity in rat CA3 hippocampal neurons.

    PubMed

    Gerber, U; Gähwiler, B H

    1994-11-01

    1. Gamma-aminobuturic acid-B (GABAB) and adenosine A1 receptors, which are expressed in hippocampal pyramidal cells, are linked to pertussis toxin-sensitive G-proteins known to be coupled negatively to the enzyme adenylyl cyclase. This study investigates the electrophysiological consequences of adenylyl cyclase inhibition in response to stimulation of these receptors. 2. Single-electrode voltage-clamp recordings were obtained from CA3 pyramidal cells in rat hippocampal slice cultures in presence of tetrodotoxin. The calcium-dependent potassium current (IAHP), which is very sensitive to intracellular levels of adenosine 3',5'-cyclic monophosphate (cAMP), was used as an electrophysiological indicator of adenylyl cyclase activity. 3. Application of baclofen (10 microM), a selective agonist at GABAB receptors, or adenosine (50 microM) each resulted in a transient decrease followed by a significant enhancement in the amplitude of evoked IAHP. The initial reduction in amplitude of IAHP probably reflects inadequacies in voltage clamp of electronically distant dendritic sites, due to the shunting caused by concomitant activation of potassium conductance by baclofen/adenosine. Comparable increases in membrane conductance in response to the GABAA agonist, muscimol, caused a similar reduction in IAHP. The enhancement of IAHP is consistent with an inhibition of constitutively active adenylyl cyclase. 4. The receptor mediating the responses to adenosine was identified as belonging to the A1 subtype on the basis of its sensitivity to the selective antagonist 8-cyclopentyl-1,3-dipropylxanthine.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Evaluation of adenosine, lidocaine, and magnesium for enhancement of platelet function during storage.

    PubMed

    Bynum, James A; Taylor, Ashley S; Peltier, Grantham C; McIntosh, Colby S; Meledeo, Michael A; Dobson, Geoffrey P; Cap, Andrew P

    2017-07-01

    The combination of adenosine, lidocaine, and magnesium (Mg2+) (ALM) has demonstrated cardioprotective and resuscitative properties in models of cardiac arrest and hemorrhagic shock that are linked to reduction of metabolic demand. Platelets play a key role in resuscitation strategies for ATC but suffer from loss of function following storage in part owing to mitochondrial exhaustion. This study evaluates whether ALM also demonstrates protective properties in stored platelet preparations. Platelets were tested at (baseline, Day 5, Day 10, and Day 15) at 22°C (room temperature) or 4°C in 100% plasma and platelet additive solution. Adenosine, lidocaine, and magnesium treatment or its individual components (A, L, M, or combinations) were added directly to the minibags at baseline for storage. Measurements consisted of blood gas and chemistry analyses, thromboelastography, impedance aggregometry, and flow cytometry. Blood gas and cell analysis, as well as flow cytometry measures, demonstrated only differences between temperature groups starting at Day 5 (p < 0.05) and no differences between treatment groups. Aggregation response to collagen (A only, M only, and ALM high dose) and thrombin receptor activation peptide (A + M, and ALM high dose) was significantly greater at Day 5 compared to respective 4°C (100% plasma) controls (p < 0.05). Thromboelastography analysis revealed significant preservation of all measures (reaction time, maximum amplitude, and angle) at Day 15 for 4°C-stored samples in 100% plasma in both controls (no ALM) and ALM treatment compared to room temperature (p < 0.05); no differences were observed between the ALM and control groups. The mechanism of ALM's protective effect remains unclear; key cellular functions may be required to provide protection. In this study, improvements in collagen and thrombin receptor activation peptide aggregation were seen when compared to 4°C-stored plasma samples although no improvements were seen when compared

  1. Inhibition of muscarinic K+ current in guinea-pig atrial myocytes by PD 81,723, an allosteric enhancer of adenosine binding to A1 receptors

    PubMed Central

    Brandts, B; Bünemann, M; Hluchy, J; Sabin, G V; Pott, L

    1997-01-01

    PD 81,723 has been shown to enhance binding of adenosine to A1 receptors by stabilizing G protein-receptor coupling (‘allosteric enhancement'). Evidence has been provided that in the perfused hearts and isolated atria PD 81,723 causes a sensitization to adenosine via this mechanism. We have studied the effect of PD 81,723 in guinea-pig isolated atrial myocytes by use of whole-cell measurement of the muscarinic K+ current (IK(ACh)) activated by different Gi-coupled receptors (A1, M2, sphingolipid). PD 81,273 caused inhibition of IK(ACh) (IC50≃5 μM) activated by either of the three receptors. Receptor-independent IK(ACh) in cells loaded with GTP-γ-S and background IK(ACh), which contributes to the resting conductance of atrial myocytes, were equally sensitive to PD 81,723. At no combination of concentrations of adenosine and PD 81,723 could an enhancing effect be detected. The compound was active from the outside only. Loading of the cells with PD 81,723 (50 μM) via the patch pipette did not affect either IK(ACh) or its sensitivity to adenosine. We suggest that PD 81,723 acts as an inhibitor of inward rectifying K+ channels; this is supported by the finding that ventricular IK1, which shares a large degree of homology with the proteins (GIRK1/GIRK4) forming IK(ACh) but is not G protein-gated, was also blocked by this compound. It is concluded that the functional effects of PD 81,723 described in the literature are not mediated by the A1 adenosine receptor-Gi-IK(ACh) pathway. PMID:9249260

  2. 5'-Monophosphate-activated protein kinase (AMPK) improves autophagic activity in diabetes and diabetic complications.

    PubMed

    Yao, Fan; Zhang, Ming; Chen, Li

    2016-01-01

    Diabetes mellitus (DM), an endocrine disorder, will be one of the leading causes of death world-wide in about two decades. Cellular injuries and disorders of energy metabolism are two key factors in the pathogenesis of diabetes, which also become the important causes for the process of diabetic complications. AMPK is a key enzyme in maintaining metabolic homeostasis and has been implicated in the activation of autophagy in distinct tissues. An increasing number of researchers have confirmed that autophagy is a potential factor to affect or induce diabetes and its complications nowadays, which could remove cytotoxic proteins and dysfunctional organelles. This review will summarize the regulation of autophagy and AMPK in diabetes and its complications, and explore how AMPK stimulates autophagy in different diabetic syndromes. A deeper understanding of the regulation and activity of AMPK in autophagy would enhance its development as a promising therapeutic target for diabetes treatment.

  3. Caffeine and adenosine.

    PubMed

    Ribeiro, Joaquim A; Sebastião, Ana M

    2010-01-01

    Caffeine causes most of its biological effects via antagonizing all types of adenosine receptors (ARs): A1, A2A, A3, and A2B and, as does adenosine, exerts effects on neurons and glial cells of all brain areas. In consequence, caffeine, when acting as an AR antagonist, is doing the opposite of activation of adenosine receptors due to removal of endogenous adenosinergic tonus. Besides AR antagonism, xanthines, including caffeine, have other biological actions: they inhibit phosphodiesterases (PDEs) (e.g., PDE1, PDE4, PDE5), promote calcium release from intracellular stores, and interfere with GABA-A receptors. Caffeine, through antagonism of ARs, affects brain functions such as sleep, cognition, learning, and memory, and modifies brain dysfunctions and diseases: Alzheimer's disease, Parkinson's disease, Huntington's disease, Epilepsy, Pain/Migraine, Depression, Schizophrenia. In conclusion, targeting approaches that involve ARs will enhance the possibilities to correct brain dysfunctions, via the universally consumed substance that is caffeine.

  4. Supplementation of exogenous adenosine 5'-triphosphate enhances mechanical properties of 3D cell-agarose constructs for cartilage tissue engineering.

    PubMed

    Gadjanski, Ivana; Yodmuang, Supansa; Spiller, Kara; Bhumiratana, Sarindr; Vunjak-Novakovic, Gordana

    2013-10-01

    Formation of tissue-engineered cartilage is greatly enhanced by mechanical stimulation. However, direct mechanical stimulation is not always a suitable method, and the utilization of mechanisms underlying mechanotransduction might allow for a highly effective and less aggressive alternate means of stimulation. In particular, the purinergic, adenosine 5'-triphosphate (ATP)-mediated signaling pathway is strongly implicated in mechanotransduction within the articular cartilage. We investigated the effects of transient and continuous exogenous ATP supplementation on mechanical properties of cartilaginous constructs engineered using bovine chondrocytes and human mesenchymal stem cells (hMSCs) encapsulated in an agarose hydrogel. For both cell types, we have observed significant increases in equilibrium and dynamic compressive moduli after transient ATP treatment applied in the fourth week of cultivation. Continuous ATP treatment over 4 weeks of culture only slightly improved the mechanical properties of the constructs, without major changes in the total glycosaminoglycan (GAG) and collagen content. Structure-function analyses showed that transiently ATP-treated constructs, and in particular those based on hMSCs, had the highest level of correlation between compositional and mechanical properties. Transiently treated groups showed intense staining of the territorial matrix for GAGs and collagen type II. These results indicate that transient ATP treatment can improve functional mechanical properties of cartilaginous constructs based on chondrogenic cells and agarose hydrogels, possibly by improving the structural organization of the bulk phase and territorial extracellular matrix (ECM), that is, by increasing correlation slopes between the content of the ECM components (GAG, collagen) and mechanical properties of the construct.

  5. Bi-stage control of dissolved oxygen to enhance cyclic adenosine monophosphate production by Arthrobacter A302.

    PubMed

    Li, Lei; Chen, Xiaochun; Cheng, Jian; Zhang, Chunwei; Bai, Jianxin; Chen, Yong; Niu, Huanqin; Ying, Hanjie

    2012-10-01

    Experiments confirmed dissolved oxygen (DO) definitely affects cyclic adenosine monophosphate (cAMP) production by Arthrobacter A302. Production of cAMP by batch fermentation was investigated under various DO conditions. A two-stage DO control strategy was proposed to achieve optimal production of cAMP based on the kinetic analysis: the DO level was controlled at 40% during the first 18 h and then switched to 30%. Relatively high cAMP production (9.9 g L(-1)) was achieved by applying this strategy. The cAMP productivity (0.14 g L(-1) h(-1)) was also successfully improved by 85.1, 59.3, 15.1 and 28.0%, compared to cases in which DO was uncontrolled or DO levels were held at 20, 30 and 40%, respectively. This is the first report of the use of a two-stage DO control strategy in cAMP production, and it was verified to be an effective method for enhancing the cAMP yield via this strain.

  6. Enhancing pigmentation via activation of A3 adenosine receptors in B16 melanoma cells and in human skin explants.

    PubMed

    Madi, Lea; Rosenberg-Haggen, Bianca; Nyska, Abraham; Korenstein, Rafi

    2013-01-01

    A3 adenosine receptor, A3AR, belongs to the Gi proteins coupled receptors. Activation of A3AR by its agonist, IB-MECA, decreases cAMP and was expected to reduce melanin level. Unexpectedly, B16 melanoma cells exposed to IB-MECA increased melanin levels in a dose-dependent manner. Human skin explants exposed to IB-MECA showed an increase in DOPA positive cells and in melanin deposition in keratinocytes. The agonist induced AKT phosphorylation, leading to a rapid translocation of the transcription factor MiTF towards the nucleus. DOPA oxidase activity and melanin levels induced by IB-MECA were further enhanced by PD98509, an inhibitor ERK signalling pathway. Our study shows that IB-MECA decreases cAMP while inducing melanogenesis. The proposed mechanism involves activation of PI3K/AKT signalling pathway by β/γ subunits of the G protein coupled to A3AR. The increase in melanin level in human skin explants suggests that IB-MECA may be a potential candidate to the treatment of hypopigmentation of skin. © 2013 John Wiley & Sons A/S.

  7. Adenosine-associated delivery systems.

    PubMed

    Kazemzadeh-Narbat, Mehdi; Annabi, Nasim; Tamayol, Ali; Oklu, Rahmi; Ghanem, Amyl; Khademhosseini, Ali

    2015-01-01

    Adenosine is a naturally occurring purine nucleoside in every cell. Many critical treatments such as modulating irregular heartbeat (arrhythmias), regulation of central nervous system (CNS) activity and inhibiting seizural episodes can be carried out using adenosine. Despite the significant potential therapeutic impact of adenosine and its derivatives, the severe side effects caused by their systemic administration have significantly limited their clinical use. In addition, due to adenosine's extremely short half-life in human blood (<10 s), there is an unmet need for sustained delivery systems to enhance efficacy and reduce side effects. In this article, various adenosine delivery techniques, including encapsulation into biodegradable polymers, cell-based delivery, implantable biomaterials and mechanical-based delivery systems, are critically reviewed and the existing challenges are highlighted.

  8. Carbamazepine and oxcarbazepine, but not eslicarbazepine, enhance excitatory synaptic transmission onto hippocampal CA1 pyramidal cells through an antagonist action at adenosine A1 receptors.

    PubMed

    Booker, Sam A; Pires, Nuno; Cobb, Stuart; Soares-da-Silva, Patrício; Vida, Imre

    2015-06-01

    This study assessed the anticonvulsant and seizure generation effects of carbamazepine (CBZ), oxcarbazepine (OXC) and eslicarbazepine (S-Lic) in wild-type mice. Electrophysiological recordings were made to discriminate potential cellular and synaptic mechanisms underlying anti- and pro-epileptic actions. The anticonvulsant and pro-convulsant effects were evaluated in the MES, the 6-Hz and the Irwin tests. Whole-cell patch-clamp recordings were used to investigate the effects on fast excitatory and inhibitory synaptic transmission in hippocampal area CA1. The safety window for CBZ, OXC and eslicarbazepine (ED50 value against the MES test and the dose that produces grade 5 convulsions in all mice), was 6.3, 6.0 and 12.5, respectively. At high concentrations the three drugs reduced synaptic transmission. CBZ and OXC enhanced excitatory postsynaptic currents (EPSCs) at low, therapeutically-relevant concentrations. These effects were associated with no change in inhibitory postsynaptic currents (IPSCs) resulting in altered balance between excitation and inhibition. S-Lic had no effect on EPSC or IPSC amplitudes over the same concentration range. The CBZ mediated enhancement of EPSCs was blocked by DPCPX, a selective antagonist, and occluded by CCPA, a selective agonist of the adenosine A1 receptor. Furthermore, reduction of endogenous adenosine by application of the enzyme adenosine deaminase also abolished the CBZ- and OXC-induced increase of EPSCs, indicating that the two drugs act as antagonists at native adenosine receptors. In conclusion, CBZ and OXC possess pro-epileptic actions at clinically-relevant concentrations through the enhancement of excitatory synaptic transmission. S-Lic by comparison has no such effect on synaptic transmission, explaining its lack of seizure exacerbation.

  9. Extracellular adenosine production by ecto-5′-nucleotidase (CD73) enhances radiation-induced lung fibrosis

    PubMed Central

    Wirsdörfer, Florian; de Leve, Simone; Cappuccini, Federica; Eldh, Therese; Meyer, Alina V.; Gau, Eva; Thompson, Linda F.; Chen, Ning-Yuan; Karmouty-Quintana, Harry; Fischer, Ute; Kasper, Michael; Klein, Diana; Ritchey, Jerry W.; Blackburn, Michael R.; Westendorf, Astrid M.; Stuschke, Martin; Jendrossek, Verena

    2016-01-01

    Radiation-induced pulmonary fibrosis is a severe side effect of thoracic irradiation, but its pathogenesis remains poorly understood and no effective treatment is available. In this study, we investigated the role of the extracellular adenosine as generated by the ecto-5'-nucleotidase CD73 in fibrosis development after thoracic irradiation. Exposure of wild-type C57BL/6 mice to a single dose (15 Gray) of whole thorax irradiation triggered a progressive increase in CD73 activity in the lung between 3 and 30 weeks post-irradiation. In parallel, adenosine levels in bronchoalveolar lavage fluid (BALF) were increased by approximately three-fold. Histological evidence of lung fibrosis was observed by 25 weeks after irradiation. Conversely, CD73-deficient mice failed to accumulate adenosine in BALF and exhibited significantly less radiation-induced lung fibrosis (P<0.010). Furthermore, treatment of wild-type mice with pegylated adenosine deaminase (PEG-ADA) or CD73 antibodies also significantly reduced radiation-induced lung fibrosis. Taken together, our findings demonstrate that CD73 potentiates radiation-induced lung fibrosis, suggesting that existing pharmacological strategies for modulating adenosine may be effective in limiting lung toxicities associated with the treatment of thoracic malignancies. PMID:26921334

  10. Intraventricular injection of agents that enhance cyclic adenosine monophosphate formation leads to inhibition of proestrous luteinizing hormone surge in rats.

    PubMed

    Taleisnik, S; Haymal, B; Caligaris, L

    1993-09-01

    The effect of increasing hypothalamic levels of 3',5'-cyclic adenosine monophosphate (cAMP) on the preovulatory surge of luteinizing hormone (LH) and ovulation was studied in cycling rats. Animals hearing chronically implanted guiding cannulae into the third ventricle were injected with agents known to enhance the cellular levels of cAMP. Hourly blood samples from the unanesthetized, unrestrained rats were obtained between 11.00 and 17.00 h through a plastic cannula inserted into the jugular vein. Intraventricular injections of serotonin (7.5 mg/ml; 2 microliters) in the morning of proestrous blocked the preovulatory surge of LH and ovulation. This effect was assigned to an increased neuronal level of cAMP because it was prevented by a serum anti-cAMP. Third-ventricle injections of 2 microliters of forskolin (0.5 mmol/l), guanosine 5'-O-(3-thiotriphosphate)(2 mmol/l) or dibutyryl-cAMP (1 mmol/l) at 11.00 h on the day of proestrus mimicked the inhibitory effect of serotonin on the proestrous release of LH. It is suggested that serotonin inhibits LH surge by acting directly on LH-releasing hormone neurons and/or on neurons that provide inputs to these neurons involving cAMP as a second messenger. Neurons releasing gamma-aminobutyric acid (GABA) may serve as interneurons sensitive to serotonin, as well as to cAMP, inasmuch as the inhibitory effect of forskolin on the release of LH was partially blocked by the GABA antagonists, picrotoxin and bicuculline.

  11. Ex vivo Perfusion with Adenosine A2A Receptor Agonist Enhances Rehabilitation of Murine Donor Lungs after Circulatory Death

    PubMed Central

    Stone, Mathew L.; Sharma, Ashish K.; Mas, Valeria. R.; Gehrau, Ricardo C.; Mulloy, Daniel P.; Zhao, Yunge; Lau, Christine L.; Kron, Irving L.; Laubach, Victor E.

    2015-01-01

    Background Ex vivo lung perfusion (EVLP) enables assessment and rehabilitation of marginal donor lungs prior to transplantation. We previously demonstrated that adenosine A2A receptor (A2AR) agonism attenuates lung ischemia-reperfusion injury. The current study utilizes a novel murine EVLP model to test the hypothesis that A2AR agonist enhances EVLP-mediated rehabilitation of donation after circulatory death (DCD) lungs. Methods Mice underwent euthanasia and 60 min warm ischemia, and lungs were flushed with Perfadex and underwent cold static preservation (CSP, 60 min). Three groups were studied: no EVLP (CSP), EVLP with Steen solution for 60 min (EVLP), and EVLP with Steen solution supplemented with ATL1223, a selective A2AR agonist (EVLP+ATL1223). Lung function, wet/dry weight, cytokines and neutrophil numbers were measured. Microarrays were performed using the Affymetrix GeneChip Mouse Genome 430A 2.0 Array. Results EVLP significantly improved lung function versus CSP, which was further, significantly improved by EVLP+ATL1223. Lung edema, cytokines and neutrophil counts were reduced after EVLP and further, significantly reduced after EVLP+ATL1223. Gene array analysis revealed differential expression of 1,594 genes after EVLP, which comprise canonical pathways involved in inflammation and innate immunity including IL-1, IL-8, IL-6 and IL-17 signaling. Several pathways were uniquely regulated by EVLP+ATL1223 including the downregulation of genes involved in IL-1 signaling such as ADCY9, ECSIT, IRAK1, MAPK12 and TOLLIP. Conclusion EVLP modulates pro-inflammatory genes and reduces pulmonary dysfunction, edema and inflammation in DCD lungs, which are further reduced by A2AR agonism. This murine EVLP model provides a novel platform to study rehabilitative mechanisms of DCD lungs. PMID:26262504

  12. Ex Vivo Perfusion With Adenosine A2A Receptor Agonist Enhances Rehabilitation of Murine Donor Lungs After Circulatory Death.

    PubMed

    Stone, Matthew L; Sharma, Ashish K; Mas, Valeria R; Gehrau, Ricardo C; Mulloy, Daniel P; Zhao, Yunge; Lau, Christine L; Kron, Irving L; Huerter, Mary E; Laubach, Victor E

    2015-12-01

    Ex vivo lung perfusion (EVLP) enables assessment and rehabilitation of marginal donor lungs before transplantation. We previously demonstrated that adenosine A2A receptor (A2AR) agonism attenuates lung ischemia-reperfusion injury. The current study utilizes a novel murine EVLP model to test the hypothesis that A2AR agonist enhances EVLP-mediated rehabilitation of donation after circulatory death (DCD) lungs. Mice underwent euthanasia and 60 minutes warm ischemia, and lungs were flushed with Perfadex and underwent cold static preservation (CSP, 60 minutes). Three groups were studied: no EVLP (CSP), EVLP with Steen solution for 60 minutes (EVLP), and EVLP with Steen solution supplemented with ATL1223, a selective A2AR agonist (EVLP + ATL1223). Lung function, wet/dry weight, cytokines and neutrophil numbers were measured. Microarrays were performed using the Affymetrix GeneChip Mouse Genome 430A 2.0 Array. Ex vivo lung perfusion significantly improved lung function versus CSP, which was further, significantly improved by EVLP + ATL1223. Lung edema, cytokines, and neutrophil counts were reduced after EVLP and further, significantly reduced after EVLP + ATL1223. Gene array analysis revealed differential expression of 1594 genes after EVLP, which comprise canonical pathways involved in inflammation and innate immunity including IL-1, IL-8, IL-6, and IL-17 signaling. Several pathways were uniquely regulated by EVLP + ATL1223 including the downregulation of genes involved in IL-1 signaling, such as ADCY9, ECSIT, IRAK1, MAPK12, and TOLLIP. Ex vivo lung perfusion modulates proinflammatory genes and reduces pulmonary dysfunction, edema, and inflammation in DCD lungs, which are further reduced by A2AR agonism. This murine EVLP model provides a novel platform to study rehabilitative mechanisms of DCD lungs.

  13. Enhanced recovery of breathing capacity from combined adenosine 2A receptor inhibition and daily acute intermittent hypoxia after chronic cervical spinal injury

    PubMed Central

    Navarrete-Opazo, A.; Dougherty, B.J.; Mitchell, G.S.

    2016-01-01

    Daily acute intermittent hypoxia (dAIH) improves breathing capacity after C2 spinal hemisection (C2HS) in rats. Since C2HS disrupts spinal serotonergic innervation below the injury, adenosine-dependent mechanisms underlie dAIH-induced functional recovery 2 weeks post-injury. We hypothesized that dAIH-induced functional recovery converts from an adenosine-dependent to a serotonin-dependent, adenosine-constrained mechanism with chronic injury. Eight weeks post-C2HS, rats began dAIH (10, 5-min episodes, 10.5% O2; 5-min intervals; 7 days) followed by AIH 3× per week (3×wAIH) for 8 additional weeks with/without systemic A2A receptor inhibition (KW6002) on each AIH exposure day. Tidal volume (VT) and bilateral diaphragm (Dia) and T2 external intercostal motor activity were assessed in unanesthetized rats breathing air and during maximum chemoreflex stimulation (MCS: 7% CO2, 10.5% O2). Nine weeks post-C2HS, dAIH increased VT versus time controls (p < 0.05), an effect enhanced by KW6002 (p < 0.05). dAIH increased bilateral Dia activity (p < 0.05), and KW6002 enhanced this effect in contralateral (p < 0.05) and ipsilateral Dia activity (p < 0.001), but not T2 inspiratory activity. Functional benefits of combined AIH plus systemic A2A receptor inhibition were maintained for 4 weeks. Thus, in rats with chronic injuries: 1) dAIH improves VT and bilateral diaphragm activity; 2) VT recovery is enhanced by A2A receptor inhibition; and 3) functional recovery with A2A receptor inhibition and AIH “reminders” last 4 weeks. Combined dAIH and A2A receptor inhibition may be a simple, safe, and effective strategy to accelerate/enhance functional recovery of breathing capacity in patients with respiratory impairment from chronic spinal injury. PMID:27079999

  14. Adenosine receptors as drug targets — what are the challenges?

    PubMed Central

    Chen, Jiang-Fan; Eltzschig, Holger K.; Fredholm, Bertil B.

    2014-01-01

    Adenosine signalling has long been a target for drug development, with adenosine itself or its derivatives being used clinically since the 1940s. In addition, methylxanthines such as caffeine have profound biological effects as antagonists at adenosine receptors. Moreover, drugs such as dipyridamole and methotrexate act by enhancing the activation of adenosine receptors. There is strong evidence that adenosine has a functional role in many diseases, and several pharmacological compounds specifically targeting individual adenosine receptors — either directly or indirectly — have now entered the clinic. However, only one adenosine receptor-specific agent — the adenosine A2A receptor agonist regadenoson (Lexiscan; Astellas Pharma) — has so far gained approval from the US Food and Drug Administration (FDA). Here, we focus on the biology of adenosine signalling to identify hurdles in the development of additional pharmacological compounds targeting adenosine receptors and discuss strategies to overcome these challenges. PMID:23535933

  15. Adenosine-Associated Delivery Systems

    PubMed Central

    Kazemzadeh-Narbat, Mehdi; Annabi, Nasim; Tamayol, Ali; Oklu, Rahmi; Ghanem, Amyl; Khademhosseini, Ali

    2016-01-01

    Adenosine is a naturally occurring purine nucleoside in every cell. Many critical treatments such as modulating irregular heartbeat (arrhythmias), regulation of central nervous system (CNS) activity, and inhibiting seizural episodes can be carried out using adenosine. Despite the significant potential therapeutic impact of adenosine and its derivatives, the severe side effects caused by their systemic administration have significantly limited their clinical use. In addition, due to adenosine’s extremely short half-life in human blood (less than 10 s), there is an unmet need for sustained delivery systems to enhance efficacy and reduce side effects. In this paper, various adenosine delivery techniques, including encapsulation into biodegradable polymers, cell-based delivery, implantable biomaterials, and mechanical-based delivery systems, are critically reviewed and the existing challenges are highlighted. PMID:26453156

  16. Adenosine receptor targets for pain.

    PubMed

    Sawynok, J

    2016-12-03

    The main focus for the development of adenosine targets as analgesics to date has been A1Rs due to its antinociceptive profile in various preclinical pain models. The usefulness of systemic A1R agonists may be limited by other effects (cardiovascular, motor), but enhanced selectivity for pain might occur with partial agonists, potent and highly selective agonists, or allosteric modulators. A2AR agonists exhibit some peripheral pronociceptive effects, but also act on immune cells to suppress inflammation and on spinal glia to suppress pain signaling and may be useful for inflammatory and neuropathic pain. A2BR agonists exhibit peripheral proinflammatory effects on immune cells, but also spinal antinociceptive effects similar to A2AR agonists. A3Rs are now demonstrated to produce antinociception in several preclinical neuropathic pain models, with mechanistic actions on glial cells, and may be useful for neuropathic pain. Endogenous adenosine levels can be augmented by inhibition of metabolism (via adenosine kinase) or increased generation (via nucleotidases), and these approaches have implications for pain. Endogenous adenosine contributes to antinociception by several pharmacological agents, herbal remedies, acupuncture, transcutaneous electrical nerve stimulation, exercise, joint mobilization, and water immersion via spinal and/or peripheral effects, such that this system appears to constitute a major pain regulatory system. Finally, caffeine inhibits A1-, A2A- and A3Rs with similar potency, and dietary caffeine intake will need attention in trials of: (a) agonists and/or modulators acting at these receptors, (b) some pharmacological and herbal analgesics, and (c) manipulations that enhance endogenous adenosine levels, all of which are inhibited by caffeine and/or A1R antagonists in preclinical studies. All adenosine receptors have effects on spinal glial cells in regulating nociception, and gender differences in the involvement of such cells in chronic

  17. Enhanced recovery of breathing capacity from combined adenosine 2A receptor inhibition and daily acute intermittent hypoxia after chronic cervical spinal injury.

    PubMed

    Navarrete-Opazo, A; Dougherty, B J; Mitchell, G S

    2017-01-01

    Daily acute intermittent hypoxia (dAIH) improves breathing capacity after C2 spinal hemisection (C2HS) in rats. Since C2HS disrupts spinal serotonergic innervation below the injury, adenosine-dependent mechanisms underlie dAIH-induced functional recovery 2weeks post-injury. We hypothesized that dAIH-induced functional recovery converts from an adenosine-dependent to a serotonin-dependent, adenosine-constrained mechanism with chronic injury. Eight weeks post-C2HS, rats began dAIH (10, 5-min episodes, 10.5% O2; 5-min intervals; 7days) followed by AIH 3× per week (3×wAIH) for 8 additional weeks with/without systemic A2A receptor inhibition (KW6002) on each AIH exposure day. Tidal volume (VT) and bilateral diaphragm (Dia) and T2 external intercostal motor activity were assessed in unanesthetized rats breathing air and during maximum chemoreflex stimulation (MCS: 7% CO2, 10.5% O2). Nine weeks post-C2HS, dAIH increased VT versus time controls (p<0.05), an effect enhanced by KW6002 (p<0.05). dAIH increased bilateral Dia activity (p<0.05), and KW6002 enhanced this effect in contralateral (p<0.05) and ipsilateral Dia activity (p<0.001), but not T2 inspiratory activity. Functional benefits of combined AIH plus systemic A2A receptor inhibition were maintained for 4weeks. Thus, in rats with chronic injuries: 1) dAIH improves VT and bilateral diaphragm activity; 2) VT recovery is enhanced by A2A receptor inhibition; and 3) functional recovery with A2A receptor inhibition and AIH "reminders" last 4weeks. Combined dAIH and A2A receptor inhibition may be a simple, safe, and effective strategy to accelerate/enhance functional recovery of breathing capacity in patients with respiratory impairment from chronic spinal injury. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. Enhancement of AMPA currents and GluR1 membrane expression through PKA-coupled adenosine A(2A) receptors.

    PubMed

    Dias, Raquel B; Ribeiro, Joaquim A; Sebastião, Ana M

    2012-02-01

    Phosphorylation of glutamate α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors by Protein Kinase A (PKA) is known to regulate AMPA receptor (AMPAR) trafficking and stabilization at the postsynaptic membrane, which in turn is one of the key mechanisms by which synaptic transmission and plasticity are tuned. However, not much is known as to how Gs-coupled receptors contribute to endogenous PKA-mediated regulation of AMPA receptor function. Here we report that activation of the excitatory A(2A) adenosine receptor by 2-[4-(2-p-carboxyethyl)phenylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680, 1-30 nM) facilitates AMPA-evoked currents in CA1 pyramidal neurons, by a mechanism dependent on PKA activation, but not on protein synthesis. This modulation of AMPA currents was mimicked by forskolin (1 μM) and did not occur in stratum radiatum interneurons. Superfusion of the A(2A) receptor agonist also caused an increase in the amplitude of miniature excitatory postsynaptic currents (mEPSCs), as well as in the membrane levels of GluR1 subunits phosphorylated at the PKA site (Ser845). The impact of this increase on GluR1-containing AMPA receptor expression was evidenced by the potentiation of LTP at the CA3-CA1 synapse that followed brief activation of A(2A) receptors. We thus propose that in conditions of increased adenosine availability, A(2A) receptor activation is responsible for setting part of the endogenous GluR1 Ser-845 phosphorylation tonus and hence, the availability of the GluR1-containing AMPA receptor extrasynaptic pool for synaptic insertion and reinforcement of synaptic strength.

  19. Adenosine kinase-deficient mutant of Saccharomyces cerevisiae accumulates S-adenosylmethionine because of an enhanced methionine biosynthesis pathway.

    PubMed

    Kanai, Muneyoshi; Masuda, Mitsunori; Takaoka, Yasumichi; Ikeda, Hiroko; Masaki, Kazuo; Fujii, Tsutomu; Iefuji, Haruyuki

    2013-02-01

    To isolate an S-adenosylmethionine (SAM)-accumulating yeast strain and to develop a more efficient method of producing SAM, we screened methionine-resistant strains using the yeast deletion library of budding yeast and isolated 123 strains. The SAM content in 81 of the 123 strains was higher than that in the parental strain BY4742. We identified ADO1 encoding adenosine kinase as one of the factors participating in high SAM accumulation. The X∆ado1 strain that was constructed from the X2180-1A strain (MAT a, ATCC 26786) could accumulate approximately 30-fold (18 mg/g dry cell weight) more SAM than the X2180-1A strain in yeast extract peptone dextrose medium. Furthermore, we attempted to identify the molecular basis underlying the differences in SAM accumulation between X∆ado1 and X2180-1A strains. DNA microarray analysis revealed that the genes involved in the methionine biosynthesis pathway, phosphate metabolism, and hexose transport were mainly overexpressed in the X∆ado1 strain compared with the X2180-1A strain. We also determined the levels of various metabolites involved in the methionine biosynthesis pathway and found increased content of SAM, tetrahydrofolate (THF), inorganic phosphate, polyphosphoric acid, and S-adenosylhomocysteine in the X∆ado1 strain. In contrast, the content of 5-methyl-THF, homocysteine, glutathione, and adenosine was decreased. These results indicated that the ∆ado1 strain could accumulate SAM because of preferential activation of the methionine biosynthesis pathway.

  20. Adenosine and Ischemic Preconditioning

    PubMed Central

    Liang, Bruce T.; Swierkosz, Tomasz A.; Herrmann, Howard C.; Kimmel, Stephen; Jacobson, Kenneth A.

    2012-01-01

    Adenosine is released in large amounts during myocardial ischemia and is capable of exerting potent cardioprotective effects in the heart. Although these observations on adenosine have been known for a long time, how adenosine acts to achieve its anti-ischemic effect remains incompletely understood. However, recent advances on the chemistry and pharmacology of adenosine receptor ligands have provided important and novel information on the function of adenosine receptor subtypes in the cardiovascular system. The development of model systems for the cardiac actions of adenosine has yielded important insights into its mechanism of action and have begun to elucidate the sequence of signalling events from receptor activation to the actual exertion of its cardioprotective effect. The present review will focus on the adenosine receptors that mediate the potent anti-ischemic effect of adenosine, new ligands at the receptors, potential molecular signalling mechanisms downstream of the receptor, mediators for cardioprotection, and possible clinical applications in cardiovascular disorders. PMID:10607860

  1. High-salt diet enhances mouse aortic relaxation through adenosine A2A receptor via CYP epoxygenases.

    PubMed

    Nayeem, Mohammed A; Ponnoth, Dovenia S; Boegehold, Matthew A; Zeldin, Darryl C; Falck, John R; Mustafa, S Jamal

    2009-03-01

    We hypothesize that A(2A) adenosine receptors (A(2A) AR) promote aortic relaxation in mice through cytochrome P450 (CYP)-epoxygenases and help to avoid salt sensitivity. Aortas from male mice maintained on a high-salt (HS; 7% NaCl) or normal-salt (NS; 0.45% NaCl) diet for 4-5 wks were used. Concentration-response curves (10(-11)-10(-5) M) for 5'-N-ethylcarboxamidoadenosine (NECA; a nonselective adenosine analog) and CGS 21680 (A(2A) AR agonist) were obtained with different antagonists including ZM 241385 (A(2A) AR antagonist; 10(-6) M), SCH 58261 (A(2A) AR antagonist; 10(-6) M), N(omega)-nitro-l-arginine methyl ester (l-NAME; endothelial nitric oxide synthase inhibitor; 10(-4) M) and inhibitors including methylsulfonyl-propargyloxyphenylhexanamide (MS-PPOH; CYP epoxygenases inhibitor; 10(-5)M), 14,15-epoxyeicosa-5(z)-enoic acid (14,15-EEZE; EET antagonist; 10(-5)M), dibromo-dodecenyl-methylsulfimide (DDMS; CYP4A inhibitor; 10(-5)M), and HET0016 (20-HETE inhibitor; 10(-5)M). At 10(-7) M of NECA, significant relaxation in HS (+22.58 +/- 3.12%) was observed compared with contraction in NS (-10.62 +/- 6.27%, P < 0.05). ZM 241385 changed the NECA response to contraction (P < 0.05) in HS. At 10(-7) M of CGS 21680, significant relaxation in HS (+32.04 +/- 3.08%) was observed compared with NS (+10.45 +/- 1.34%, P < 0.05). SCH 58261, l-NAME, MS-PPOH, and 14,15-EEZE changed the CGS 21680-induced relaxation to contraction (P < 0.05) in HS. Interestingly, DDMS and HET0016 changed CGS 21680 response to relaxation (P < 0.05) in NS; however, there was no significant difference found between DDMS, HET0016-treated HS and NS vs. nontreated HS group (P > 0.05). CYP2C29 protein was 55% and 74% upregulated in HS vs. NS (P < 0.05) mice aorta and kidney, respectively. CYP4A protein was 30.30% and 35.70% upregulated in NS vs. HS (P < 0.05) mice aorta and kidneys, respectively. A(1) AR was downregulated, whereas A(2A) AR was upregulated in HS compared with NS. These data suggest that HS

  2. High-salt diet enhances mouse aortic relaxation through adenosine A2A receptor via CYP epoxygenases

    PubMed Central

    Nayeem, Mohammed A.; Ponnoth, Dovenia S.; Boegehold, Matthew A.; Zeldin, Darryl C.; Falck, John R.; Mustafa, S. Jamal

    2009-01-01

    We hypothesize that A2A adenosine receptors (A2A AR) promote aortic relaxation in mice through cytochrome P450 (CYP)-epoxygenases and help to avoid salt sensitivity. Aortas from male mice maintained on a high-salt (HS; 7% NaCl) or normal-salt (NS; 0.45% NaCl) diet for 4–5 wks were used. Concentration-response curves (10−11–10−5 M) for 5′-N-ethylcarboxamidoadenosine (NECA; a nonselective adenosine analog) and CGS 21680 (A2A AR agonist) were obtained with different antagonists including ZM 241385 (A2A AR antagonist; 10−6 M), SCH 58261 (A2A AR antagonist; 10−6 M), Nω-nitro-l-arginine methyl ester (l-NAME; endothelial nitric oxide synthase inhibitor; 10−4 M) and inhibitors including methylsulfonyl-propargyloxyphenylhexanamide (MS-PPOH; CYP epoxygenases inhibitor; 10−5M), 14,15-epoxyeicosa-5(z)-enoic acid (14,15-EEZE; EET antagonist; 10−5M), dibromo-dodecenyl-methylsulfimide (DDMS; CYP4A inhibitor; 10−5M), and HET0016 (20-HETE inhibitor; 10−5M). At 10−7 M of NECA, significant relaxation in HS (+22.58 ± 3.12%) was observed compared with contraction in NS (−10.62 ± 6.27%, P < 0.05). ZM 241385 changed the NECA response to contraction (P < 0.05) in HS. At 10−7 M of CGS 21680, significant relaxation in HS (+32.04 ± 3.08%) was observed compared with NS (+10.45 ± 1.34%, P < 0.05). SCH 58261, l-NAME, MS-PPOH, and 14,15-EEZE changed the CGS 21680-induced relaxation to contraction (P < 0.05) in HS. Interestingly, DDMS and HET0016 changed CGS 21680 response to relaxation (P < 0.05) in NS; however, there was no significant difference found between DDMS, HET0016-treated HS and NS vs. nontreated HS group (P > 0.05). CYP2C29 protein was 55% and 74% upregulated in HS vs. NS (P < 0.05) mice aorta and kidney, respectively. CYP4A protein was 30.30% and 35.70% upregulated in NS vs. HS (P < 0.05) mice aorta and kidneys, respectively. A1 AR was downregulated, whereas A2A AR was upregulated in HS compared with NS. These data suggest that HS may activate CYP2

  3. Enhanced Actions of Adenosine in Medial Entorhinal Cortex Layer II Stellate Neurons in Temporal Lobe Epilepsy are Mediated via A1 Receptor Activation

    PubMed Central

    Hargus, Nicholas J.; Jennings, Conor; Perez-Reyes, Edward; Bertram, Edward H.; Patel, Manoj K.

    2011-01-01

    Summary Purpose The adenosinergic system is known to exert an inhibitory affect in the brain and as such adenosine has been considered an endogenous anticonvulsant. Entorhinal cortex (EC) layer II neurons, which serve as the primary input to the hippocampus, are spared in temporal lobe epilepsy (TLE) and become hyperexcitable. Since these neurons also express adenosine receptors, the activity of these neurons may be controlled by adenosine, specifically during seizure activity when adenosine levels are thought to rise. In light of this, we determined if the actions of adenosine on medial EC (mEC) layer II stellate neurons are augmented in TLE and by which receptor subtype. Methods Horizontal brain slices were prepared from rats exhibiting spontaneous seizures (TLE) induced by electrical stimulation and compared with age matched control rats. mEC layer II stellate neurons were visually identified and action potentials (AP) evoked by either a series of depolarizing current injection steps or via presynaptic stimulation of mEC deep layers. The effects of adenosine were compared with actions of adenosine A1 and A2A receptor-specific agonists (CPA and CGS 21680) and antagonists (DPCPX and ZM241385) respectively. Immunohistochemical and qPCR techniques were also employed to assess relative adenosine A1 receptor message and expression. Key Findings mEC layer II stellate neurons were hyper-excitable in TLE, evoking a higher frequency of AP's when depolarized and generating bursts of AP's when synaptically stimulated. Adenosine reduced AP frequency and synaptically evoked AP's in a dose dependent manner (500 nM – 100 μM); however, in TLE, the inhibitory actions of adenosine occurred at concentrations that were without affect in control neurons. In both cases, the inhibitory actions of adenosine were mediated via activation of the A1 and not the A2A receptor subtype. qPCR and immunohistochemical experiments revealed an up-regulation of the adenosine A1 mRNA and an

  4. Caffeine intensifies taste of certain sweeteners: role of adenosine receptor.

    PubMed

    Schiffman, S S; Diaz, C; Beeker, T G

    1986-03-01

    Caffeine, a potent antagonist of adenosine receptors, potentiates the taste of some but not all sweeteners. It significantly enhances the taste of acesulfam-K, neohesperidin dihydrochalcone, d-tryptophan, thaumatin, stevioside, and sodium saccharin. Adenosine reverses the enhancement. Caffeine has no effect on aspartame, sucrose, fructose, and calcium cyclamate. These results suggest that the inhibitory A1 adenosine receptor plays an important local role in modulating the taste intensity of certain sweeteners and that several transduction mechanisms mediate sweet taste.

  5. Neurabin scaffolding of adenosine receptor and RGS4 regulates anti-seizure effect of endogenous adenosine

    PubMed Central

    Chen, Yunjia; Liu, Yin; Cottingham, Christopher; McMahon, Lori; Jiao, Kai; Greengard, Paul; Wang, Qin

    2012-01-01

    Endogenous adenosine is an essential protective agent against neural damage by various insults to the brain. However, the therapeutic potential of adenosine receptor-directed ligands for neuroprotection is offset by side effects in peripheral tissues and organs. An increase in adenosine receptor responsiveness to endogenous adenosine would enhance neuroprotection while avoiding the confounding effects of exogenous ligands. Here we report novel regulation of adenosine-evoked responses by a neural tissue-specific protein, neurabin. Neurabin attenuated adenosine A1 receptor (A1R) signaling by assembling a complex between the A1R and the regulator of G protein signaling 4 (RGS4), a protein known to turn off G protein signaling. Inactivation of the neurabin gene enhanced A1R signaling and promoted the protective effect of adenosine against excitotoxic seizure and neuronal death in mice. Furthermore, administration of a small molecule inhibitor of RGS4 significantly attenuated seizure severity in mice. Notably, the dose of kainate capable of inducing an ~50% rate of death in WT mice did not affect neurabin null mice or WT mice co-treated with an RGS4 inhibitor. The enhanced anti-seizure and neuroprotective effect achieved by disruption of the A1R/neurabin/RGS4 complex is elicited by the on-site and on-demand release of endogenous adenosine, and does not require administration of A1R ligands. These data identify neurabin-RGS4 as a novel tissue-selective regulatory mechanism for fine-tuning adenosine receptor function in the nervous system. Moreover, these findings implicate the A1R/neurabin/RGS4 complex as a valid therapeutic target for specifically manipulating the neuroprotective effects of endogenous adenosine. PMID:22357852

  6. Neurabin scaffolding of adenosine receptor and RGS4 regulates anti-seizure effect of endogenous adenosine.

    PubMed

    Chen, Yunjia; Liu, Yin; Cottingham, Christopher; McMahon, Lori; Jiao, Kai; Greengard, Paul; Wang, Qin

    2012-02-22

    Endogenous adenosine is an essential protective agent against neural damage by various insults to the brain. However, the therapeutic potential of adenosine receptor-directed ligands for neuroprotection is offset by side effects in peripheral tissues and organs. An increase in adenosine receptor responsiveness to endogenous adenosine would enhance neuroprotection while avoiding the confounding effects of exogenous ligands. Here we report novel regulation of adenosine-evoked responses by a neural tissue-specific protein, neurabin. Neurabin attenuated adenosine A(1) receptor (A1R) signaling by assembling a complex between the A1R and the regulator of G-protein signaling 4 (RGS4), a protein known to turn off G-protein signaling. Inactivation of the neurabin gene enhanced A1R signaling and promoted the protective effect of adenosine against excitotoxic seizure and neuronal death in mice. Furthermore, administration of a small molecule inhibitor of RGS4 significantly attenuated seizure severity in mice. Notably, the dose of kainate capable of inducing an ∼50% rate of death in wild-type (WT) mice did not affect neurabin-null mice or WT mice cotreated with an RGS4 inhibitor. The enhanced anti-seizure and neuroprotective effect achieved by disruption of the A1R/neurabin/RGS4 complex is elicited by the on-site and on-demand release of endogenous adenosine, and does not require administration of A1R ligands. These data identify neurabin-RGS4 as a novel tissue-selective regulatory mechanism for fine-tuning adenosine receptor function in the nervous system. Moreover, these findings implicate the A1R/neurabin/RGS4 complex as a valid therapeutic target for specifically manipulating the neuroprotective effects of endogenous adenosine.

  7. Adenosine and preeclampsia.

    PubMed

    Salsoso, Rocío; Farías, Marcelo; Gutiérrez, Jaime; Pardo, Fabián; Chiarello, Delia I; Toledo, Fernando; Leiva, Andrea; Mate, Alfonso; Vázquez, Carmen M; Sobrevia, Luis

    2017-06-01

    Adenosine is an endogenous nucleoside with pleiotropic effects in different physiological processes including circulation, renal blood flow, immune function, or glucose homeostasis. Changes in adenosine membrane transporters, adenosine receptors, and corresponding intracellular signalling network associate with development of pathologies of pregnancy, including preeclampsia. Preeclampsia is a cause of maternal and perinatal morbidity and mortality affecting 3-5% of pregnancies. Since the proposed mechanisms of preeclampsia development include adenosine-dependent biological effects, adenosine membrane transporters and receptors, and the associated signalling mechanisms might play a role in the pathophysiology of preeclampsia. Preeclampsia associates with increased adenosine concentration in the maternal blood and placental tissue, likely due to local hypoxia and ischemia (although not directly demonstrated), microthrombosis, increased catecholamine release, and platelet activation. In addition, abnormal expression and function of equilibrative nucleoside transporters is described in foetoplacental tissues from preeclampsia; however, the role of adenosine receptors in the aetiology of this disease is not well understood. Adenosine receptors activation may be related to abnormal trophoblast invasion, angiogenesis, and ischemia/reperfusion mechanisms in the placenta from preeclampsia. These mechanisms may explain only a low fraction of the associated abnormal transformation of spiral arteries in preeclampsia, triggering cellular stress and inflammatory mediators release from the placenta to the maternal circulation. Although increased adenosine concentration in preeclampsia may be a compensatory or adaptive mechanism favouring placental angiogenesis, a poor angiogenic state is found in preeclampsia. Thus, preeclampsia-associated complications might affect the cell response to adenosine due to altered expression and activity of adenosine receptors, membrane transporters

  8. Adenosine Monophosphate (AMP)-Activated Protein Kinase: A New Target for Nutraceutical Compounds

    PubMed Central

    Marín-Aguilar, Fabiola; Pavillard, Luis E.; Giampieri, Francesca; Bullón, Pedro; Cordero, Mario D.

    2017-01-01

    Adenosine monophosphate-activated protein kinase (AMPK) is an important energy sensor which is activated by increases in adenosine monophosphate (AMP)/adenosine triphosphate (ATP) ratio and/or adenosine diphosphate (ADP)/ATP ratio, and increases different metabolic pathways such as fatty acid oxidation, glucose transport and mitochondrial biogenesis. In this sense, AMPK maintains cellular energy homeostasis by induction of catabolism and inhibition of ATP-consuming biosynthetic pathways to preserve ATP levels. Several studies indicate a reduction of AMPK sensitivity to cellular stress during aging and this could impair the downstream signaling and the maintenance of the cellular energy balance and the stress resistance. However, several diseases have been related with an AMPK dysfunction. Alterations in AMPK signaling decrease mitochondrial biogenesis, increase cellular stress and induce inflammation, which are typical events of the aging process and have been associated to several pathological processes. In this sense, in the last few years AMPK has been identified as a very interesting target and different nutraceutical compounds are being studied for an interesting potential effect on AMPK induction. In this review, we will evaluate the interaction of the different nutraceutical compounds to induce the AMPK phosphorylation and the applications in diseases such as cancer, type II diabetes, neurodegenerative diseases or cardiovascular diseases. PMID:28146060

  9. Different mechanisms of extracellular adenosine accumulation by reduction of the external Ca(2+) concentration and inhibition of adenosine metabolism in spinal astrocytes.

    PubMed

    Eguchi, Ryota; Akao, Sanae; Otsuguro, Ken-ichi; Yamaguchi, Soichiro; Ito, Shigeo

    2015-05-01

    Extracellular adenosine is a neuromodulator in the central nervous system. Astrocytes mainly participate in adenosine production, and extracellular adenosine accumulates under physiological and pathophysiological conditions. Inhibition of intracellular adenosine metabolism and reduction of the external Ca(2+) concentration ([Ca(2+)]e) participate in adenosine accumulation, but the precise mechanisms remain unclear. This study investigated the mechanisms underlying extracellular adenosine accumulation in cultured rat spinal astrocytes. The combination of adenosine kinase and deaminase (ADK/ADA) inhibition and a reduced [Ca(2+)]e increased the extracellular adenosine level. ADK/ADA inhibitors increased the level of extracellular adenosine but not of adenine nucleotides, which was suppressed by inhibition of equilibrative nucleoside transporter (ENT) 2. Unlike ADK/ADA inhibition, a reduced [Ca(2+)]e increased the extracellular level not only of adenosine but also of ATP. This adenosine increase was enhanced by ENT2 inhibition, and suppressed by sodium polyoxotungstate (ecto-nucleoside triphosphate diphosphohydrolase inhibitor). Gap junction inhibitors suppressed the increases in adenosine and adenine nucleotide levels by reduction of [Ca(2+)]e. These results indicate that extracellular adenosine accumulation by ADK/ADA inhibition is due to the adenosine release via ENT2, while that by reduction of [Ca(2+)]e is due to breakdown of ATP released via gap junction hemichannels, after which ENT2 incorporates adenosine into the cells.

  10. Adenosine A₂A-receptor antagonist istradefylline enhances the motor response of L-DOPA without worsening dyskinesia in MPTP-treated common marmosets.

    PubMed

    Uchida, Shin-ichi; Tashiro, Tomomi; Kawai-Uchida, Mika; Mori, Akihisa; Jenner, Peter; Kanda, Tomoyuki

    2014-01-01

    The adenosine A₂A-receptor antagonist istradefylline decreases OFF time in patients with Parkinson's disease who are already treated with optimal doses of dopaminergic medication but can cause an increase in non-troublesome dyskinesia. Preclinical experiments have shown that A₂A antagonists are most effective in potentiating motor function when combined with sub-maximal doses of L-DOPA. However, the effects of combining istradefylline with sub-optimal L-DOPA treatment on established dyskinesia have not been studied. We now examine the effects of acute and repeated administration of istradefylline on dyskinesia in MPTP-treated common marmosets previously primed to exhibit involuntary movements by prior exposure to L-DOPA. In these animals, single dose acute oral administration of istradefylline (10 mg/kg) enhanced and prolonged the anti-parkinsonian effects of a sub-optimal dose of L-DOPA (2.5 mg/kg). The chronic co-administration of istradefylline (10 mg/kg) with L-DOPA (2.5 mg/kg) for 21 days did not worsen the severity of existing dyskinesia. Rather, the severity of dyskinesia tended to be reduced over the 21-day treatment period. These results suggest that istradefylline can be used to potentiate the effects of sub-optimal doses of L-DOPA in the treatment of Parkinson's disease without causing or worsening dyskinesia.

  11. Histamine H3 Receptor Activation Counteracts Adenosine A2A Receptor-Mediated Enhancement of Depolarization-Evoked [3H]-GABA Release from Rat Globus Pallidus Synaptosomes

    PubMed Central

    2014-01-01

    High levels of histamine H3 receptors (H3Rs) are found in the globus pallidus (GP), a neuronal nucleus in the basal ganglia involved in the control of motor behavior. By using rat GP isolated nerve terminals (synaptosomes), we studied whether H3R activation modified the previously reported enhancing action of adenosine A2A receptor (A2AR) stimulation on depolarization-evoked [3H]-GABA release. At 3 and 10 nM, the A2AR agonist CGS-21680 enhanced [3H]-GABA release induced by high K+ (20 mM) and the effect of 3 nM CGS-21680 was prevented by the A2AR antagonist ZM-241385 (100 nM). The presence of presynaptic H3Rs was confirmed by the specific binding of N-α-[methyl-3H]-histamine to membranes from GP synaptosomes (maximum binding, Bmax, 1327 ± 79 fmol/mg protein; dissociation constant, Kd, 0.74 nM), which was inhibited by the H3R ligands immepip, clobenpropit, and A-331440 (inhibition constants, Ki, 0.28, 8.53, and 316 nM, respectively). Perfusion of synaptosomes with the H3R agonist immepip (100 nM) had no effect on K+-evoked [3H]-GABA release, but inhibited the stimulatory action of A2AR activation. In turn, the effect of immepip was blocked by the H3R antagonist clobenpropit, which had no significant effect of its own on K+-induced [3H]-GABA release. These data indicate that H3R activation selectively counteracts the facilitatory action of A2AR stimulation on GABA release from striato-pallidal projections. PMID:24884070

  12. Activation of the A2B adenosine receptor in B16 melanomas induces CXCL12 expression in FAP-positive tumor stromal cells, enhancing tumor progression

    PubMed Central

    Sorrentino, Claudia; Miele, Lucio; Porta, Amalia; Pinto, Aldo; Morello, Silvana

    2016-01-01

    The A2B receptor (A2BR) can mediate adenosine-induced tumor proliferation, immunosuppression and angiogenesis. Targeting the A2BR has proved to be therapeutically effective in some murine tumor models, but the mechanisms of these effects are still incompletely understood. Here, we report that pharmacologic inhibition of A2BR with PSB1115, which inhibits tumor growth, decreased the number of fibroblast activation protein (FAP)-expressing cells in tumors in a mouse model of melanoma. This effect was associated with reduced expression of fibroblast growth factor (FGF)-2. Treatment of melanoma-associated fibroblasts with the A2BR agonist Bay60-6583 enhanced CXCL12 and FGF2 expression. This effect was abrogated by PSB1115. The A2AR agonist CGS21680 did not induce CXCL12 or FGF2 expression in tumor associated fibroblasts. Similar results were obtained under hypoxic conditions in skin-derived fibroblasts, which responded to Bay60-6583 in an A2BR-dependent manner, by stimulating pERK1/2. FGF2 produced by Bay60-6583-treated fibroblasts directly enhanced the proliferation of melanoma cells. This effect could be reversed by PSB1115 or an anti-FGF2 antibody. Interestingly, melanoma growth in mice receiving Bay60-6583 was attenuated by inhibition of the CXCL12/CXCR4 pathway with AMD3100. CXCL12 and its receptor CXCR4 are involved in angiogenesis and immune-suppression. Treatment of mice with AMD3100 reduced the number of CD31+ cells induced by Bay60-6583. Conversely, CXCR4 blockade did not affect the accumulation of tumor-infiltrating MDSCs or Tregs. Together, our data reveal an important role for A2BR in stimulating FGF2 and CXCL12 expression in melanoma-associated fibroblasts. These factors contribute to create a tumor-promoting microenvironment. Our findings support the therapeutic potential of PSB1115 for melanoma. PMID:27590504

  13. Influence of ischemic injury on vein graft remodeling: role of cyclic adenosine monophosphate second messenger pathway in enhanced vein graft preservation.

    PubMed

    Sakaguchi, Taichi; Asai, Tomohiro; Belov, Dmitri; Okada, Morihito; Pinsky, David J; Schmidt, Ann Marie; Naka, Yoshifumi

    2005-01-01

    Endothelial injury during the harvest of saphenous vein grafts might play an important role in the development of vein graft disease after coronary artery bypass grafting. Using a murine autologous arterialized vein patch model, we tested whether the initial ischemic insult of vein grafts was linked to the later development of graft neointimal hyperplasia and whether the restoration of the cyclic adenosine monophosphate second messenger pathway would attenuate the development of neointimal hyperplasia. A segment of the external jugular vein of a mouse was grafted onto its abdominal aorta. Three weeks after the operation, the degree of neointimal hyperplasia of the implanted graft was compared among (1) grafts without preservation, (2) grafts with 2 hours of preservation (25 degrees C) in heparinized saline, and (3) grafts with 2 hours of preservation in heparinized saline in the presence of a cyclic adenosine monophosphate analog. In addition, cyclic adenosine monophosphate contents of vein grafts and leukocyte adherence to the graft endothelium were assessed. Cyclic adenosine monophosphate contents were significantly decreased after 2 hours of preservation (212 +/- 8 vs 156 +/- 5 pmol/L, P < .01). The grafts preserved for 2 hours showed greater neointimal hyperplasia compared with the grafts without preservation (neointimal expansion, 68.7% +/- 9.6% vs 46.1% +/- 4.8%; P < .01). The addition of a cyclic adenosine monophosphate analog to the preservation solution significantly suppressed neointimal hyperplasia of grafts preserved for 2 hours (44.3% +/- 5.0%). Inhibiting the cyclic adenosine monophosphate-dependent protein kinase by adding Rp-cAMPS abrogated the beneficial effects. Furthermore, grafts preserved for 2 hours had significantly more leukocytes adhering to the graft endothelium 24 hours after the operation compared with nonpreserved grafts, which was significantly reduced by the cyclic adenosine monophosphate treatment. Ischemic insult during vein graft

  14. Supplementation of Exogenous Adenosine 5′-Triphosphate Enhances Mechanical Properties of 3D Cell–Agarose Constructs for Cartilage Tissue Engineering

    PubMed Central

    Gadjanski, Ivana; Yodmuang, Supansa; Spiller, Kara; Bhumiratana, Sarindr

    2013-01-01

    Formation of tissue-engineered cartilage is greatly enhanced by mechanical stimulation. However, direct mechanical stimulation is not always a suitable method, and the utilization of mechanisms underlying mechanotransduction might allow for a highly effective and less aggressive alternate means of stimulation. In particular, the purinergic, adenosine 5′-triphosphate (ATP)-mediated signaling pathway is strongly implicated in mechanotransduction within the articular cartilage. We investigated the effects of transient and continuous exogenous ATP supplementation on mechanical properties of cartilaginous constructs engineered using bovine chondrocytes and human mesenchymal stem cells (hMSCs) encapsulated in an agarose hydrogel. For both cell types, we have observed significant increases in equilibrium and dynamic compressive moduli after transient ATP treatment applied in the fourth week of cultivation. Continuous ATP treatment over 4 weeks of culture only slightly improved the mechanical properties of the constructs, without major changes in the total glycosaminoglycan (GAG) and collagen content. Structure–function analyses showed that transiently ATP-treated constructs, and in particular those based on hMSCs, had the highest level of correlation between compositional and mechanical properties. Transiently treated groups showed intense staining of the territorial matrix for GAGs and collagen type II. These results indicate that transient ATP treatment can improve functional mechanical properties of cartilaginous constructs based on chondrogenic cells and agarose hydrogels, possibly by improving the structural organization of the bulk phase and territorial extracellular matrix (ECM), that is, by increasing correlation slopes between the content of the ECM components (GAG, collagen) and mechanical properties of the construct. PMID:23651296

  15. Chromium Propionate Enhances Adipogenic Differentiation of Bovine Intramuscular Adipocytes

    PubMed Central

    Tokach, Rebecca J.; Ribeiro, Flavio R. B.; Chung, Ki Yong; Rounds, Whitney; Johnson, Bradley J.

    2015-01-01

    In vitro experiments were performed to determine the effects of increasing concentrations of chromium propionate (CrPro) on mRNA and protein abundance of different enzymes and receptors. Intramuscular (IM) and subcutaneous (SC) preadipocytes and bovine satellite cells were isolated from the longissimus muscle to determine the effect of treatment on glucose transporter type 4 (GLUT4) and peroxisome proliferator-activated receptor γ mRNA and GLUT4 protein abundance. Preadipocyte cultures were treated with differentiation media plus either sodium propionate or different concentrations of CrPro for 96, 120, and 144 h before harvest. This study indicated that adipogenesis of the bovine IM adipocytes were more sensitive to the treatment of CrPro as compared to SC adipocytes. Enhancement of adenosine monophosphate-activated protein kinase and GLUT4 mRNA by CrPro treatment may enhance glucose uptake in IM adipocytes. CrPro decreased GLUT4 protein levels in muscle cell cultures suggesting that those cells have increased efficiency of glucose uptake due to exposure to increased levels of CrPro. In contrast, each of the two adipogenic lines had opposing responses to the CrPro. It appeared that CrPro had the most stimulative effect of GLUT4 response in the IM adipocytes as compared to SC adipocytes. These findings indicated opportunities to potentially augment marbling in beef cattle fed CrPro during the finishing phase. PMID:26664955

  16. Chromium Propionate Enhances Adipogenic Differentiation of Bovine Intramuscular Adipocytes.

    PubMed

    Tokach, Rebecca J; Ribeiro, Flavio R B; Chung, Ki Yong; Rounds, Whitney; Johnson, Bradley J

    2015-01-01

    In vitro experiments were performed to determine the effects of increasing concentrations of chromium propionate (CrPro) on mRNA and protein abundance of different enzymes and receptors. Intramuscular (IM) and subcutaneous (SC) preadipocytes and bovine satellite cells were isolated from the longissimus muscle to determine the effect of treatment on glucose transporter type 4 (GLUT4) and peroxisome proliferator-activated receptor γ mRNA and GLUT4 protein abundance. Preadipocyte cultures were treated with differentiation media plus either sodium propionate or different concentrations of CrPro for 96, 120, and 144 h before harvest. This study indicated that adipogenesis of the bovine IM adipocytes were more sensitive to the treatment of CrPro as compared to SC adipocytes. Enhancement of adenosine monophosphate-activated protein kinase and GLUT4 mRNA by CrPro treatment may enhance glucose uptake in IM adipocytes. CrPro decreased GLUT4 protein levels in muscle cell cultures suggesting that those cells have increased efficiency of glucose uptake due to exposure to increased levels of CrPro. In contrast, each of the two adipogenic lines had opposing responses to the CrPro. It appeared that CrPro had the most stimulative effect of GLUT4 response in the IM adipocytes as compared to SC adipocytes. These findings indicated opportunities to potentially augment marbling in beef cattle fed CrPro during the finishing phase.

  17. Effect of adenosine and adenosine analogues on cyclic AMP accumulation in cultured mesangial cells and isolated glomeruli of the rat.

    PubMed Central

    Olivera, A.; Lopez-Novoa, J. M.

    1992-01-01

    1. Changes in intracellular levels of adenosine 3':5'-cyclic monophosphate (cyclic AMP) were studied in rat isolated glomeruli and cultured glomerular mesangial cells exposed to adenosine and to the preferential A1 receptor agonist N6-R-1-methyl-2-phenylethyl adenosine (R-PIA), or the potent A2 adenosine receptor agonist 5-(N-ethylcarboxamide)adenosine (NECA). 2. Whereas NECA and adenosine triggered a dose-dependent increase in cyclic AMP values with EC50 values of approximately 10(-6) M and 3 x 10(-5) M respectively, R-PIA lowered cyclic AMP levels at concentrations of 10(-6) M or less and increased them at higher concentrations. 3. The time-course of the increase induced by 10(-6) M NECA was slower than that induced by 10(-4) M adenosine. Adenosine produced a maximal stimulation within the first minute, whereas the effect of NECA in both glomeruli and mesangial cells was noticeable only from the second minute of incubation. 4. The effects of the agonists R-PIA and NECA on the cyclic AMP system were blocked respectively by the A1 adenosine receptor antagonist, 8-cyclopentyl-1, 3-dipropylxanthihe (DPCPX) at 10(-6) M and the A2 antagonist N-(2-dimethylaminoethyl)-N-methyl-4-(2, 3, 6, 7-tetrahydro-2,b-dioxo-1, 3-dipropyl-1H-purin-8-yl) benzene sulphonamide (PD115,199) at 10(-6) M. Theophylline, a known antagonist of adenosine receptors, inhibited the action of adenosine on cyclic AMP in mesangial cells. Dipyridamole, an inhibitor of the uptake of adenosine by the cells, enhanced the response to adenosine.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1330173

  18. Dietary effects of adenosine monophosphate to enhance growth, digestibility, innate immune responses and stress resistance of juvenile red sea bream, Pagrus major.

    PubMed

    Hossain, Md Sakhawat; Koshio, Shunsuke; Ishikawa, Manabu; Yokoyama, Saichiro; Sony, Nadia Mahjabin

    2016-09-01

    Our study explored the dietary effects of adenosine monophosphate (AMP) to enhance growth, digestibility, innate immune responses and stress resistance of juvenile red sea bream. A semi-purified basal diet supplemented with 0% (Control), 0.1% (AMP-0.1), 0.2% (AMP-0.2), 0.4% (AMP-0.4) and 0.8% (AMP-0.8) purified AMP to formulate five experimental diets. Each diet was randomly allocated to triplicate groups of fish (mean initial weight 3.4 g) for 56 days. The results indicated that dietary AMP supplements tended to improve growth performances. One of the best ones was found in diet group AMP-0.2, followed by diet groups AMP-0.1, AMP-0.4 and AMP-0.8. The Apparent digestibility coefficients (dry matter, protein and lipid) also improved by AMP supplementation and the significantly highest dry matter digestibility was observed in diet group AMP-0.2. Fish fed diet groups AMP-0.2 and AMP-0.4 had significantly higher peroxidase and bactericidal activities than fish fed the control diet. Nitro-blue-tetrazolium (NBT) activity was found to be significantly (P < 0.05) greater in fish fed diet groups AMP-0.4 and AMP-0.8. Total serum protein, lysozyme activity and agglutination antibody titer were also increased (P > 0.05) by dietary supplementation. In contrast, catalase activity decreased with AMP supplementation. Moreover, the fish fed AMP supplemented diets had better improvement (P < 0.05) in body lipid contents, condition factor, hematocrit content and glutamyl oxaloacetic transaminase (GOT) level than the control group. Supplementation also improved both freshwater and oxidative stress resistances. Interestingly, the fish fed diet groups AMP-0.2 and AMP-0.4 showed the least oxidative stress condition. Finally it is concluded that, dietary AMP supplementation enhanced the growth, digestibility, immune response and stress resistance of red sea bream. The regression analysis revealed that a dietary AMP supplementation between 0.2 and 0.4% supported weight gain and

  19. Adenosine triphosphate in milk.

    PubMed

    Zulak, I M; Patton, S; Hammerstedt, R H

    1976-08-01

    Freshly secreted goat's milk contains a number of viable metabolic pathways including those for the synthesis of triglycerides and phospholipids. Toward understanding this matter, amounts of the fundamentally important energy substrate, adenosine triphosphate, in goat's milk were evaluated. Milk left in the goat udder overnight had less adenosine triphosphate (12.4 muM) than fresh secreted milk (37.6 muM). In similar experiments the skim milk derived from whole accumulating in the udder overnight was lower in adenosine triphosphate (14.2 muM) than skim milk from freshly secreted milk (26.0 muM). To determine changes in quantities of adenosine triphosphate after milking, milks were divided into two parts, one containing only milk and the other milk plus 2,4-dinitrophenol and sodium arsenate, inhibitors of oxidative and substrate level phosphorylation. Adenosine triphosphate in milk decreased during 4-h in vitro incubations, and the rate of decline was markedly greater in the presence of the inhibitors. Thus, freshly secreted goat's milk contains significant amounts of adenosine triphosphate, and more can be synthesized therein after removal from the udder. In limited samples human and bovine milks contained much lower concentrations of the compound than those in goat's milk.

  20. Temporal and mechanistic dissociation of ATP and adenosine release during ischaemia in the mammalian hippocampus1

    PubMed Central

    Frenguelli, Bruno G; Wigmore, Geoffrey; Llaudet, Enrique; Dale, Nicholas

    2007-01-01

    Abstract Adenosine is well known to be released during cerebral metabolic stress and is believed to be neuroprotective. ATP release under similar circumstances has been much less studied. We have now used biosensors to measure and compare in real time the release of ATP and adenosine during in vitro ischaemia in hippocampal slices. ATP release only occurred following the anoxic depolarisation, whereas adenosine release was apparent almost immediately after the onset of ischaemia. ATP release required extracellular Ca2+. By contrast adenosine release was enhanced by removal of extracellular Ca2+, whilst TTX had no effect on either ATP release or adenosine release. Blockade of ionotropic glutamate receptors substantially enhanced ATP release, but had only a modest effect on adenosine release. Carbenoxolone, an inhibitor of gap junction hemichannels, also greatly enhanced ischaemic ATP release, but had little effect on adenosine release. The ecto-ATPase inhibitor ARL 67156, whilst modestly enhancing the ATP signal detected during ischaemia, had no effect on adenosine release. Adenosine release during ischaemia was reduced by pre-treament with homosysteine thiolactone suggesting an intracellular origin. Adenosine transport inhibitors did not inhibit adenosine release, but instead they caused a twofold increase of release. Our data suggest that ATP and adenosine release during ischaemia are for the most part independent processes with distinct underlying mechanisms. These two purines will consequently confer temporally distinct influences on neuronal and glial function in the ischaemic brain. PMID:17459147

  1. Lungs donated after circulatory death and prolonged warm ischemia are transplanted successfully after enhanced ex vivo lung perfusion using adenosine A2B receptor antagonism.

    PubMed

    Charles, Eric J; Mehaffey, J Hunter; Sharma, Ashish K; Zhao, Yunge; Stoler, Mark H; Isbell, James M; Lau, Christine L; Tribble, Curtis G; Laubach, Victor E; Kron, Irving L

    2017-04-12

    The current supply of acceptable donor lungs is not sufficient for the number of patients awaiting transplantation. We hypothesized that ex vivo lung perfusion (EVLP) with targeted drug therapy would allow successful rehabilitation and transplantation of donation after circulatory death lungs exposed to 2 hours of warm ischemia. Donor porcine lungs were procured after 2 hours of warm ischemia postcardiac arrest and subjected to 4 hours of cold preservation or EVLP. ATL802, an adenosine A2B receptor antagonist, was administered to select groups. Four groups (n = 4/group) were randomized: cold preservation (Cold), cold preservation with ATL802 during reperfusion (Cold + ATL802), EVLP (EVLP), and EVLP with ATL802 during ex vivo perfusion (EVLP + ATL802). Lungs subsequently were transplanted, reperfused, and assessed by measuring dynamic lung compliance and oxygenation capacity. EVLP + ATL802 significantly improved dynamic lung compliance compared with EVLP (25.0 ± 1.8 vs 17.0 ± 2.4 mL/cmH2O, P = .04), and compared with cold preservation (Cold: 12.2 ± 1.3, P = .004; Cold + ATL802: 10.6 ± 2.0 mL/cmH2O, P = .002). Oxygenation capacity was highest in EVLP (440.4 ± 37.0 vs Cold: 174.0 ± 61.3 mm Hg, P = .037). No differences in oxygenation or pulmonary edema were observed between EVLP and EVLP + ATL802. A significant decrease in interleukin-12 expression in tissue and bronchoalveolar lavage was identified between groups EVLP and EVLP + ATL802, along with less neutrophil infiltration. Severely injured donation after circulatory death lungs subjected to 2 hours of warm ischemia are transplanted successfully after enhanced EVLP with targeted drug therapy. Increased use of lungs after uncontrolled donor cardiac death and prolonged warm ischemia may be possible and may improve transplant wait list times and mortality. Copyright © 2017 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.

  2. WITHDRAWN: Anti-adipogenic effects of centipede grass extract in 3T3-L1 adipocytes and high fat diet induced obesity mice through activating adenosine monophosphate activated protein kinase.

    PubMed

    Kim, Sokho; Oh, Myung-Hoon; Kwon, Jungkee

    2013-11-02

    This article has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy.

  3. Adenosine receptor neurobiology: overview.

    PubMed

    Chen, Jiang-Fan; Lee, Chien-fei; Chern, Yijuang

    2014-01-01

    Adenosine is a naturally occurring nucleoside that is distributed ubiquitously throughout the body as a metabolic intermediary. In the brain, adenosine functions as an important upstream neuromodulator of a broad spectrum of neurotransmitters, receptors, and signaling pathways. By acting through four G-protein-coupled receptors, adenosine contributes critically to homeostasis and neuromodulatory control of a variety of normal and abnormal brain functions, ranging from synaptic plasticity, to cognition, to sleep, to motor activity to neuroinflammation, and cell death. This review begun with an overview of the gene and genome structure and the expression pattern of adenosine receptors (ARs). We feature several new developments over the past decade in our understanding of AR functions in the brain, with special focus on the identification and characterization of canonical and noncanonical signaling pathways of ARs. We provide an update on functional insights from complementary genetic-knockout and pharmacological studies on the AR control of various brain functions. We also highlight several novel and recent developments of AR neurobiology, including (i) recent breakthrough in high resolution of three-dimension structure of adenosine A2A receptors (A2ARs) in several functional status, (ii) receptor-receptor heterodimerization, (iii) AR function in glial cells, and (iv) the druggability of AR. We concluded the review with the contention that these new developments extend and strengthen the support for A1 and A2ARs in brain as therapeutic targets for neurologic and psychiatric diseases. © 2014 Elsevier Inc. All rights reserved.

  4. The adenosine A2A receptor antagonist ZM241385 enhances neuronal survival after oxygen-glucose deprivation in rat CA1 hippocampal slices

    PubMed Central

    Pugliese, AM; Traini, C; Cipriani, S; Gianfriddo, M; Mello, T; Giovannini, MG; Galli, A; Pedata, F

    2009-01-01

    Background and purpose: Activation of adenosine A2A receptors in the CA1 region of rat hippocampal slices during oxygen-glucose deprivation (OGD), a model of cerebral ischaemia, was investigated. Experimental approach: We made extracellular recordings of CA1 field excitatory postsynaptic potentials (fepsps) followed by histochemical and immunohistochemical techniques coupled to Western blots. Key results: OGD (7 or 30 min duration) elicited an irreversible loss of fepsps invariably followed by the appearance of anoxic depolarization (AD), an unambiguous sign of neuronal damage. The application of the selective adenosine A2A receptor antagonist, ZM241385 (4-(2-[7-amino-2-{2-furyl}{1,2,4}triazolo{2,3-a}{1,3,5}triazin-5-ylamino]ethyl)phenol; 100–500 nmol·L−1) prevented or delayed AD appearance induced by 7 or 30 min OGD and protected from the irreversible fepsp depression elicited by 7 min OGD. Two different selective adenosine A2A receptor antagonists, SCH58261 and SCH442416, were less effective than ZM241385 during 7 min OGD. The extent of CA1 cell injury was assessed 3 h after the end of 7 min OGD by propidium iodide. Substantial CA1 pyramidal neuronal damage occurred in untreated slices, exposed to OGD, whereas injury was significantly prevented by 100 nmol·L−1 ZM241385. Glial fibrillary acid protein (GFAP) immunostaining showed that 3 h after 7 min OGD, astrogliosis was appreciable. Western blot analysis indicated an increase in GFAP 30 kDa fragment which was significantly reduced by treatment with 100 nmol·L−1 ZM241385. Conclusions and implications: In the CA1 hippocampus, antagonism of A2A adenosine receptors by ZM241385 was protective during OGD (a model of cerebral ischaemia) by delaying AD appearance, decreasing astrocyte activation and improving neuronal survival. PMID:19422385

  5. Methotrexate enhances the anti-inflammatory effect of CF101 via up-regulation of the A3 adenosine receptor expression

    PubMed Central

    Ochaion, Avivit; Bar-Yehuda, Sara; Cohn, Shira; Del Valle, Luis; Perez-Liz, Georginia; Madi, Lea; Barer, Faina; Farbstein, Motti; Fishman-Furman, Sari; Reitblat, Tatiana; Reitblat, Alexander; Amital, Howard; Levi, Yair; Molad, Yair; Mader, Reuven; Tishler, Moshe; Langevitz, Pnina; Zabutti, Alexander; Fishman, Pnina

    2006-01-01

    Methotrexate (MTX) exerts an anti-inflammatory effect via its metabolite adenosine, which activates adenosine receptors. The A3 adenosine receptor (A3AR) was found to be highly expressed in inflammatory tissues and peripheral blood mononuclear cells (PBMCs) of rats with adjuvant-induced arthritis (AIA). CF101 (IB-MECA), an A3AR agonist, was previously found to inhibit the clinical and pathological manifestations of AIA. The aim of the present study was to examine the effect of MTX on A3AR expression level and the efficacy of combined treatment with CF101 and MTX in AIA rats. AIA rats were treated with MTX, CF101, or both agents combined. A3AR mRNA, protein expression and exhibition were tested in paw and PBMC extracts from AIA rats utilizing immunohistochemistry staining, RT-PCR and Western blot analysis. A3AR level was tested in PBMC extracts from patients chronically treated with MTX and healthy individuals. The effect of CF101, MTX and combined treatment on A3AR expression level was also tested in PHA-stimulated PBMCs from healthy individuals and from MTX-treated patients with rheumatoid arthritis (RA). Combined treatment with CF101 and MTX resulted in an additive anti-inflammatory effect in AIA rats. MTX induced A2AAR and A3AR over-expression in paw cells from treated animals. Moreover, increased A3AR expression level was detected in PBMCs from MTX-treated RA patients compared with cells from healthy individuals. MTX also increased the protein expression level of PHA-stimulated PBMCs from healthy individuals. The increase in A3AR level was counteracted in vitro by adenosine deaminase and mimicked in vivo by dipyridamole, demonstrating that receptor over-expression was mediated by adenosine. In conclusion, the data presented here indicate that MTX induces increased A3AR expression and exhibition, thereby potentiating the inhibitory effect of CF101 and supporting combined use of these drugs to treat RA. PMID:17101059

  6. Adenosine signaling in normal and sickle erythrocytes and beyond

    PubMed Central

    Zhang, Yujin; Xia, Yang

    2012-01-01

    Sickle cell disease (SCD) is a debilitating hemolytic genetic disorder with high morbidity and mortality affecting millions of individuals worldwide. Although SCD was discovered more than a century ago, no effective mechanism-based prevention and treatment are available due to poorly understood molecular basis of sickling, the fundamental pathogenic process of the disease. SCD patients constantly face hypoxia. One of the best-known signaling molecules to be induced under hypoxic conditions is adenosine. Recent studies demonstrate that hypoxia-mediated elevated adenosine signaling plays an important role in normal erythrocyte physiology. In contrast, elevated adenosine signaling contributes to sickling and multiple life threatening complications including tissue damage, pulmonary dysfunction and priapism. Here, we summarize recent research on the role of adenosine signaling in normal and sickle erythrocytes, progression of the disease and therapeutic implications. In normal erythrocytes, both genetic and pharmacological studies demonstrate that adenosine can enhance 2,3-bisphosphoglycerate (2,3-BPG) production via A2B receptor (ADORA2B) activation, suggesting that elevated adenosine has an unrecognized role in normal erythrocytes to promote O2 release and prevent acute ischemic tissue injury. However, in sickle erythrocytes, the beneficial role of excessive adenosine-mediated 2,3-BPG induction becomes detrimental by promoting deoxygenation, polymerization of sickle hemoglobin and subsequent sickling. Additionally, adenosine signaling via the A2A receptor (ADORA2A) on invariant natural killer T (iNKT) cells inhibits iNKT cell activation and attenuates pulmonary dysfunction in SCD mice. Finally, elevated adenosine coupled with ADORA2BR activation is responsible for priapism, a dangerous complication seen in SCD. Overall, the research reviewed here reveals a differential role of elevated adenosine in normal erythrocytes, sickle erythrocytes, iNK cells and progression

  7. Adenosine and sleep

    SciTech Connect

    Yanik, G.M. Jr.

    1987-01-01

    Behavioral and biochemical approaches have been used to determine the relative contribution of endogenous adenosine and adenosine receptors to the sleep-wake cycle in the rat. Adenosine concentrations in specific areas of the rat brain were not affected by 24 hours of total sleep deprivation, or by 24 or 48 hours of REM sleep deprivation. In order to assess the effect of REM sleep deprivation on adenosine A/sub 1/ receptors, /sup 3/H-L-PIA binding was measured. The Bmax values for /sup 3/H-L-PIA binding to membrane preparations of the cortices and corpus striata from 48 hour REM sleep-deprived animals were increased 14.8% and 23%, respectively. These increases were not maintained following the cessation of sleep deprivation and recovered within 2 hours. The results of a 96 hour REM deprivation experiment were similar to those of the 48 hour REM sleep deprivation experiment. However, these increases were not evident in similar structures taken from stress control animals, and conclusively demonstrated that the changes in /sup 3/H-L-PIA binding resulted from REM sleep deprivation and not from stress.

  8. Adenosine reduces postbypass transfusion requirements in humans after heart surgery.

    PubMed Central

    Mentzer, R M; Rahko, P S; Canver, C C; Chopra, P S; Love, R B; Cook, T D; Hegge, M O; Lasley, R D

    1996-01-01

    OBJECTIVE: The objective of this study was to determine the effect, if any, of adenosine blood cardioplegia on blood component usage after heart surgery. SUMMARY BACKGROUND DATA: The most common cause of nonsurgical postcardiopulmonary bypass bleeding is platelet dysfunction. For this reason, pharmacologic agents are under investigation in an effort to reduce the need for transfusion in this setting. METHODS: A posthoc analysis of blood product usage was performed in data obtained from a Phase I, single center, open label, randomized study performed in 63 patients. The trial was designed to test the safety and tolerance of adenosine when added to blood cardioplegia in increasing doses to enhance myocardial protection. The database provided information regarding the effect of adenosine cardioplegia on venous plasma adenosine concentrations, the amount of platelets, fresh frozen plasma and packed erythrocytes used, and the association between the adenosine dose and postoperative thoracic drainage. RESULTS: The postoperative thoracic drainage at 6 hours, 24 hours, and at the time of chest tube removal in the high-dose adenosine cardioplegia group was 68%, 76%, and 75% of the placebo and low-dose adenosine cardioplegia group (p < 0.05). The highest dose of adenosine studied increased baseline adenosine venous plasma levels 360-fold, from 0.17 +/- 0.09 mumol/L to 42.30 +/- 11.20 mumol/L (p < 0.05). This marked increase was associated with a 68%, 56%, and 58% reduction in platelet, fresh frozen plasma, and packed erythrocyte usage, respectively (p < 0.05). CONCLUSIONS: In addition to enhancing the heart's tolerance to ischemia, adenosine-supplemented cardioplegic solution also may reduce bleeding after cardiopulmonary bypass. PMID:8857856

  9. Tween 20-stabilized gold nanoparticles combined with adenosine triphosphate-BODIPY conjugates for the fluorescence detection of adenosine with more than 1000-fold selectivity.

    PubMed

    Hung, Szu-Ying; Shih, Ya-Chen; Tseng, Wei-Lung

    2015-02-01

    This study describes the development of a simple, enzyme-free, label-free, sensitive, and selective system for detecting adenosine based on the use of Tween 20-stabilized gold nanoparticles (Tween 20-AuNPs) as an efficient fluorescence quencher for boron dipyrromethene-conjugated adenosine 5'-triphosphate (BODIPY-ATP) and as a recognition element for adenosine. BODIPY-ATP can interact with Tween 20-AuNPs through the coordination between the adenine group of BODIPY-ATP and Au atoms on the NP surface, thereby causing the fluorescence quenching of BODIPY-ATP through the nanometal surface energy transfer (NSET) effect. When adenosine attaches to the NP surface, the attached adenosine exhibits additional electrostatic attraction to BODIPY-ATP. As a result, the presence of adenosine enhances the efficiency of AuNPs in fluorescence quenching of BODIPY-ATP. The AuNP-induced fluorescence quenching of BODIPY-ATP progressively increased with an increase in the concentration of adenosine; the detection limit at a signal-to-noise ratio of 3 for adenosine was determined to be 60nM. The selectivity of the proposed system was more than 1000-fold for adenosine over any adenosine analogs and other nucleotides. The proposed system combined with a phenylboronic acid-containing column was successfully applied to the determination of adenosine in urine.

  10. Rat cardiac myocyte adenosine transport and metabolism

    SciTech Connect

    Ford, D.A.; Rovetto, M.J.

    1987-01-01

    Based on the importance of myocardial adenosine and adenine nucleotide metabolism, the adenosine salvage pathway in ventricular myocytes was studied. Accurate estimates of transport rates, separate from metabolic fllux, were determined. Adenosine influx was constant between 3 and 60 s. Adenosine metabolism maintained intracellular adenosine concentrations < 10% of the extracellular adenosine concentrations and thus unidirectional influx could be measured. Myocytes transported adenosine via saturable and nonsaturable processes. A minimum estimate of the V/sub max/ of myocytic adenosine kinase indicated the saturable component of adenosine influx was independent of adenosine kinase activity. Saturable transport was inhibited by nitrobenzylthioinosine and verapamil. Extracellular adenosine taken up myocytes was rapidly phosphorylated to adenine taken up by myocytes was rapidly phosphorylated to adenine nucleotides. Not all extracellular adenosine, though, was phosphorylated on entering myocytes, since free, as opposed to protein-bound, intracellular adenosine was detected after digitonin extraction of cells in the presence of 1 mM ethylene-diaminetetraacetic acid.

  11. Loss of Liver Kinase B1 (LKB1) in Beta Cells Enhances Glucose-stimulated Insulin Secretion Despite Profound Mitochondrial Defects.

    PubMed

    Swisa, Avital; Granot, Zvi; Tamarina, Natalia; Sayers, Sophie; Bardeesy, Nabeel; Philipson, Louis; Hodson, David J; Wikstrom, Jakob D; Rutter, Guy A; Leibowitz, Gil; Glaser, Benjamin; Dor, Yuval

    2015-08-21

    The tumor suppressor liver kinase B1 (LKB1) is an important regulator of pancreatic β cell biology. LKB1-dependent phosphorylation of distinct AMPK (adenosine monophosphate-activated protein kinase) family members determines proper β cell polarity and restricts β cell size, total β cell mass, and glucose-stimulated insulin secretion (GSIS). However, the full spectrum of LKB1 effects and the mechanisms involved in the secretory phenotype remain incompletely understood. We report here that in the absence of LKB1 in β cells, GSIS is dramatically and persistently improved. The enhancement is seen both in vivo and in vitro and cannot be explained by altered cell polarity, increased β cell number, or increased insulin content. Increased secretion does require membrane depolarization and calcium influx but appears to rely mostly on a distal step in the secretion pathway. Surprisingly, enhanced GSIS is seen despite profound defects in mitochondrial structure and function in LKB1-deficient β cells, expected to greatly diminish insulin secretion via the classic triggering pathway. Thus LKB1 is essential for mitochondrial homeostasis in β cells and in parallel is a powerful negative regulator of insulin secretion. This study shows that β cells can be manipulated to enhance GSIS to supra-normal levels even in the face of defective mitochondria and without deterioration over months.

  12. Intracerebral adenosine infusion improves neurological outcome after transient focal ischemia in rats.

    PubMed

    Kitagawa, Hisashi; Mori, Atsushi; Shimada, Jun; Mitsumoto, Yasuhide; Kikuchi, Tetsuro

    2002-04-01

    Second Institute of New Drug Research, Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan In order to elucidate the role of adenosine in brain ischemia, the possible protective effects of adenosine on ischemic brain injury were investigated in a rat model of brain ischemia both in vitro and in vivo. Exogenous adenosine dose-dependently rescued cortical neuronal cells from injury after glucose deprivation in vitro. Adenosine (1 mM) also significantly reduced hypoglycemia/hypoxia-induced glutamate release from the hippocampal slice. In a rat model of transient middle cerebral artery occlusion (MCAO), extracellular adenosine concentration was increased immediately after occlusion, and then returned to the baseline by 30 min after reperfusion. Adenosine infusion through a microdialysis probe into the ipsilateral striatum (1 mM adenosine, 2 microl min(-1), total 4.5 h from the occlusion to 3 h after reperfusion) showed a significant improvement in the neurological outcome, and about 25% reduction of infarct volume, although the effect did not reach statistical significance, compared with the vehicle-treated group at 20 h after 90 min of MCAO. These results demonstrated the neuroprotective effect of adenosine against ischemic brain injury both in vitro and in vivo, suggesting the possible therapeutic application of adenosine regulating agents, which inhibit adenosine uptake or metabolism to enhance or maintain extracellular endogenous adenosine levels, for stroke treatment.

  13. Possible mechanism of adenosine protection in carbon tetrachloride acute hepatotoxicity. Role of adenosine by-products and glutathione peroxidase.

    PubMed

    Chagoya de Sánchez, V; Hernández-Muñoz, R; Yáñez, L; Vidrio, S; Díaz-Muñoz, M

    1995-02-01

    Adenosine proved to be an effective hepatoprotector increasing the survival rate of rats receiving lethal doses of CCl4. Searching for the mechanism of action, we found that adenosine transiently prevents the necrotic liver damage associated to an acute CCl4 treatment. The antilipoperoxidative action of the nucleoside was evidenced by a decrease of TBA-reactive products and the diene conjugates elicited by the hepatotoxin. Adenosine's protective effect was demonstrated by reverting the decrease of cytochrome P-450 while preserved intact the activity of the microsomal enzyme glucose-6-phosphatase. CCl4 promoted an increase in the oxidant stress through an enhancement in oxidized glutathione levels. This action was also completely counteracted by the nucleoside. Adenosine was unable to prevent CCl4 activation and, even, increased .CCl3 formation in the presence of PBN in vivo. However, in the presence of the nucleoside, irreversible binding of 14CCl4 to the microsomal lipid fraction of the treated animals was decreased. These results suggest that adenosine protective action might be exerted at the level of the propagation reaction following CCl4 activation. Two possible mechanisms were associated to the nucleoside protection: (1) the peroxide-metabolyzed enzymes, GSH-per, showed a marked increase after 30 minutes of adenosine treatment, which was potentiated by the hepatotoxin, suggesting an important role of this enzyme in the nucleoside's action; (2) the adenosine catabolism induced an increase in uric acid level, and allopurinol, a purine metabolism inhibitor, prevented such elevation as well as the antilipoperoxidative action of adenosine and the increase of GSH-per associated with the nucleoside treatment. These facts strongly suggest that the protective effect elicited by adenosine is not a direct one, but rather is related to its catabolic products, such as uric acid, which has been recognized as a free radical scavenger.

  14. Adenosine thallium 201 myocardial perfusion scintigraphy

    SciTech Connect

    Verani, M.S. )

    1991-07-01

    Pharmacologic coronary vasodilation as an adjunct to myocardial perfusion imaging has become increasingly important in the evaluation of patients with coronary artery disease, in view of the large number of patients who cannot perform an adequate exercise test or in whom contraindications render exercise inappropriate. Adenosine is a very potent coronary vasodilator and when combined with thallium 201 scintigraphy produces images of high quality, with the added advantages of a very short half-life (less than 10 seconds) and the ability to adjust the dose during the infusion, which may enhance safety and curtail the duration of side effects. The reported sensitivity and specificity of adenosine thallium 201 scintigraphy for the detection of coronary artery disease are high and at least comparable with imaging after exercise or dipyridamole administration. 23 refs.

  15. Feedback Effects of Host-Derived Adenosine on Enteropathogenic Escherichia coli

    PubMed Central

    Crane, John K.; Shulgina, Irina

    2009-01-01

    Enteropathogenic Escherichia coli (EPEC) is a common cause of watery diarrhea in children in developing countries. After adhering intimately to small intestinal cells, EPEC secretes effector proteins into host cells, altering host cell functions and causing cell damage and death. We previously showed that EPEC infection triggers the release of adenosine triphosphate (ATP) from host cells and that ATP is broken down to ADP, AMP, and adenosine. Adenosine produced from the breakdown of extracellular ATP triggers fluid secretion in cultured intestinal monolayers and may be an important mediator of EPEC-induced diarrhea. In this study we examined whether adenosine has any effects on EPEC bacteria themselves. Adenosine stimulated EPEC growth in several types of media in vitro. Adenosine also altered the pattern of EPEC adherence to cultured cells from a classic localized adherence pattern to a more diffuse adherence pattern. Adenosine changed the pattern of expression of virulence factors in EPEC, inhibiting the expression of the bundle-forming pilus (BFP) and enhancing expression of the EPEC secreted proteins (Esps). The ability of adenosine to inhibit BFP was dependent on the Plasmid-encoded Regulator (Per). In vivo, experimental manipulations of adenosine levels had strong effects on the outcome of EPEC infection in rabbit intestinal loops. Reduction in adenosine levels by addition of exogenous adenosine deaminase (ADA) reduced numbers of EPEC bacteria recovered by over 10-fold in rabbit intestine in vivo. Conversely, inhibitors of ADA increased EPEC-induced fluid secretion, the number of EPEC bacteria recovered from intestinal fluid, and increased the in vivo expression of espA and espB. In addition to its previously reported effects on host cells and tissues, adenosine also has strong effects on EPEC bacteria, stimulating EPEC growth, altering its adherence pattern, and changing the expression of several important virulence genes. Adenosine is released from host

  16. trans-Cinnamaldehyde stimulates mitochondrial biogenesis through PGC-1α and PPARβ/δ leading to enhanced GLUT4 expression.

    PubMed

    Gannon, Nicholas P; Schnuck, Jamie K; Mermier, Christine M; Conn, Carole A; Vaughan, Roger A

    2015-12-01

    Type 2 diabetes is characterized by insulin resistance and chronic hyperglycemia, and is increasing in incidence and severity. This work explored the effects of trans-cinnamaldehyde (CA) on carbohydrate metabolism, mitochondrial content, and related metabolic gene and protein expression in cultured myotubes treated with various concentrations of CA for up to 24 h. CA treatment increased myotube myocyte enhancer factor 2 (MEF2) along with glucose transporter 4 (GLUT4) content. CA treatment also significantly increased expression of markers of improved oxidative metabolism including 5' adenosine monophosphate-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ coactivator 1 α (PGC-1α), cytochrome c (CytC), as well as peroxisome proliferator-activated receptor α (PPARα) and PPARβ/δ. Despite increased expression of proteins associated with improved oxidative metabolism and glucose uptake, CA-treated myotubes exhibited significantly reduced oxidative metabolism compared with controlled cells. Additionally, CA treatment increased markers of glucose-mediated lipid biosynthesis without elevated PPARγ and sterol receptor element binding protein 1c (SREBP-1c) expression. The ability of CA to stimulate mitochondrial biogenesis and GLUT4 expression suggests CA may offer possible benefits for metabolic disease. However, increases in markers of fatty acid synthesis with simultaneously reduced oxidative metabolism suggest CA may have counterproductive effects for metabolic disease, warranting a need for further investigation. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  17. Multidrug Resistance-Associated Protein 2 Expression Is Upregulated by Adenosine 5’-Triphosphate in Colorectal Cancer Cells and Enhances Their Survival to Chemotherapeutic Drugs

    PubMed Central

    Vinette, Valérie; Placet, Morgane; Arguin, Guillaume; Gendron, Fernand-Pierre

    2015-01-01

    Extracellular adenosine 5’-triphosphate (ATP) is a signaling molecule that induces a plethora of effects ranging from the regulation of cell proliferation to modulation of cancerous cell behavior. In colorectal cancer, ATP was reported to stimulate epithelial cell proliferation and possibly promote resistance to anti-cancer treatments. However, the exact role of this danger-signaling molecule on cancerous intestinal epithelial cells (IECs) in response to chemotherapeutic agents remains unknown. To address how ATP may influence the response of cancerous IECs to chemotherapeutic agents, we used Caco-2 cells, which display enterocyte-like features, to determine the effect of ATP on the expression of multidrug resistance-associated protein 2 (MRP2). Gene and protein expression were determined by quantitative real-time PCR (qRT-PCR) and Western blotting. Resistance to etoposide, cisplatin and doxorubicin was determined by MTT assays in response to ATP stimulation of Caco-2 cells and in cells for which MRP2 expression was down-regulated by shRNA. ATP increased the expression of MRP2 at both the mRNA and protein levels. MRP2 expression involved an ATP-dependent stimulation of the MEK/ERK signaling pathway that was associated with an increase in relative resistance of Caco-2 cells to etoposide. Abolition of MRP2 expression using shRNA significantly reduced the protective effect of MRP2 toward etoposide as well as to cisplatin and doxorubicin. This study describes the mechanism by which ATP may contribute to the chemoresistance of cancerous IECs in colorectal cancer. Given the heterogeneity of colorectal adenocarcinoma responses to anti-cancer drugs, these findings call for further study to understand the role of P2 receptors in cancer drug therapy and to develop novel therapies aimed at regulating P2 receptor activity. PMID:26295158

  18. E2F1 enhances 8-chloro-adenosine-induced G2/M arrest and apoptosis in A549 and H1299 lung cancer cells.

    PubMed

    Duan, Hong-Ying; Cao, Ji-Xiang; Qi, Jun-Juan; Wu, Guo-Sheng; Li, Shu-Yan; An, Guo-Shun; Jia, Hong-Ti; Cai, Wang-Wei; Ni, Ju-Hua

    2012-03-01

    The E2F1 transcription factor is a well known regulator of cell proliferation and apoptosis, but its role in response to DNA damage is less clear. 8-Chloro-adenosine (8-Cl-Ado), a nucleoside analog, can inhibit proliferation in a variety of human tumor cells. However, it is still elusive how the agent acts on tumors. Here we show that A549 and H1299 cells formed DNA double-strand breaks after 8-Cl-Ado exposure, accompanied by E2F1 upregulation at protein level. Overexpressed wild-type (E2F1-wt) colocalized with double-strand break marker γ-H2AX and promoted G2/M arrest in 8-Cl-Ado-exposed A549 and H1299, while expressed S31A mutant of E2F1 (E2F1-mu) significantly reduced ability to accumulate at sites of DNA damage and G2/M arrest, suggesting that E2F1 is required for activating G2/M checkpoint pathway upon DNA damage. Transfection of either E2F1-wt or E2F1-mu plasmid promoted apoptosis in 8-Cl-Ado-exposed cells, indicating that 8-Cl-Ado may induce apoptosis in E2F1-dependent and E2F1-independent ways. These findings demonstrate that E2F1 plays a crucial role in 8-Cl-Ado-induced G2/M arrest but is dispensable for 8-Cl-Ado-induced apoptosis. These data also suggest that the mechanism of 8-Cl-Ado action is complicated.

  19. Imaging Adenosine Triphosphate (ATP)

    PubMed Central

    Rajendran, Megha; Dane, Eric; Conley, Jason; Tantama, Mathew

    2016-01-01

    Adenosine triphosphate (ATP) is a universal mediator of metabolism and signaling across unicellular and multicellular species. There is a fundamental interdependence between the dynamics of ATP and the physiology that occurs inside and outside the cell. Characterizing and understanding ATP dynamics provides valuable mechanistic insight into processes that range from neurotransmission to the chemotaxis of immune cells. Therefore, we require the methodology to interrogate both temporal and spatial components of ATP dynamics from the subcellular to organismal levels in live specimens. Over the last several decades, a number of molecular probes that are specific for ATP have been developed. These probes have been combined with imaging approaches, particularly optical microscopy, to enable qualitative and quantitative detection of this critical molecule. In this review, we survey current examples of technologies that are available to visualize ATP in living cells and identify areas where new tools and approaches are needed to expand our capabilities. PMID:27638696

  20. Feed-Forward Inhibition of CD73 and Upregulation of Adenosine Deaminase Contribute to the Loss of Adenosine Neuromodulation in Postinflammatory Ileitis

    PubMed Central

    Magalhães-Cardoso, Maria Teresa; Ferreirinha, Fátima; Dias, Ana Sofia; Pelletier, Julie

    2014-01-01

    Purinergic signalling is remarkably plastic during gastrointestinal inflammation. Thus, selective drugs targeting the “purinome” may be helpful for inflammatory gastrointestinal diseases. The myenteric neuromuscular transmission of healthy individuals is fine-tuned and controlled by adenosine acting on A2A excitatory receptors. Here, we investigated the neuromodulatory role of adenosine in TNBS-inflamed longitudinal muscle-myenteric plexus of the rat ileum. Seven-day postinflammation ileitis lacks adenosine neuromodulation, which may contribute to acceleration of gastrointestinal transit. The loss of adenosine neuromodulation results from deficient accumulation of the nucleoside at the myenteric synapse despite the fact that the increases in ATP release were observed. Disparity between ATP outflow and adenosine deficit in postinflammatory ileitis is ascribed to feed-forward inhibition of ecto-5′-nucleotidase/CD73 by high extracellular ATP and/or ADP. Redistribution of NTPDase2, but not of NTPDase3, from ganglion cell bodies to myenteric nerve terminals leads to preferential ADP accumulation from released ATP, thus contributing to the prolonged inhibition of muscle-bound ecto-5′-nucleotidase/CD73 and to the delay of adenosine formation at the inflamed neuromuscular synapse. On the other hand, depression of endogenous adenosine accumulation may also occur due to enhancement of adenosine deaminase activity. Both membrane-bound and soluble forms of ecto-5′-nucleotidase/CD73 and adenosine deaminase were detected in the inflamed myenteric plexus. These findings provide novel therapeutic targets for inflammatory gut motility disorders. PMID:25210228

  1. Purine metabolism in adenosine deaminase deficiency.

    PubMed Central

    Mills, G C; Schmalstieg, F C; Trimmer, K B; Goldman, A S; Goldblum, R M

    1976-01-01

    Purine and pyrimidine metabolites were measured in erythrocytes, plasma, and urine of a 5-month-old infant with adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) deficiency. Adenosine and adenine were measured using newly devised ion exchange separation techniques and a sensitive fluorescence assay. Plasma adenosine levels were increased, whereas adenosine was normal in erythrocytes and not detectable in urine. Increased amounts of adenine were found in erythrocytes and urine as well as in the plasma. Erythrocyte adenosine 5'-monophosphate and adenosine diphosphate concentrations were normal, but adenosine triphosphate content was greatly elevated. Because of the possibility of pyrimidine starvation, pyrimidine nucleotides (pyrimidine coenzymes) in erythrocytes and orotic acid in urine were measured. Pyrimidine nucleotide concentrations were normal, while orotic acid was not detected. These studies suggest that the immune deficiency associated with adenosine deaminase deficiency may be related to increased amounts of adenine, adenosine, or adenine nucleotides. PMID:1066699

  2. Potentiation of the depression by adenosine of rat cerebral cortical neurones by progestational agents.

    PubMed Central

    Phillis, J. W.

    1986-01-01

    The effects of four progestational agents pregnenolone sulphate, cyproterone acetate, norethindrone acetate and progesterone, on adenosine-evoked depression of the firing of rat cerebral cortical neurones have been studied. When applied iontophoretically, pregnenolone sulphate, cyproterone, and norethindrone enhanced the actions of iontophoretically applied adenosine and failed to potentiate the depressant effects of adenosine 5'-N-ethylcarboxamide and gamma-aminobutyric acid. Cyproterone acetate (50 micrograms kg-1) and progesterone (200 micrograms kg-1) administered intravenously enhanced the depressant actions of iontophoretically applied adenosine. When applied by large currents, cyproterone, and less frequently norethindrone, depressed the firing of cerebral cortical neurones. The depressant effects of cyproterone were antagonized by caffeine. Pregnenolone sulphate tended to excite cortical neurones but neither this action, nor its potentiation of adenosine were reproduced by application of sulphate ions. It is hypothesized that some of the psychotropic actions of progestational agents may involve an enhancement of 'purinergic' tone in the central nervous system. PMID:3814905

  3. Supplementation of chitosan alleviates high-fat diet-enhanced lipogenesis in rats via adenosine monophosphate (AMP)-activated protein kinase activation and inhibition of lipogenesis-associated genes.

    PubMed

    Chiu, Chen-Yuan; Chan, Im-Lam; Yang, Tsung-Han; Liu, Shing-Hwa; Chiang, Meng-Tsan

    2015-03-25

    This study investigated the role of chitosan in lipogenesis in high-fat diet-induced obese rats. The lipogenesis-associated genes and their upstream regulatory proteins were explored. Diet supplementation of chitosan efficiently decreased the increased weights in body, livers, and adipose tissues in high-fat diet-fed rats. Chitosan supplementation significantly raised the lipolysis rate; attenuated the adipocyte hypertrophy, triglyceride accumulation, and lipoprotein lipase activity in epididymal adipose tissues; and decreased hepatic enzyme activities of lipid biosynthesis. Chitosan supplementation significantly activated adenosine monophosphate (AMP)-activated protein kinase (AMPK) phosphorylation and attenuated high-fat diet-induced protein expressions of lipogenic transcription factors (PPAR-γ and SREBP1c) in livers and adipose tissues. Moreover, chitosan supplementation significantly inhibited the expressions of downstream lipogenic genes (FAS, HMGCR, FATP1, and FABP4) in livers and adipose tissues of high-fat diet-fed rats. These results demonstrate for the first time that chitosan supplementation alleviates high-fat diet-enhanced lipogenesis in rats via AMPK activation and lipogenesis-associated gene inhibition.

  4. Enhancement of muscle glucose uptake by the vasopeptidase inhibitor, omapatrilat, is independent of insulin signaling and the AMP kinase pathway.

    PubMed

    Wong, Victor; Szeto, Linda; Uffelman, Kristine; Fantus, I George; Lewis, Gary F

    2006-08-01

    Omapatrilat (OMA), a vasopeptidase inhibitor (VPI), presently being tested in clinical trials for its antihypertensive properties, inhibits both angiotensin-converting enzyme and neutral endopeptidase, and raises tissue bradykinin levels. Recent studies from our laboratory and those of others have demonstrated that VPIs enhance muscle glucose uptake in animal models, and this effect is mediated by the bradykinin-nitric oxide pathway. The mechanism of the effect of OMA on muscle glucose uptake, however, is presently unknown. To investigate the effect of OMA on insulin signaling, soleus muscle was isolated 2 or 5 min after an i.v. bolus of insulin or saline from male Zucker fatty rats (8-10 weeks of age), following a 5-day treatment period of oral OMA (15 mg/kg per day) or drug vehicle (placebo). OMA resulted in significantly lower systolic blood pressure compared with the placebo-treated group (84.4+/- 7.52 mmHg in OMA vs 112+/-2.18 mmHg in controls, P<0.01). Immunoprecipitation and Western blot analysis of insulin receptor substrate 1 (IRS-1) revealed no changes in protein mass with OMA treatment. OMA did not enhance basal or insulin-stimulated IRS-1 tyrosine phosphorylation or its subsequent association with the p85 regulatory subunit of phosphatidylinositol 3-kinase. Under basal and insulin-stimulated conditions, OMA treatment did not alter the protein mass or the phosphorylation of Akt/protein kinase B, p42/44 extracellular signal-regulated kinase or adenosine monophosphate-activated protein kinase, or GLUT4 protein expression. We conclude that the ability of OMA to enhance whole body and specifically muscle glucose uptake in Zucker fatty rats is not mediated by enhancing insulin or AMPK signaling. Future studies should examine whether hemodynamic effects of the drug, independent of insulin signaling, enhance glucose uptake in insulin-resistant skeletal muscle.

  5. An Anti-Parkinson's Disease Drug via Targeting Adenosine A2A Receptor Enhances Amyloid-β Generation and γ-Secretase Activity.

    PubMed

    Lu, Jing; Cui, Jin; Li, Xiaohang; Wang, Xin; Zhou, Yue; Yang, Wenjuan; Chen, Ming; Zhao, Jian; Pei, Gang

    2016-01-01

    γ-secretase mediates the intramembranous proteolysis of amyloid precursor protein (APP) and determines the generation of Aβ which is associated with Alzheimer's disease (AD). Here we identified that an anti-Parkinson's disease drug, Istradefylline, could enhance Aβ generation in various cell lines and primary neuronal cells of APP/PS1 mouse. Moreover, the increased generation of Aβ42 was detected in the cortex of APP/PS1 mouse after chronic treatment with Istradefylline. Istradefylline promoted the activity of γ-secretase which could lead to increased Aβ production. These effects of Istradefylline were reduced by the knockdown of A2AR but independent of A2AR-mediated G protein- or β-arrestin-dependent signal pathway. We further observed that A2AR colocalized with γ-secretase in endosomes and physically interacted with the catalytic subunit presenilin-1 (PS1). Interestingly, Istradefylline attenuated the interaction in time- and dosage-dependent manners. Moreover the knockdown of A2AR which in theory would release PS1 potentiated both Aβ generation and γ-secretase activity. Thus, our study implies that the association of A2AR could modulate γ-secretase activity. Istradefylline enhance Aβ generation and γ-secretase activity possibly via modulating the interaction between A2AR and γ-secretase, which may bring some undesired effects in the central nervous system (CNS).

  6. An Anti-Parkinson’s Disease Drug via Targeting Adenosine A2A Receptor Enhances Amyloid-β Generation and γ-Secretase Activity

    PubMed Central

    Li, Xiaohang; Wang, Xin; Zhou, Yue; Yang, Wenjuan; Chen, Ming; Zhao, Jian; Pei, Gang

    2016-01-01

    γ-secretase mediates the intramembranous proteolysis of amyloid precursor protein (APP) and determines the generation of Aβ which is associated with Alzheimer’s disease (AD). Here we identified that an anti-Parkinson’s disease drug, Istradefylline, could enhance Aβ generation in various cell lines and primary neuronal cells of APP/PS1 mouse. Moreover, the increased generation of Aβ42 was detected in the cortex of APP/PS1 mouse after chronic treatment with Istradefylline. Istradefylline promoted the activity of γ-secretase which could lead to increased Aβ production. These effects of Istradefylline were reduced by the knockdown of A2AR but independent of A2AR-mediated G protein- or β-arrestin-dependent signal pathway. We further observed that A2AR colocalized with γ-secretase in endosomes and physically interacted with the catalytic subunit presenilin-1 (PS1). Interestingly, Istradefylline attenuated the interaction in time- and dosage-dependent manners. Moreover the knockdown of A2AR which in theory would release PS1 potentiated both Aβ generation and γ-secretase activity. Thus, our study implies that the association of A2AR could modulate γ-secretase activity. Istradefylline enhance Aβ generation and γ-secretase activity possibly via modulating the interaction between A2AR and γ-secretase, which may bring some undesired effects in the central nervous system (CNS). PMID:27835671

  7. Adenosine hypothesis of schizophrenia –opportunities for pharmacotherapy

    PubMed Central

    Boison, Detlev; Singer, Philipp; Shen, Hai-Ying; Feldon, Joram; Yee, Benjamin K.

    2011-01-01

    Pharmacotherapy of schizophrenia based on the dopamine hypothesis remains unsatisfactory for the negative and cognitive symptoms of the disease. Enhancing N-methyl-d-aspartate receptors (NMDAR) function is expected to alleviate such persistent symptoms, but successful development of novel clinically effective compounds remains challenging. Adenosine is a homeostatic bioenergetic network modulator that is able to affect complex networks synergistically at different levels (receptor dependent pathways, biochemistry, bioenergetics, and epigenetics). By affecting brain dopamine and glutamate activities it represents a promising candidate for restoring the functional imbalance in these neurotransmitter systems believed to underlie the genesis of schizophrenia symptoms, as well as restoring homeostasis of bioenergetics. Suggestion of an adenosine hypothesis of schizophrenia further posits that adenosinergic dysfunction might contribute to the emergence of multiple neurotransmitter dysfunctionscharacteristic of schizophrenia via diverse mechanisms. Given the importance of adenosine in early brain development and regulation of brain immune response, it also bears direct relevance to the aetiology of schizophrenia. Here, we provide an overview of the rationale and evidence in support of the therapeutic potential of multiple adenosinergic targets, including the high-affinity adenosine receptors (A1R and A2AR), and the regulatory enzyme adenosine kinase (ADK). Key preliminary clinical data and preclinical findings are reviewed. PMID:21315743

  8. Molecular implications of adenosine in obesity.

    PubMed

    Pardo, Fabián; Villalobos-Labra, Roberto; Chiarello, Delia I; Salsoso, Rocío; Toledo, Fernando; Gutierrez, Jaime; Leiva, Andrea; Sobrevia, Luis

    2017-06-01

    Adenosine has broad activities in organisms due to the existence of multiple receptors, the differential adenosine concentrations necessary to activate these receptors and the presence of proteins able to synthetize, degrade or transport this nucleoside. All adenosine receptors have been reported to be involved in glucose homeostasis, inflammation, adipogenesis, insulin resistance, and thermogenesis, indicating that adenosine could participate in the process of obesity. Since adenosine seems to be associated with several effects, it is plausible that adenosine participates in the initiation and development of obesity or may function to prevent it. Thus, the purpose of this review was to explore the involvement of adenosine in adipogenesis, insulin resistance and thermogenesis, with the aim of understanding how adenosine could be used to avoid, treat or improve the metabolic state of obesity. Treatment with specific agonists and/or antagonists of adenosine receptors could reverse the obesity state, since adenosine receptors normalizes several mechanisms involved in obesity, such as lipolysis, insulin sensitivity and thermogenesis. Furthermore, obesity is a preventable state, and the specific activation of adenosine receptors could aid in the prevention of obesity. Nevertheless, for the treatment of obesity and its consequences, more studies and therapeutic strategies in addition to adenosine are necessary. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Preparation and general properties of a soluble adenosine triphosphatase from mitochondria

    PubMed Central

    Selwyn, M. J.

    1967-01-01

    1. The purification of an adenosine triphosphatase present in aqueous extracts of acetone-dried ox-heart mitochondria is described. 2. No evidence was found for the presence of more than one protein having adenosine-triphosphatase activity in these extracts. 3. The enzyme is less stable at 0° than at 25° but is stabilized by glycerol. 4. The activity is dependent on the presence of Mg2+ or certain other bivalent metal cations. 5. The adenosine-triphosphatase activity of the Mg2+-activated enzyme is enhanced by 2,4-dinitrophenol. 6. The kinetics of Mg2+ activation indicate that the ATP–Mg2+ complex is the important substrate: free ATP and Mg2+ are inhibitory. 7. This preparation of mitochondrial adenosine triphosphatase has many properties in common with the adenosine triphosphatase coupling factor from mitochondria (Racker, 1961). PMID:4228455

  10. Role of adenosine in postprandial and reactive hyperemia in canine jejunum.

    PubMed

    Sawmiller, D R; Chou, C C

    1992-10-01

    The role of adenosine in postprandial jejunal hyperemia was investigated by determining the effect of placement of predigested food into the jejunal lumen on blood flow and oxygen consumption before and during intra-arterial infusion of dipyridamole (1.5 microM arterial concn) or adenosine deaminase (9 U/ml arterial concn) in anesthetized dogs. Neither drug significantly altered resting jejunal blood flow and oxygen consumption. Before dipyridamole or deaminase, food placement increased blood flow by 30-36%, 26-42%, and 21-46%, and oxygen consumption by 13-22%, 21-22%, and 26-29%, during 0- to 3-, 4- to 7-, and 8- to 11-min placement periods, respectively. Adenosine deaminase abolished the entire 11-min hyperemia, whereas dipyridamole significantly enhanced the initial 7-min hyperemia (45-49%). Both drugs abolished the initial 7-min food-induced increase in oxygen consumption. Dipyridamole attenuated (14%), whereas deaminase did not alter (28%), the increased oxygen consumption that occurred at 8-11 min. Adenosine deaminase also prevented the food-induced increase in venoarterial adenosine concentration difference. In separate series of experiments, luminal placement of food significantly increased jejunal lymphatic adenosine concentration and release. Also, reactive hyperemia was accompanied by an increase in venous adenosine concentration and release. This study provides further evidence to support the thesis that adenosine plays a role in postprandial and reactive hyperemia in the canine jejunum.

  11. Adenosine receptors and asthma in humans.

    PubMed

    Wilson, C N

    2008-10-01

    According to an executive summary of the GINA dissemination committee report, it is now estimated that approximately 300 million people (5% of the global population or 1 in 20 persons) have asthma. Despite the scientific progress made over the past several decades toward improving our understanding of the pathophysiology of asthma, there is still a great need for improved therapies, particularly oral therapies that enhance patient compliance and that target new mechanisms of action. Adenosine is an important signalling molecule in human asthma. By acting on extracellular G-protein-coupled ARs on a number of different cell types important in the pathophysiology of human asthma, adenosine affects bronchial reactivity, inflammation and airway remodelling. Four AR subtypes (A(1), A(2a), A(2b) and A(3)) have been cloned in humans, are expressed in the lung, and are all targets for drug development for human asthma. This review summarizes what is known about these AR subtypes and their function in human asthma as well as the pros and cons of therapeutic approaches to these AR targets. A number of molecules with high affinity and high selectivity for the human AR subtypes have entered clinical trials or are poised to enter clinical trials as anti-asthma treatments. With the availability of these molecules for testing in humans, the function of ARs in human asthma, as well as the safety and efficacy of approaches to the different AR targets, can now be determined.

  12. Intracoronary adenosine versus intravenous adenosine during primary PCI for ST-elevation myocardial infarction: which one offers better outcomes in terms of microvascular obstruction?

    PubMed

    Doolub, Gemina; Dall'armellina, Erica

    2013-01-01

    Aims. Previous studies have suggested that intravenous administration of adenosine improves myocardial reperfusion and reduces infarct size in ST-elevation myocardial infarction (STEMI) patients. Intracoronary administration of adenosine has shown conflicting results. Methods. In this retrospective, single-centre, blinded clinical study, we assessed whether selective intracoronary administration of adenosine distal to the occlusion site immediately before initial balloon inflation reduces microvascular obstruction (MVO) as assessed with cardiac magnetic resonance imaging (MRI). Using contrast-enhanced sequences, microvascular obstruction (MVO) was calculated. We found 81 patients presenting with STEMI within 12 h from symptom onset who were eligible for the study. In 80/81 (100%) patients receiving the study drug, MRI was performed on Day 1 after primary angioplasty. Results. The prevalence of MVO was reduced in the patients treated with intracoronary adenosine, (45%) compared to 85% of patients who were administered intravenous adenosine (P = 0.0043). We found that the size of MVO in patients receiving intracoronary adenosine was significantly reduced compared to 0.91 g in the intravenous-treated group (P = 0.027). There was no statistically significant difference in TIMI flow and clinical outcomes after primary PCI. Conclusion. We found significant evidence that selective high-dose intracoronary administration of adenosine distal to the occlusion site of the culprit lesion in STEMI patients results in a decrease in microvascular obstruction.

  13. Regulation of adenosine levels during cerebral ischemia

    PubMed Central

    Chu, Stephanie; Xiong, Wei; Zhang, Dali; Soylu, Hanifi; Sun, Chao; Albensi, Benedict C; Parkinson, Fiona E

    2013-01-01

    Adenosine is a neuromodulator with its level increasing up to 100-fold during ischemic events, and attenuates the excitotoxic neuronal injury. Adenosine is produced both intracellularly and extracellularly, and nucleoside transport proteins transfer adenosine across plasma membranes. Adenosine levels and receptor-mediated effects of adenosine are regulated by intracellular ATP consumption, cellular release of ATP, metabolism of extracellular ATP (and other adenine nucleotides), adenosine influx, adenosine efflux and adenosine metabolism. Recent studies have used genetically modified mice to investigate the relative contributions of intra- and extracellular pathways for adenosine formation. The importance of cortical or hippocampal neurons as a source or a sink of adenosine under basal and hypoxic/ischemic conditions was addressed through the use of transgenic mice expressing human equilibrative nucleoside transporter 1 (hENT1) under the control of a promoter for neuron-specific enolase. From these studies, we conclude that ATP consumption within neurons is the primary source of adenosine in neuronal cultures, but not in hippocampal slices or in vivo mice exposed to ischemic conditions. PMID:23064722

  14. K+ depolarization evokes ATP, adenosine and glutamate release from glia in rat hippocampus: a microelectrode biosensor study

    PubMed Central

    Heinrich, A; Andó, RD; Túri, G; Rózsa, B; Sperlágh, B

    2012-01-01

    BACKGROUND AND PURPOSE This study was undertaken to characterize the ATP, adenosine and glutamate outflow evoked by depolarization with high K+ concentrations, in slices of rat hippocampus. EXPERIMENTAL APPROACH We utilized the microelectrode biosensor technique and extracellular electrophysiological recording for the real-time monitoring of the efflux of ATP, adenosine and glutamate. KEY RESULTS ATP, adenosine and glutamate sensors exhibited transient and reversible current during depolarization with 25 mM K+, with distinct kinetics. The ecto-ATPase inhibitor ARL67156 enhanced the extracellular level of ATP and inhibited the prolonged adenosine efflux, suggesting that generation of adenosine may derive from the extracellular breakdown of ATP. Stimulation-evoked ATP, adenosine and glutamate efflux was inhibited by tetrodotoxin, while exposure to Ca2+-free medium abolished ATP and adenosine efflux from hippocampal slices. Extracellular elevation of ATP and adenosine were decreased in the presence of NMDA receptor antagonists, D-AP-5 and ifenprodil, whereas non-NMDA receptor blockade by CNQX inhibited glutamate but not ATP and adenosine efflux. The gliotoxin fluoroacetate and P2X7 receptor antagonists inhibited the K+-evoked ATP, adenosine and glutamate efflux, while carbenoxolone in low concentration and probenecid decreased only the adenosine efflux. CONCLUSIONS AND IMPLICATIONS Our results demonstrated activity-dependent gliotransmitter release in the hippocampus in response to ongoing neuronal activity. ATP and glutamate were released by P2X7 receptor activation into extracellular space. Although the increased extracellular levels of adenosine did derive from released ATP, adenosine might also be released directly via pannexin hemichannels. LINKED ARTICLE This article is commented on by Sershen, pp. 1000–1002 of this issue. To view this commentary visit http://dx.doi.org/10.1111/j.1476-5381.2012.02072.x PMID:22394324

  15. Sleep-wake sensitive mechanisms of adenosine release in the basal forebrain of rodents: an in vitro study.

    PubMed

    Sims, Robert Edward; Wu, Houdini Ho Tin; Dale, Nicholas

    2013-01-01

    Adenosine acting in the basal forebrain is a key mediator of sleep homeostasis. Extracellular adenosine concentrations increase during wakefulness, especially during prolonged wakefulness and lead to increased sleep pressure and subsequent rebound sleep. The release of endogenous adenosine during the sleep-wake cycle has mainly been studied in vivo with microdialysis techniques. The biochemical changes that accompany sleep-wake status may be preserved in vitro. We have therefore used adenosine-sensitive biosensors in slices of the basal forebrain (BFB) to study both depolarization-evoked adenosine release and the steady state adenosine tone in rats, mice and hamsters. Adenosine release was evoked by high K(+), AMPA, NMDA and mGlu receptor agonists, but not by other transmitters associated with wakefulness such as orexin, histamine or neurotensin. Evoked and basal adenosine release in the BFB in vitro exhibited three key features: the magnitude of each varied systematically with the diurnal time at which the animal was sacrificed; sleep deprivation prior to sacrifice greatly increased both evoked adenosine release and the basal tone; and the enhancement of evoked adenosine release and basal tone resulting from sleep deprivation was reversed by the inducible nitric oxide synthase (iNOS) inhibitor, 1400 W. These data indicate that characteristics of adenosine release recorded in the BFB in vitro reflect those that have been linked in vivo to the homeostatic control of sleep. Our results provide methodologically independent support for a key role for induction of iNOS as a trigger for enhanced adenosine release following sleep deprivation and suggest that this induction may constitute a biochemical memory of this state.

  16. Pathological overproduction: the bad side of adenosine.

    PubMed

    Borea, Pier Andrea; Gessi, Stefania; Merighi, Stefania; Vincenzi, Fabrizio; Varani, Katia

    2017-03-02

    Adenosine is an endogenous ubiquitous purine nucleoside, which is increased by hypoxia, ischaemia and tissue damage and mediates a number of physiopathological effects by interacting with four GPCRs, identified as A1 , A2A , A2B and A3 . Physiological and acutely increased adenosine is mostly associated with beneficial effects that include vasodilatation and a decrease in inflammation. In contrast, chronic overproduction of adenosine occurs in important pathological states, where long-lasting increases in the nucleoside levels are responsible for the bad side of adenosine associated with chronic inflammation, fibrosis and organ damage. In this review, we describe and critically discuss the pathological overproduction of adenosine and analyse when, where and how adenosine exerts its detrimental effects throughout the body.

  17. Torsades de pointes after adenosine administration.

    PubMed

    Teodorovich, Nicholay; Margolin, Elena; Kogan, Yonatan; Paz, Ofir; Swissa, Moshe

    2016-01-01

    Adenosine can produce arrhythmias, which are generally short living. It may induce PACs and PVCs, sinus bradycardia, and atrial fibrillation. There have been reports of transient polymorphic VT (torsades de pointes) in patients with LQTS and others in people with normal QT interval. We report a case of a long episode of polymorphic VT induced by adenosine. A 27 year old woman received 6 mg adenosine for PSVT, which terminated and torsades de pointes developed, persisting for 17 seconds and terminated spontaneously. This is the longest described duration of the torsades after adenosine administration in patients with normal QT interval.

  18. Partial separation of platelet and placental adenosine receptors from adenosine A2-like binding protein

    SciTech Connect

    Zolnierowicz, S.; Work, C.; Hutchison, K.; Fox, I.H. )

    1990-04-01

    The ubiquitous adenosine A2-like binding protein obscures the binding properties of adenosine receptors assayed with 5'-N-({sup 3}H)ethylcarboxamidoadenosine (({sup 3}H)NECA). To solve this problem, we developed a rapid and simple method to separate adenosine receptors from the adenosine A2-like binding protein. Human platelet and placental membranes were solubilized with 1% 3-((3-cholamidopropyl)dimethylammonio)-1-propanesulfonate. The soluble platelet extract was precipitated with polyethylene glycol and the fraction enriched in adenosine receptors was isolated from the precipitate by differential centrifugation. The adenosine A2-like binding protein was removed from the soluble placental extract with hydroxylapatite and adenosine receptors were precipitated with polyethylene glycol. The specificity of the ({sup 3}H)NECA binding is typical of an adenosine A2 receptor for platelets and an adenosine A1 receptor for placenta. This method leads to enrichment of adenosine A2 receptors for platelets and adenosine A1 receptors for placenta. This provides a useful preparation technique for pharmacologic studies of adenosine receptors.

  19. Autophagy occurs within an hour of adenosine triphosphate treatment after nerve cell damage: the neuroprotective effects of adenosine triphosphate against apoptosis

    PubMed Central

    Lu, Na; Wang, Baoying; Deng, Xiaohui; Zhao, Honggang; Wang, Yong; Li, Dongliang

    2014-01-01

    After hypoxia, ischemia, or inflammatory injuries to the central nervous system, the damaged cells release a large amount of adenosine triphosphate, which may cause secondary neuronal death. Autophagy is a form of cell death that also has neuroprotective effects. Cell Counting Kit assay, monodansylcadaverine staining, flow cytometry, western blotting, and real-time PCR were used to determine the effects of exogenous adenosine triphosphate treatment at different concentrations (2, 4, 6, 8, 10 mmol/L) over time (1, 2, 3, and 6 hours) on the apoptosis and autophagy of SH-SY5Y cells. High concentrations of extracellular adenosine triphosphate induced autophagy and apoptosis of SH-SY5Y cells. The enhanced autophagy first appeared, and peaked at 1 hour after treatment with adenosine triphosphate. Cell apoptosis peaked at 3 hours, and persisted through 6 hours. With prolonged exposure to the adenosine triphosphate treatment, the fraction of apoptotic cells increased. These data suggest that the SH-SY5Y neural cells initiated autophagy against apoptosis within an hour of adenosine triphosphate treatment to protect themselves against injury. PMID:25368646

  20. Autophagy occurs within an hour of adenosine triphosphate treatment after nerve cell damage: the neuroprotective effects of adenosine triphosphate against apoptosis.

    PubMed

    Lu, Na; Wang, Baoying; Deng, Xiaohui; Zhao, Honggang; Wang, Yong; Li, Dongliang

    2014-09-01

    After hypoxia, ischemia, or inflammatory injuries to the central nervous system, the damaged cells release a large amount of adenosine triphosphate, which may cause secondary neuronal death. Autophagy is a form of cell death that also has neuroprotective effects. Cell Counting Kit assay, monodansylcadaverine staining, flow cytometry, western blotting, and real-time PCR were used to determine the effects of exogenous adenosine triphosphate treatment at different concentrations (2, 4, 6, 8, 10 mmol/L) over time (1, 2, 3, and 6 hours) on the apoptosis and autophagy of SH-SY5Y cells. High concentrations of extracellular adenosine triphosphate induced autophagy and apoptosis of SH-SY5Y cells. The enhanced autophagy first appeared, and peaked at 1 hour after treatment with adenosine triphosphate. Cell apoptosis peaked at 3 hours, and persisted through 6 hours. With prolonged exposure to the adenosine triphosphate treatment, the fraction of apoptotic cells increased. These data suggest that the SH-SY5Y neural cells initiated autophagy against apoptosis within an hour of adenosine triphosphate treatment to protect themselves against injury.

  1. Adenosine actions on CA1 pyramidal neurones in rat hippocampal slices.

    PubMed

    Greene, R W; Haas, H L

    1985-09-01

    Intracellular recordings with a bridge amplifier of CA1 pyramidal neurones in vitro were employed to study the mechanisms of action of exogenously applied adenosine in the hippocampal slice preparation of the rat. Adenosine enhanced the calcium-dependent, long-duration after-hyperpolarization (a.h.p.) at least in part by a reduction in the rate of decay of the a.h.p. Both the reduced rate of decay and that of the control can be described with a single exponential. Antagonism of the calcium-dependent potassium current (and as a result, the a.h.p.) by bath application of CdCl2 or intracellular injection of EGTA (ethyleneglycolbis-(beta-aminoethyl ether)N,N'-tetraacetic acid) did not reduce the adenosine-evoked hyperpolarization or decrease in input resistance. Similarly, TEA (tetraethylammonium), which antagonizes both the voltage- and calcium-sensitive, delayed, outward rectification, had no effect on the adenosine-evoked changes in resting membrane properties. Adenosine did not affect the early, transient, outward rectification. During exposure to 4-aminopyridine (4-AP) in concentrations sufficient to antagonize this early rectification, the changes in resting membrane properties evoked by adenosine were unaffected. We conclude that the enhancement of the a.h.p. and accommodation by adenosine may be mediated by a change in the regulation of intracellular calcium. However, the mechanism responsible for the hyperpolarization and decrease in input resistance evoked by adenosine is both calcium and voltage insensitive. Thus, it appears distinct from that mediating the enhancement of the a.h.p. and accommodation.

  2. Adenosine actions on CA1 pyramidal neurones in rat hippocampal slices.

    PubMed Central

    Greene, R W; Haas, H L

    1985-01-01

    Intracellular recordings with a bridge amplifier of CA1 pyramidal neurones in vitro were employed to study the mechanisms of action of exogenously applied adenosine in the hippocampal slice preparation of the rat. Adenosine enhanced the calcium-dependent, long-duration after-hyperpolarization (a.h.p.) at least in part by a reduction in the rate of decay of the a.h.p. Both the reduced rate of decay and that of the control can be described with a single exponential. Antagonism of the calcium-dependent potassium current (and as a result, the a.h.p.) by bath application of CdCl2 or intracellular injection of EGTA (ethyleneglycolbis-(beta-aminoethyl ether)N,N'-tetraacetic acid) did not reduce the adenosine-evoked hyperpolarization or decrease in input resistance. Similarly, TEA (tetraethylammonium), which antagonizes both the voltage- and calcium-sensitive, delayed, outward rectification, had no effect on the adenosine-evoked changes in resting membrane properties. Adenosine did not affect the early, transient, outward rectification. During exposure to 4-aminopyridine (4-AP) in concentrations sufficient to antagonize this early rectification, the changes in resting membrane properties evoked by adenosine were unaffected. We conclude that the enhancement of the a.h.p. and accommodation by adenosine may be mediated by a change in the regulation of intracellular calcium. However, the mechanism responsible for the hyperpolarization and decrease in input resistance evoked by adenosine is both calcium and voltage insensitive. Thus, it appears distinct from that mediating the enhancement of the a.h.p. and accommodation. PMID:3932644

  3. Effects of adenosine, exercise, and moderate acute hypoxia on energy substrate utilization of human skeletal muscle.

    PubMed

    Heinonen, Ilkka; Kemppainen, Jukka; Kaskinoro, Kimmo; Peltonen, Juha E; Sipilä, Hannu T; Nuutila, Pirjo; Knuuti, Juhani; Boushel, Robert; Kalliokoski, Kari K

    2012-02-01

    Glucose metabolism increases in hypoxia and can be influenced by endogenous adenosine, but the role of adenosine for regulating glucose metabolism at rest or during exercise in hypoxia has not been elucidated in humans. We studied the effects of exogenous adenosine on human skeletal muscle glucose uptake and other blood energy substrates [free fatty acid (FFA) and lactate] by infusing adenosine into the femoral artery in nine healthy young men. The role of endogenous adenosine was studied by intra-arterial adenosine receptor inhibition (aminophylline) during dynamic one-leg knee extension exercise in normoxia and acute hypoxia corresponding to ∼3,400 m of altitude. Extraction and release of energy substrates were studied by arterial-to-venous (A-V) blood samples, and total uptake or release was determined by the product of A-V differences and muscle nutritive perfusion measured by positron emission tomography. The results showed that glucose uptake increased from a baseline value of 0.2 ± 0.2 to 2.0 ± 2.2 μmol·100 g(-1)·min(-1) during adenosine infusion (P < 0.05) at rest. Although acute hypoxia enhanced arterial FFA levels, it did not affect muscle substrate utilization at rest. During exercise, glucose uptake was higher (195%) during acute hypoxia compared with normoxia (P = 0.058), and aminophylline had no effect on energy substrate utilization during exercise, despite that arterial FFA levels were increased. In conclusion, exogenous adenosine at rest and acute moderate hypoxia during low-intensity knee-extension exercise increases skeletal muscle glucose uptake, but the increase in hypoxia appears not to be mediated by adenosine.

  4. Passive targeting of ischemic-reperfused myocardium with adenosine-loaded silica nanoparticles

    PubMed Central

    Galagudza, Michael; Korolev, Dmitry; Postnov, Viktor; Naumisheva, Elena; Grigorova, Yulia; Uskov, Ivan; Shlyakhto, Eugene

    2012-01-01

    Pharmacological agents suggested for infarct size limitation have serious side effects when used at cardioprotective doses which hinders their translation into clinical practice. The solution to the problem might be direct delivery of cardioprotective drugs into ischemic-reperfused myocardium. In this study, we explored the potential of silica nanoparticles for passive delivery of adenosine, a prototype cardioprotective agent, into ischemic-reperfused heart tissue. In addition, the biodegradation of silica nanoparticles was studied both in vitro and in vivo. Immobilization of adenosine on the surface of silica nanoparticles resulted in enhancement of adenosine-mediated infarct size limitation in the rat model. Furthermore, the hypotensive effect of adenosine was attenuated after its adsorption on silica nanoparticles. We conclude that silica nanoparticles are biocompatible materials that might potentially be used as carriers for heart-targeted drug delivery. PMID:22619519

  5. Adenosine modulates cell growth in the human breast cancer cells via adenosine receptors.

    PubMed

    Panjehpour, Mojtaba; Karami-Tehrani, Fatemeh

    2007-01-01

    Adenosine modulates the proliferation, survival, and apoptosis of many different cell types. The present study was performed to investigate the role of adenosine receptors in the human breast cancer cell lines MCF-7 and MDA-MB468. The biological effects of adenosine on the cells were analyzed by adenylyl cyclase and cell viability assay as well as RT-PCR of adenosine receptors. RT-PCR results show the expression of the transcript of all adenosine receptors in both cell lines. By using adenosine and selective adenosine receptor agonists or antagonists, we found that A3 stimulation reduced cell viability, which was abolished by pretreatment with A3 receptor antagonist. Moreover, we demonstrated that adenosine (natural agonist) triggers a cytotoxic signal via A3 receptor activation that was not seen for other subclasses of adenosine receptors. Intracellular cAMP concentration was changed significantly only for A3 and A2B receptor-selective agonists, which indicates the functional form of these receptors on the cell surface. In conclusion, our findings revealed the role of adenosine receptors in breast cancer cell lines on growth modulation role of A3 and functional form of A2B, although its involvement in cell growth modulation was not seen. Theses findings as well as data by others may provide a possible application of adenosine receptor agonists/antagonists in breast malignancies.

  6. Green tea catechins enhance norepinephrine-induced lipolysis via a protein kinase A-dependent pathway in adipocytes.

    PubMed

    Chen, Shu; Osaki, Noriko; Shimotoyodome, Akira

    2015-05-22

    Green tea catechins have been shown to attenuate obesity in animals and humans. The catechins activate adenosine monophosphate-activated protein kinase (AMPK), and thereby increase fatty acid oxidation in liver and skeletal muscles. Green tea catechins have also been shown to reduce body fat in humans. However, the effect of the catechins on lipolysis in adipose tissue has not been fully understood. The aim of this study was to clarify the effect of green tea catechins on lipolysis in adipocytes and to elucidate the underlying mechanism. Differentiated mouse adipocyte cell line (3T3-L1) was stimulated with green tea catechins in the presence or absence of norepinephrine. Glycerol and free fatty acids in the media were measured. Phosphorylation of hormone-sensitive lipase (HSL) was determined by Western blotting, and the mRNA expression levels of HSL, adipose triglyceride lipase (ATGL), and perilipin were determined by quantitative RT-PCR. The cells were treated with inhibitors of protein kinase A (PKA), protein kinase C (PKC), protein kinase G (PKG), or mitogen-activated protein kinase (MAPK) to determine the responsible pathway. Treatment of 3T3-L1 adipocytes with green tea catechins increased the level of glycerol and free fatty acids released into the media in the presence, but not absence, of norepinephrine, and increased the level of phosphorylated HSL in the cells. The catechins also increased mRNA and protein levels of HSL and ATGL. PKA inhibitor (H89) attenuated the catechin-induced increase in glycerol release and HSL phosphorylation. The results demonstrate that green tea catechins enhance lipolysis in the presence of norepinephrine via a PKA-dependent pathway in 3T3-L1 adipocytes, providing a potential mechanism by which green tea catechins could reduce body fat. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis

    SciTech Connect

    Dong, Chenglong; Zheng, Haining; Huang, Shanshan; You, Na; Xu, Jiarong; Ye, Xiaolong; Zhu, Qun; Feng, Yamin; You, Qiang; Miao, Heng; Ding, Dafa; Lu, Yibing

    2015-10-01

    Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12–24 h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy. - Highlights: • High glucose leads to increased autophagy in podocytes at an early stage. • The early autophagic response protects against high glucose-induced apoptosis. • Heme oxygenase-1 enhances autophagy and decreases high glucose -mediated apoptosis. • Heme oxygenase-1 induces autophagy through the activation of AMPK.

  8. Adenosine signaling promotes regeneration of pancreatic β-cells in vivo

    PubMed Central

    Andersson, Olov; Adams, Bruce A.; Yoo, Daniel; Ellis, Gregory C.; Gut, Philipp; Anderson, Ryan M.; German, Michael S.; Stainier, Didier Y. R.

    2012-01-01

    Diabetes can be controlled with insulin injections, but a curative approach that restores the number of insulin-producing β-cells is still needed. Using a zebrafish model of diabetes, we screened ~7000 small molecules to identify enhancers of β-cell regeneration. The compounds we identified converge on the adenosine signaling pathway and include exogenous agonists and compounds that inhibit degradation of endogenously produced adenosine. The most potent enhancer of β-cell regeneration was the adenosine agonist 5′-N-Ethylcarboxamidoadenosine (NECA), which acting through the adenosine receptor A2aa increased β-cell proliferation and accelerated restoration of normoglycemia in zebrafish. Despite markedly stimulating β-cell proliferation during regeneration, NECA had only a modest effect during development. The proliferative and glucose-lowering effect of NECA was confirmed in diabetic mice, suggesting an evolutionarily conserved role for adenosine in β-cell regeneration. With this whole-organism screen, we identified components of the adenosine pathway that could be therapeutically targeted for the treatment of diabetes. PMID:22608007

  9. Enzymatic regeneration of adenosine triphosphate cofactor

    NASA Technical Reports Server (NTRS)

    Marshall, D. L.

    1974-01-01

    Regenerating adenosine triphosphate (ATP) from adenosine diphosphate (ADP) by enzymatic process which utilizes carbamyl phosphate as phosphoryl donor is technique used to regenerate expensive cofactors. Process allows complex enzymatic reactions to be considered as candidates for large-scale continuous processes.

  10. Endogenous adenosine and adenosine receptors localized to ganglion cells of the retina

    SciTech Connect

    Braas, K.M.; Zarbin, M.A.; Snyder, S.H.

    1987-06-01

    Using specific sensitive antisera against adenosine, we have immunocytochemically localized endogenous adenosine to specific layers of rat, guinea pig, monkey, and human retina. Highest adenosine immunoreactivity was observed in ganglion cells and their processes in the optic nerve fiber layer. Substantial staining was also found throughout the inner plexiform layer and in select cells in the inner nuclear layer. Adenosine A1 receptors, labeled with the agonists L-(/sup 3/H)phenylisopropyladenosine and /sup 125/I-labeled hydroxy-phenylisopropyladenosine, were autoradiographically localized. The highest levels of binding sites occurred in the nerve fiber, ganglion cell, and inner plexiform layers of the retina in all the species examined. The distribution of adenosine A1 receptor sites closely parallels that of retinal neurons and fibers containing immunoreactive adenosine. These results suggest a role for endogenous adenosine as a coneurotransmitter in ganglion cells and their fibers in the optic nerve.

  11. Insulin/adenosine axis linked signalling.

    PubMed

    Silva, Luis; Subiabre, Mario; Araos, Joaquín; Sáez, Tamara; Salsoso, Rocío; Pardo, Fabián; Leiva, Andrea; San Martín, Rody; Toledo, Fernando; Sobrevia, Luis

    2017-06-01

    Regulation of blood flow depends on systemic and local release of vasoactive molecules such as insulin and adenosine. These molecules cause vasodilation by activation of plasma membrane receptors at the vascular endothelium. Adenosine activates at least four subtypes of adenosine receptors (A1AR, A2AAR, A2BAR, A3AR), of which A2AAR and A2BAR activation leads to increased cAMP level, generation of nitric oxide, and relaxation of the underlying smooth muscle cell layer. Vasodilation caused by adenosine also depends on plasma membrane hyperpolarization due to either activation of intermediate-conductance Ca(2+)-activated K(+) channels in vascular smooth muscle or activation of ATP-activated K(+) channels in the endothelium. Adenosine also causes vasoconstriction via a mechanism involving A1AR activation resulting in lower cAMP level and increased thromboxane release. Insulin has also a dual effect causing NO-dependent vasodilation, but also sympathetic activity- and increased endothelin 1 release-dependent vasoconstriction. Interestingly, insulin effects require or are increased by activation or inactivation of adenosine receptors. This is phenomenon described for d-glucose and l-arginine transport where A2AAR and A2BAR play a major role. Other studies show that A1AR activation could reduce insulin release from pancreatic β-cells. Whether adenosine modulation of insulin biological effect is a phenomenon that depends on co-localization of adenosine receptors and insulin receptors, and adenosine plasma membrane transporters is something still unclear. This review summarizes findings addressing potential involvement of adenosine receptors to modulate insulin effect via insulin receptors with emphasis in the human vasculature. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Halobacterial adenosine triphosphatases and the adenosine triphosphatase from Halobacterium saccharovorum

    NASA Technical Reports Server (NTRS)

    Kristjansson, Hordur; Sadler, Martha H.; Hochstein, Lawrence I.

    1986-01-01

    Membranes prepared from various members of the genus Halobacterium contained a Triton X-l00 activated adenosine triphosphatase. The enzyme from Halobacterium saccharovorum was unstable in solutions of low ionic strength and maximally active in the presence of 3.5 M NaCl. A variety of nucleotide triphosphates was hydrolyzed. MgADP, the product of ATP hydrolysis, was not hydrolyzed and was a competitive inhibitor with respect to MgATP. The enzyme from H. saccharovorum was composed of at least 2 and possibly 4 subunits. The 83-kDa and 60-kDa subunits represented about 90 percent of total protein. The 60-kDa subunit reacted with dicyclohexyl-carbodiimide when inhibition was carried out in an acidic medium. The enzyme from H. saccharovorum, possesses properties of an F(1)F(0) as well as an E(1)E(2) ATPase.

  13. Halobacterial adenosine triphosphatases and the adenosine triphosphatase from Halobacterium saccharovorum

    NASA Technical Reports Server (NTRS)

    Kristjansson, Hordur; Sadler, Martha H.; Hochstein, Lawrence I.

    1986-01-01

    Membranes prepared from various members of the genus Halobacterium contained a Triton X-l00 activated adenosine triphosphatase. The enzyme from Halobacterium saccharovorum was unstable in solutions of low ionic strength and maximally active in the presence of 3.5 M NaCl. A variety of nucleotide triphosphates was hydrolyzed. MgADP, the product of ATP hydrolysis, was not hydrolyzed and was a competitive inhibitor with respect to MgATP. The enzyme from H. saccharovorum was composed of at least 2 and possibly 4 subunits. The 83-kDa and 60-kDa subunits represented about 90 percent of total protein. The 60-kDa subunit reacted with dicyclohexyl-carbodiimide when inhibition was carried out in an acidic medium. The enzyme from H. saccharovorum, possesses properties of an F(1)F(0) as well as an E(1)E(2) ATPase.

  14. Impairment of ATP hydrolysis decreases adenosine A1 receptor tonus favoring cholinergic nerve hyperactivity in the obstructed human urinary bladder.

    PubMed

    Silva-Ramos, M; Silva, I; Faria, M; Magalhães-Cardoso, M T; Correia, J; Ferreirinha, F; Correia-de-Sá, P

    2015-12-01

    This study was designed to investigate whether reduced adenosine formation linked to deficits in extracellular ATP hydrolysis by NTPDases contributes to detrusor neuromodulatory changes associated with bladder outlet obstruction in men with benign prostatic hyperplasia (BPH). The kinetics of ATP catabolism and adenosine formation as well as the role of P1 receptor agonists on muscle tension and nerve-evoked [(3)H]ACh release were evaluated in mucosal-denuded detrusor strips from BPH patients (n = 31) and control organ donors (n = 23). The neurogenic release of ATP and [(3)H]ACh was higher (P < 0.05) in detrusor strips from BPH patients. The extracellular hydrolysis of ATP and, subsequent, adenosine formation was slower (t (1/2) 73 vs. 36 min, P < 0.05) in BPH detrusor strips. The A(1) receptor-mediated inhibition of evoked [(3)H]ACh release by adenosine (100 μM), NECA (1 μM), and R-PIA (0.3 μM) was enhanced in BPH bladders. Relaxation of detrusor contractions induced by acetylcholine required 30-fold higher concentrations of adenosine. Despite VAChT-positive cholinergic nerves exhibiting higher A(1) immunoreactivity in BPH bladders, the endogenous adenosine tonus revealed by adenosine deaminase is missing. Restoration of A1 inhibition was achieved by favoring (1) ATP hydrolysis with apyrase (2 U mL(-1)) or (2) extracellular adenosine accumulation with dipyridamole or EHNA, as these drugs inhibit adenosine uptake and deamination, respectively. In conclusion, reduced ATP hydrolysis leads to deficient adenosine formation and A(1) receptor-mediated inhibition of cholinergic nerve activity in the obstructed human bladder. Thus, we propose that pharmacological manipulation of endogenous adenosine levels and/or A(1) receptor activation might be useful to control bladder overactivity in BPH patients.

  15. Homeostatic control of synaptic activity by endogenous adenosine is mediated by adenosine kinase.

    PubMed

    Diógenes, Maria José; Neves-Tomé, Raquel; Fucile, Sergio; Martinello, Katiuscia; Scianni, Maria; Theofilas, Panos; Lopatár, Jan; Ribeiro, Joaquim A; Maggi, Laura; Frenguelli, Bruno G; Limatola, Cristina; Boison, Detlev; Sebastião, Ana M

    2014-01-01

    Extracellular adenosine, a key regulator of neuronal excitability, is metabolized by astrocyte-based enzyme adenosine kinase (ADK). We hypothesized that ADK might be an upstream regulator of adenosine-based homeostatic brain functions by simultaneously affecting several downstream pathways. We therefore studied the relationship between ADK expression, levels of extracellular adenosine, synaptic transmission, intrinsic excitability, and brain-derived neurotrophic factor (BDNF)-dependent synaptic actions in transgenic mice underexpressing or overexpressing ADK. We demonstrate that ADK: 1) Critically influences the basal tone of adenosine, evaluated by microelectrode adenosine biosensors, and its release following stimulation; 2) determines the degree of tonic adenosine-dependent synaptic inhibition, which correlates with differential plasticity at hippocampal synapses with low release probability; 3) modulates the age-dependent effects of BDNF on hippocampal synaptic transmission, an action dependent upon co-activation of adenosine A2A receptors; and 4) influences GABAA receptor-mediated currents in CA3 pyramidal neurons. We conclude that ADK provides important upstream regulation of adenosine-based homeostatic function of the brain and that this mechanism is necessary and permissive to synaptic actions of adenosine acting on multiple pathways. These mechanistic studies support previous therapeutic studies and implicate ADK as a promising therapeutic target for upstream control of multiple neuronal signaling pathways crucial for a variety of neurological disorders.

  16. Halogenated pyrrolopyrimidine analogues of adenosine from marine organisms: pharmacological activities and potent inhibition of adenosine kinase.

    PubMed

    Davies, L P; Jamieson, D D; Baird-Lambert, J A; Kazlauskas, R

    1984-02-01

    Two novel halogenated pyrrolopyrimidine analogues of adenosine, isolated from marine sources, have been examined for pharmacological and biochemical activities. 4-Amino-5-bromo-pyrrolo[2,3-d]pyrimidine, from a sponge of the genus Echinodictyum, had bronchodilator activity at least as potent as theophylline but with a different biochemical profile; unlike theophylline it had no antagonist activity at CNS adenosine receptors and it was quite a potent inhibitor of adenosine uptake and adenosine kinase in brain tissue. 5'-Deoxy-5-iodotubercidin, isolated from the red alga Hypnea valentiae, caused potent muscle relaxation and hypothermia when injected into mice. This compound was a very potent inhibitor of adenosine uptake into rat and guinea-pig brain slices and an extremely potent inhibitor of adenosine kinase from guinea-pig brain and rat brain and liver. Neither of these two pyrrolopyrimidine analogues was a substrate for, or an inhibitor of, adenosine deaminase. Neither compound appeared to have any direct agonist activity on guinea-pig brain adenosine-stimulated adenylate cyclase (A2 adenosine receptors). 5'-Deoxy-5-iodotubercidin is unique in two respects: it appears to be the first naturally-occurring example of a 5'-deoxyribosyl nucleoside and is the first example of a specifically iodinated nucleoside from natural sources. It may be the most potent adenosine kinase inhibitor yet described and, by virtue of its structure, may prove to be the most specific.

  17. Adenosine administration accelerates progression of the cell cycle during rat liver regeneration induced by one-third hepatectomy.

    PubMed

    Mendieta-Condado, Edgar; Pichardo-Olvera, Mariana; Sánchez-Sevilla, Lourdes; Chagoya de Sánchez, Victoria; Hernández-Muñoz, Rolando

    2009-10-01

    We have shown that adenosine administration is capable of reversing fibrosis in the carbon tetrachloride-induced rat cirrhotic liver, stimulating the diminished proliferative potential of the cirrhotic liver. To characterize adenosine actions on liver cellular proliferation, we used rats subjected to one-third partial hepatectomy (PH). In PH animals acutely administered with adenosine (25-200 mg/kg b.w.), parameters indicative of cell proliferation were determined. In addition, hepatocyte growth factor (HGF), epidermal growth factor, and transforming growth factor-alpha, cyclins, members of the E2F family, proto-oncogenes, and adenosine-receptors were determined through Western blot analyses. Adenosine (100 mg/kg body weight) induced an earlier increase in liver cell proliferation as evidenced by enhanced levels of proliferating cell nuclear antigen, nuclear Ki-67 antigen, and those for cyclins (D1, E, A, and B1), as well as by an increased mitotic index. These effects were also accompanied for a long-lasting increase of serum and liver levels of HGF and liver expression of c-Met and HGF liver activator. Adenosine effects on cell proliferation could be mediated by an early increase in E2F-1 and by that of c-Myc, despite the fact that phosphorylation of the Rb protein and expression of E2F-3 were decreased. Moreover, the liver amount of specific receptors for adenosine was not significantly changed by PH and/or adenosine treatment. In conclusion, these data suggest that adenosine actions can accelerate and increase proliferation in a "primed" liver, mainly through enhancing c-Myc, E2F family, cell-cycle cyclins, and HGF expression. Therefore, these pharmacological adenosine effects suggest a modulating role for the nucleoside on mitogenic events once the liver has been triggered to proliferate.

  18. Adenosine receptor control of cognition in normal and disease.

    PubMed

    Chen, Jiang-Fan

    2014-01-01

    Adenosine and adenosine receptors (ARs) are increasingly recognized as important therapeutic targets for controlling cognition under normal and disease conditions for its dual roles of neuromodulation as well as of homeostatic function in the brain. This chapter first presents the unique ability of adenosine, by acting on the inhibitory A1 and facilitating A2A receptor, to integrate dopamine, glutamate, and BNDF signaling and to modulate synaptic plasticity (e.g., long-term potentiation and long-term depression) in brain regions relevant to learning and memory, providing the molecular and cellular bases for adenosine receptor (AR) control of cognition. This led to the demonstration of AR modulation of social recognition memory, working memory, reference memory, reversal learning, goal-directed behavior/habit formation, Pavlovian fear conditioning, and effort-related behavior. Furthermore, human and animal studies support that AR activity can also, through cognitive enhancement and neuroprotection, reverse cognitive impairments in animal models of Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease, and schizophrenia. Lastly, epidemiological evidence indicates that regular human consumption of caffeine, the most widely used psychoactive drug and nonselective AR antagonists, is associated with the reduced cognitive decline in aging and AD patients, and with the reduced risk in developing PD. Thus, there is a convergence of the molecular studies revealing AR as molecular targets for integrating neurotransmitter signaling and controlling synaptic plasticity, with animal studies demonstrating the strong procognitive impact upon AR antagonism in normal and disease brains and with epidemiological and clinical evidences in support of caffeine and AR drugs for therapeutic modulation of cognition. Since some of adenosine A2A receptor antagonists are already in phase III clinical trials for motor benefits in PD patients with remarkable safety profiles

  19. Optical Aptasensors for Adenosine Triphosphate

    PubMed Central

    Ng, Stella; Lim, Hui Si; Ma, Qian; Gao, Zhiqiang

    2016-01-01

    Nucleic acids are among the most researched and applied biomolecules. Their diverse two- and three-dimensional structures in conjunction with their robust chemistry and ease of manipulation provide a rare opportunity for sensor applications. Moreover, their high biocompatibility has seen them being used in the construction of in vivo assays. Various nucleic acid-based devices have been extensively studied as either the principal element in discrete molecule-like sensors or as the main component in the fabrication of sensing devices. The use of aptamers in sensors - aptasensors, in particular, has led to improvements in sensitivity, selectivity, and multiplexing capacity for a wide verity of analytes like proteins, nucleic acids, as well as small biomolecules such as glucose and adenosine triphosphate (ATP). This article reviews the progress in the use of aptamers as the principal component in sensors for optical detection of ATP with an emphasis on sensing mechanism, performance, and applications with some discussion on challenges and perspectives. PMID:27446501

  20. The adenosine neuromodulation system in schizophrenia.

    PubMed

    Rial, Daniel; Lara, Diogo R; Cunha, Rodrigo A

    2014-01-01

    The management of schizophrenia endophenotypes, namely positive, negative, and cognitive symptoms is still an open goal, justifying the search of novel therapeutic avenues. We now review the evidence supporting the interest in targeting the adenosine modulation system to counteract the core features of schizophrenia. This interest is forwarded by the combined ability of strategies aimed at bolstering adenosine levels together with the increasingly recognized impact of adenosine A2A receptors to control dopaminergic signaling, working memory, and behavioral sensitization; this is further heralded by the suggested clinical effectiveness of therapies increasing extracellular adenosine such as dipyridamole and allopurinol and the emergent recognition of a role for adenosine in neurodevelopment. Finally, the combined role of A1 and A2A receptors in assisting the implementation of adaptive changes and encoding of information salience in neuronal circuits together with the adaptive alterations of A1 and A2A receptor density upon brain dysfunction prompts the novel working hypothesis that the parallel imbalance of adenosine formation and of A1 and A2A receptors blurs the adequate encoding of information salience in neuronal circuits, which we propose to be a core pathogenic feature in the development of schizophrenia endophenotypes. This proposal should also provide a rationale to assist the design of future therapeutic intervention targeting the adenosine modulation system to manage schizophrenia endophenotypes: these should not be based only on an attempt to target adenosine kinase-A1 receptors or only A2A receptors, but should instead simultaneously target these two arms of the adenosine modulation system. © 2014 Elsevier Inc. All rights reserved.

  1. Adenosine Kinase: Exploitation for Therapeutic Gain

    PubMed Central

    2013-01-01

    Adenosine kinase (ADK; EC 2.7.1.20) is an evolutionarily conserved phosphotransferase that converts the purine ribonucleoside adenosine into 5′-adenosine-monophosphate. This enzymatic reaction plays a fundamental role in determining the tone of adenosine, which fulfills essential functions as a homeostatic and metabolic regulator in all living systems. Adenosine not only activates specific signaling pathways by activation of four types of adenosine receptors but it is also a primordial metabolite and regulator of biochemical enzyme reactions that couple to bioenergetic and epigenetic functions. By regulating adenosine, ADK can thus be identified as an upstream regulator of complex homeostatic and metabolic networks. Not surprisingly, ADK dysfunction is involved in several pathologies, including diabetes, epilepsy, and cancer. Consequently, ADK emerges as a rational therapeutic target, and adenosine-regulating drugs have been tested extensively. In recent attempts to improve specificity of treatment, localized therapies have been developed to augment adenosine signaling at sites of injury or pathology; those approaches include transplantation of stem cells with deletions of ADK or the use of gene therapy vectors to downregulate ADK expression. More recently, the first human mutations in ADK have been described, and novel findings suggest an unexpected role of ADK in a wider range of pathologies. ADK-regulating strategies thus represent innovative therapeutic opportunities to reconstruct network homeostasis in a multitude of conditions. This review will provide a comprehensive overview of the genetics, biochemistry, and pharmacology of ADK and will then focus on pathologies and therapeutic interventions. Challenges to translate ADK-based therapies into clinical use will be discussed critically. PMID:23592612

  2. AMID mediates adenosine-induced caspase-independent HuH-7 cell apoptosis.

    PubMed

    Yang, Dongqin; Yaguchi, Takahiro; Nagata, Tetsu; Gotoh, Akinobu; Dovat, Sinisa; Song, Chunhua; Nishizaki, Tomoyuki

    2011-01-01

    The mechanism underlying extracellular adenosine-induced caspase-independent apoptosis in HuH-7 human hepatoma cells is not fully understood. The present study investigated the role for apoptosis-inducing factor (AIF)-homologous mitochondrion-associated inducer of death (AMID) in the pathway. To see the implication of AMID in adenosine-induced HuH-7 cell apoptosis, real-time reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescent cytochemistry, time-laps GFP monitoring, cell cycle analysis, flow cytometry, Western blotting, cell viability assay, and TUNEL staining were carried out. Adenosine upregulated AMID expression in HuH-7 cells, and translocated AMID from the cytosol into the nucleus. Adenosine induced HuH-7 cell apoptosis, and the effect was further enhanced by overexpressing AMID. Adenosine-induced HuH-7 cell apoptosis, alternatively, was inhibited by knocking-down AMID. The results of the present study provide evidence for AMID as a critical factor for adenosine-induced caspase-independent HuH-7 cell apoptosis. Copyright © 2011 S. Karger AG, Basel.

  3. Responsiveness of renal glomeruli to adenosine in streptozotocin-induced diabetic rats dependent on hyperglycaemia level.

    PubMed

    Szczepańska-Konkel, M; Jankowski, M; Stiepanow-Trzeciak, A; Rudzik, A; Pawełczyk, T; Angielski, S

    2003-03-01

    Glomerular filtration rate (GFR) in response to adenosine precursor, NAD, and glomeruli contractility in response to adenosine were evaluated in streptozotocin-induced diabetic rats with severe (blood glucose 27.8 +/- 1.2 mmol/L) and moderate hyperglycaemia (18.2 +/- 0.9 mmol/L) compared with nondiabetic (ND)-rats. In anaesthetised rats, basal GFR was greater in moderately diabetic rats compared with severely diabetic rats (p < 0.05) and ND-rats (p < 0.02). Intravenous infusion of 5 nmol x min(-1) x kg(-1) NAD reduced GFR and renal plasma flow (RPF) in diabetic rats but had no effect on these parameters in ND-rats. Moreover, NAD-induced reduction of GFR and RPF was greater in rats with severe diabetes (41% and 30%, respectively) than in with moderate diabetes (25% and 26%, respectively). Theophylline (0.2 micromol x min(-1) x kg(-1) ) abolished renal response to NAD. Isolated glomeruli contraction in response to adenosine, assessed by glomerular 3H-inulin space reduction, was lowered in moderately diabetic-group and enhanced in severely diabetic-group. compared with ND-group (p < 0.05). Adenosine A1-receptor antagonist DPCPX inhibited adenosine-induced glomeruli contraction. This differential response of diabetic renal glomeruli to adenosine suggests that impaired glomerular contractility in response to adenosine could be responsible for hyperfiltration in moderate diabets, whereas, the increased adenosine-dependent contractility of glomeruli in severe diabetes may increase the risk of acute renal failure in this condition.

  4. Polarized fibronectin secretion induced by adenosine regulates bacterial–epithelial interaction in human intestinal epithelial cells

    PubMed Central

    2004-01-01

    Fibronectin (FN) is a multifunctional protein that plays important roles in many biological processes including cell adhesion and migration, wound healing and inflammation. Cellular FNs are produced by a wide variety of cell types including epithelial cells, which secrete them and often organize them into extensive extracellular matrices at their basal surface. However, regulation of FN synthesis and the polarity of FN secretion by intestinal epithelial cells have not been investigated. In the present study we investigated the role of adenosine, whose levels are up-regulated during inflammation, in modulating FN synthesis, the polarity of FN secretion and the downstream effects of the secreted FN. Polarized monolayers of T84 cells were used as an intestinal epithelial model. Adenosine added to either the apical or basolateral aspect of the cells led to a time- and dose-dependent accumulation of FN in the culture supernatants, polarized to the apical compartment and reached maximal levels 24 h after apical or basolateral addition of adenosine. Confocal microscopy confirmed that FN localized to the apical domain of model intestinal epithelial cells stimulated with apical or basolateral adenosine. The induction of FN was significantly down-regulated in response to the adenosine receptor antagonist alloxazine and was inhibited by cycloheximide. Moreover, adenosine increased FN promoter activity (3.5-fold compared with unstimulated controls) indicating that FN induction is, in part, transcriptionally regulated. Interestingly, we demonstrated that adenosine, as well as apical FN, significantly enhanced the adherence and invasion of Salmonella typhimurium into cultured epithelial cells. In summary, we have shown for the first time that FN, a classic extracellular matrix protein, is secreted into the apical compartment of epithelial cells in response to adenosine. FN may be a critical host factor that modulates adherence and invasion of bacteria, thus playing a key role

  5. Repeated administration of adenosine increases its cardiovascular effects in rats.

    PubMed

    Vidrio, H; García-Márquez, F; Magos, G A

    1987-01-20

    Hypotensive and negative chronotropic responses to adenosine in anesthetized rats increased after previous administration of the nucleoside. Bradycardia after adenosine in the isolated perfused rat heart was also potentiated after repeated administration at short intervals. This self-potentiation could be due to extracellular accumulation of adenosine and persistent stimulation of receptors caused by saturation or inhibition of cellular uptake of adenosine.

  6. Adenosine-to-Inosine RNA Editing in Health and Disease.

    PubMed

    Gatsiou, Aikaterini; Vlachogiannis, Nikolaos; Lunella, Federica Francesca; Sachse, Marco; Stellos, Konstantinos

    2017-09-26

    Adenosine deamination in transcriptome results in the formation of inosine, a process that is called A-to-I RNA editing. Adenosine deamination is one of the more than 140 described RNA modifications. A-to-I RNA editing is catalyzed by adenosine deaminase acting on RNA (ADAR) enzymes and is essential for life. Recent Advances: Accumulating evidence supports a critical role of RNA editing in all aspects of RNA metabolism, including mRNA stability, splicing, nuclear export, and localization, as well as in recoding of proteins. These advances have significantly enhanced the understanding of mechanisms involved in development and in homeostasis. Furthermore, recent studies have indicated that RNA editing may be critically involved in cancer, aging, neurological, autoimmune, or cardiovascular diseases. This review summarizes recent and significant achievements in the field of A-to-I RNA editing and discusses the importance and translational value of this RNA modification for gene expression, cellular, and organ function, as well as for disease development. Elucidation of the exact RNA editing-dependent mechanisms in a single-nucleotide level may pave the path toward the development of novel therapeutic strategies focusing on modulation of ADAR function in the disease context. Antioxid. Redox Signal. 00, 000-000.

  7. Some aspects of adenosine triphosphate synthesis from adenine and adenosine in human red blood cells

    PubMed Central

    Whittam, R.; Wiley, J. S.

    1968-01-01

    1. The synthesis of ATP has been studied in human erythrocytes. Fresh cells showed no net synthesis of ATP when incubated with adenine or adenosine, although labelled adenine was incorporated into ATP in small amounts. 2. Cold-stored cells (3-6 weeks old) became progressively depleted of adenine nucleotides but incubation with adenosine or adenine plus inosine restored the ATP concentration to normal within 4 hr. Incorporation of labelled adenine or adenosine into the ATP of incubated stored cells corresponded to net ATP synthesis by these cells. 3. Synthesis of ATP from adenosine plus adenine together was 75% derived from adenine and only 25% from adenosine, indicating that nucleotide synthesis from adenine inhibits the simultaneous synthesis of nucleotide from adenosine. PMID:5723519

  8. Impact on monoclonal antibody production in murine hybridoma cell cultures of adenosine receptor antagonists and phosphodiesterase inhibitors.

    PubMed

    Kelso, Geoffrey F; Kazi, Shahid A; Harris, Simon J; Boysen, Reinhard I; Chowdhury, Jamil; Hearn, Milton T W

    2016-01-15

    The effects of different adenosine receptor antagonists and cyclic nucleotide phosphodiesterase (PDE) inhibitors on monoclonal antibody (mAb) titer and cell viability of murine hybridoma cells in culture were measured as part of our investigations to discover additives that enhance mAb production. Specific adenosine receptor antagonists and PDE inhibitors were found to enhance or decrease the titer of immunoglobulin G1 (IgG1) mAbs relative to negative controls, depending on the specific compound and cell line employed. The observed enhancements or decreases in IgG1 mAb titer appeared to be mainly due to an increase or decrease in specific productivity rates (ngmAb/cell), respectively. The different effects of the selective adenosine antagonists suggest that antagonism at the level of the adenosine A2A and A1 or the adenosine A3 receptors result in either enhancement or suppression of IgG1 mAb production by hybridoma cells. Overall, these studies have identified hitherto unknown activities of specific adenosine antagonists and PDE inhibitors which indicate they may have valuable roles as cell culture additives in industrial biomanufacturing processes designed to enhance the yields of mAbs or other recombinant proteins produced by mammalian cell culture procedures.

  9. Genetics Home Reference: adenosine deaminase 2 deficiency

    MedlinePlus

    ... This Page Bras J, Guerreiro R, Santo GC. Mutant ADA2 in vasculopathies. N Engl J Med. 2014 ... M, Anikster Y, King MC, Levy-Lahad E. Mutant adenosine deaminase 2 in a polyarteritis nodosa vasculopathy. ...

  10. [Adenosine deaminase in experimental trypanosomiasis: future implications].

    PubMed

    Pérez-Aguilar, Mary Carmen; Rondón-Mercado, Rocío

    2015-09-01

    The adenosine deaminase represents a control point in the regulation of extracellular adenosine levels, thus playing a critical role in the modulation of purinergic responses to certain pathophysiological events. Several studies have shown that serum and plasma enzyme levels are elevated in some diseases caused by microorganisms, which may represent a compensatory mechanism due to the elevated levels of adenosine and the release of inflammatory mediators. Recent research indicates that adenosine deaminase activity decreases and affects hematological parameters of infected animals with Trypanosoma evansi, so that such alterations could have implications in the pathogenesis of the disease. In addition, the enzyme has been detected in this parasite; allowing the inference that it could be associated with the vital functions of the same, similar to what occurs in mammals. This knowledge may be useful in the association of chemotherapy with specific inhibitors of the enzyme in future studies.

  11. Role of adenosine receptors in caffeine tolerance

    SciTech Connect

    Holtzman, S.G.; Mante, S.; Minneman, K.P. )

    1991-01-01

    Caffeine is a competitive antagonist at adenosine receptors. Receptor up-regulation during chronic drug treatment has been proposed to be the mechanism of tolerance to the behavioral stimulant effects of caffeine. This study reassessed the role of adenosine receptors in caffeine tolerance. Separate groups of rats were given scheduled access to drinking bottles containing plain tap water or a 0.1% solution of caffeine. Daily drug intake averaged 60-75 mg/kg and resulted in complete tolerance to caffeine-induced stimulation of locomotor activity, which could not be surmounted by increasing the dose of caffeine. 5'-N-ethylcarboxamidoadenosine (0.001-1.0 mg/kg) dose dependently decreased the locomotor activity of caffeine-tolerant rats and their water-treated controls but was 8-fold more potent in the latter group. Caffeine (1.0-10 mg/kg) injected concurrently with 5-N-ethylcarboxamidoadenosine antagonized the decreases in locomotor activity comparably in both groups. Apparent pA2 values for tolerant and control rats also were comparable: 5.05 and 5.11. Thus, the adenosine-antagonist activity of caffeine was undiminished in tolerant rats. The effects of chronic caffeine administration on parameters of adenosine receptor binding and function were measured in cerebral cortex. There were no differences between brain tissue from control and caffeine-treated rats in number and affinity of adenosine binding sites or in receptor-mediated increases (A2 adenosine receptor) and decreases (A1 adenosine receptor) in cAMP accumulation. These results are consistent with theoretical arguments that changes in receptor density should not affect the potency of a competitive antagonist. Experimental evidence and theoretical considerations indicate that up-regulation of adenosine receptors is not the mechanism of tolerance to caffeine-induced stimulation of locomotor activity.

  12. Control of basal extracellular adenosine concentration in rat cerebellum

    PubMed Central

    Wall, Mark J; Atterbury, Alison; Dale, Nicholas

    2007-01-01

    To re-examine how the basal extracellular concentration of adenosine is regulated in acutely isolated cerebellar slices we have combined electrophysiological and microelectrode biosensor measurements. In almost all cases, synaptic transmission was tonically inhibited by adenosine acting via A1 receptors. By contrast, in most slices, the biosensors did not measure an adenosine tone but did record a spatially non-uniform extracellular tone of the downstream metabolites (inosine and hypoxanthine). Most of the extracellular hypoxanthine arose from the metabolism of inosine by ecto-purine nucleoside phosphorylase (PNP). Adenosine kinase was the major determinant of adenosine levels, as its inhibition increased both adenosine concentration and A1 receptor-mediated synaptic inhibition. Breakdown of adenosine by adenosine deaminase was the major source of the inosine/hypoxanthine tone. However adenosine deaminase played a minor role in determining the level of adenosine at synapses, suggesting a distal location. Blockade of adenosine transport (by NBTI/dipyridamole) had inconsistent effects on basal levels of adenosine and synaptic transmission. Unexpectedly, application of NBTI/dipyridamole prevented the efflux of adenosine resulting from block of adenosine kinase at only a subset of synapses. We conclude that there is spatial variation in the functional expression of NBTI/dipyridamole-sensitive transporters. The increased spatial and temporal resolution of the purine biosensor measurements has revealed the complexity of the control of adenosine and purine tone in the cerebellum. PMID:17446223

  13. Adenosine Kinase of Trypanosoma brucei and Its Role in Susceptibility to Adenosine Antimetabolites▿ †

    PubMed Central

    Lüscher, Alexandra; Önal, Pinar; Schweingruber, Anne-Marie; Mäser, Pascal

    2007-01-01

    Trypanosoma brucei cannot synthesize purines de novo and relies on purine salvage from its hosts to build nucleic acids. With adenosine being a preferred purine source of bloodstream-form trypanosomes, adenosine kinase (AK; EC 2.7.1.20) is likely to be a key player in purine salvage. Adenosine kinase is also of high pharmacological interest, since for many adenosine antimetabolites, phosphorylation is a prerequisite for activity. Here, we cloned and functionally characterized adenosine kinase from T. brucei (TbAK). TbAK is a tandem gene, expressed in both procyclic- and bloodstream-form trypanosomes, whose product localized to the cytosol of the parasites. The RNA interference-mediated silencing of TbAK suggested that the gene is nonessential under standard growth conditions. Inhibition or downregulation of TbAK rendered the trypanosomes resistant to cordycepin (3′-deoxyadenosine), demonstrating a role for TbAK in the activation of adenosine antimetabolites. The expression of TbAK in Saccharomyces cerevisiae complemented a null mutation in the adenosine kinase gene ado1. The concomitant expression of TbAK with the T. brucei adenosine transporter gene TbAT1 allowed S. cerevisiae ado1 ade2 double mutants to grow on adenosine as the sole purine source and, at the same time, sensitized them to adenosine antimetabolites. The coexpression of TbAK and TbAT1 in S. cerevisiae ado1 ade2 double mutants proved to be a convenient tool for testing nucleoside analogues for uptake and activation by T. brucei adenosine salvage enzymes. PMID:17698621

  14. Comparative transcriptome analysis of Bacillus subtilis responding to dissolved oxygen in adenosine fermentation.

    PubMed

    Yu, Wen-Bang; Gao, Shu-Hong; Yin, Chun-Yun; Zhou, Ying; Ye, Bang-Ce

    2011-01-01

    Dissolved oxygen (DO) is an important factor for adenosine fermentation. Our previous experiments have shown that low oxygen supply in the growth period was optimal for high adenosine yield. Herein, to better understand the link between oxygen supply and adenosine productivity in B. subtilis (ATCC21616), we sought to systematically explore the effect of DO on genetic regulation and metabolism through transcriptome analysis. The microarrays representing 4,106 genes were used to study temporal transcript profiles of B. subtilis fermentation in response to high oxygen supply (agitation 700 r/min) and low oxygen supply (agitation 450 r/min). The transcriptome data analysis revealed that low oxygen supply has three major effects on metabolism: enhance carbon metabolism (glucose metabolism, pyruvate metabolism and carbon overflow), inhibit degradation of nitrogen sources (glutamate family amino acids and xanthine) and purine synthesis. Inhibition of xanthine degradation was the reason that low oxygen supply enhanced adenosine production. These provide us with potential targets, which can be modified to achieve higher adenosine yield. Expression of genes involved in energy, cell type differentiation, protein synthesis was also influenced by oxygen supply. These results provided new insights into the relationship between oxygen supply and metabolism.

  15. Comparative Transcriptome Analysis of Bacillus subtilis Responding to Dissolved Oxygen in Adenosine Fermentation

    PubMed Central

    Yin, Chun-Yun; Zhou, Ying; Ye, Bang-Ce

    2011-01-01

    Dissolved oxygen (DO) is an important factor for adenosine fermentation. Our previous experiments have shown that low oxygen supply in the growth period was optimal for high adenosine yield. Herein, to better understand the link between oxygen supply and adenosine productivity in B. subtilis (ATCC21616), we sought to systematically explore the effect of DO on genetic regulation and metabolism through transcriptome analysis. The microarrays representing 4,106 genes were used to study temporal transcript profiles of B. subtilis fermentation in response to high oxygen supply (agitation 700 r/min) and low oxygen supply (agitation 450 r/min). The transcriptome data analysis revealed that low oxygen supply has three major effects on metabolism: enhance carbon metabolism (glucose metabolism, pyruvate metabolism and carbon overflow), inhibit degradation of nitrogen sources (glutamate family amino acids and xanthine) and purine synthesis. Inhibition of xanthine degradation was the reason that low oxygen supply enhanced adenosine production. These provide us with potential targets, which can be modified to achieve higher adenosine yield. Expression of genes involved in energy, cell type differentiation, protein synthesis was also influenced by oxygen supply. These results provided new insights into the relationship between oxygen supply and metabolism. PMID:21625606

  16. Mucosal adenosine stimulates chloride secretion in canine tracheal epithelium

    SciTech Connect

    Pratt, A.D.; Clancy, G.; Welsh, M.J.

    1986-08-01

    Adenosine is a local regulator of a variety of physiological functions in many tissues and has been observed to stimulate secretion in several Cl-secreting epithelia. In canine tracheal epithelium the authors found that adenosine stimulates Cl secretion from both the mucosal and submucosal surfaces. Addition of adenosine, or its analogue 2-chloroadenosine, to the mucosal surface potently stimulated Cl secretion with no effect on the rate of Na absorption. Stimulation resulted from an interaction of adenosine with adenosine receptors, because it was blocked by the adenosine receptor blocker, 8-phenyltheophylline. The adenosine receptor was a stimulatory receptor as judged by the rank-order potency of adenosine and its analogues and by the increase in cellular adenosine 3',5'-cyclic monophosphate levels produced by 2-chloroadenosine. Adenosine also stimulated Cl secretion when it was added to the submucosal surface, although the maximal increase in secretion was less and it was much less potent. The observation that mucosal 8-phenyletheophylline blocked the effect of submucosal 2-chloroadenosine, whereas submucosal 8-phenyltheophylline did not prevent a response to mucosal or submucosal 2-chloroadenosine, suggests that adenosine receptors are located on the mucosal surface. Thus submucosal adenosine may stimulate secretion by crossing the epithelium and interacting with receptors located on the mucosal surface. Because adenosine can be released from mast cells located in the airway lumen in response to inhaled material, and because adenosine stimulated secretion from the mucosal surface, it may be in a unique position to control the epithelium on a regional level.

  17. Adenosine promotes burst activity in guinea-pig geniculocortical neurones through two different ionic mechanisms.

    PubMed Central

    Pape, H C

    1992-01-01

    1. The mechanisms of action of adenosine were examined in relay neurones of the dorsal lateral geniculate nucleus (LGND) using in vitro intracellular recording techniques in guinea-pig thalamic slices. 2. Adenosine hyperpolarized LGND relay neurones due to an increase in membrane potassium conductance. The K+ currents generated by near maximal stimulation of adenosine and GABAB receptors were non-additive. 3. Blockage of membrane K+ conductances by barium unmasked a second response to adenosine; an outward shift of the current versus voltage relationship negative to -65 mV associated with an increase in membrane input resistance. The beta-adrenoceptor agonist isoprenaline elicited an inward current in the same voltage range, which was inhibited and replaced by an outward current during activation of adenosine receptors. The effects of adenosine were due to a decrease in amplitude and rate of rise of the hyperpolarization-activated cation current, Ih. Maximal reduction by 66% of Ih amplitude occurred near the range of half-activation. 4. Both responses to adenosine were mimicked by the selective A1 receptor agonists N6-cyclopentyladenosine or N6-cyclohexyladenosine, and reversibly blocked by the selective A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). 5. The decrease in Ih by adenosine may be mediated by an inhibition of adenylyl cyclase activity and hence a decrease in the intracellular level of cyclic AMP, since local application of the adenylyl cyclase inhibitor 2',3'-dideoxyadenosine imitated the decrease in Ih. Local application of the adenylyl cyclase stimulant forskolin or 8-bromo-cyclic AMP resulted in an enhancement in Ih, and forskolin inhibited the action on Ih evoked by N6-cyclopentyladenosine. 6. The adenosine-induced effects interacted with the intrinsic electrophysiological properties of LGND neurones in that (i) the hyperpolarization due to an increase in K+ conductance inhibited single spike firing and promoted calcium

  18. Transcapillary adenosine transport and interstitial adenosine concentration in guinea pig hearts

    PubMed Central

    WANGLER, ROGER D.; GORMAN, MARK W.; WANG, C. Y.; DEWITT, DONALD F.; CHAN, I. S.; BASSINGTHWAIGHTE, JAMES B.; SPARKS, HARVEY V.

    2010-01-01

    We used the multiple-indicator-dilution technique to observe the capillary transport of adenosine in isolated Krebs-Henseleit-perfused guinea pig hearts. Tracer concentrations of radiolabeled albumin, sucrose, and adenosine were injected into the coronary inflow; outflow samples were collected for 10-25 s and analyzed by high-performance liquid chromatography (HPLC) and by γ- and β-counting. The albumin data define the intravascular transport characteristics; the sucrose data define permeation through interendothelial clefts and dilution in interstitial fluid (ISF). Parameters calculated from adenosine data include permeability-surface area products for endothelial cell uptake at the luminal and abluminal membranes and intraendothelial metabolism. We found that in situ endothelial cells avidly take up and metabolize adenosine. Tracer adenosine in the capillary lumen is twice as likely to enter an endothelial cell as it is to permeate the clefts. There was no adenosine in the arterial perfusate. Under control conditions, the steady-state venous adenosine concentration was 3.6 ± 0.8 nM, which from the flow and the parameters estimated from the tracer data gave a calculated ISF concentration of 6.8 ± 1.5 nM. During dipyridamole infusion (10 μM) at constant pressure, the cell permeabilities went essentially to zero, whereas the venous adenosine concentration increased to 44.0 ± 12.6 nM, giving an estimated ISF concentration of 191 ± 53 nM. With constant flow perfusion, venous concentration during dipyridamole infusion was 30.9 ± 6.3 nM, and estimated ISF concentration was 88 ± 20 mM. We conclude that in this preparation, at rest, the ISF adenosine concentration is about twice the venous concentration and the ISF adenosine concentration increases with dipyridamole administration. PMID:2750952

  19. Dual Effect of Adenosine A1 Receptor Activation on Renal O2 Consumption.

    PubMed

    Babich, Victor; Vadnagara, Komal; Di Sole, Francesca

    2015-12-01

    The high requirement of O2 in the renal proximal tubule stems from a high rate of Na(+) transport. Adenosine A1 receptor (A1R) activation regulates Na(+) transport in this nephron segment. Thus, the effect of the acute activation and the mechanisms of A1R on the rate of O2 consumption were evaluated. The A1R-antagonist, 8-cyclopentyl-1,3-dipropylxanthine (CPX) and adenosine deaminase (ADA), which metabolize endogenous adenosine, reduced O2 consumption (40-50%). Replacing Na(+) in the buffer reversed the ADA- or CPX-mediated reduction of O2 consumption. Blocking the Na/H-exchanger activity, which decreases O2 usage per se, did not enhance the ADA- or CPX-induced inhibition of O2 consumption. These data indicate that endogenous adenosine increases O2 usage via the activation of Na(+) transport. In the presence of endogenous adenosine, A1R was further activated by the A1R-agonist N(6)-cyclopentyladenosine (CPA); CPA inhibited O2 usage (30%) and this effect also depended on Na(+) transport. Moreover, a low concentration of CPA activated O2 usage in tissue pretreated with ADA, whereas a high concentration of CPA inhibited O2 usage; both effects depended on Na(+). Protein kinase C signaling mediated the inhibitory effect of A1R, while adenylyl cyclase mediated its stimulatory effect on O2 consumption. In summary, increasing the local concentrations of adenosine can either activate or inhibit O2 consumption via A1R, and this mechanism depends on Na(+) transport. The inhibition of O2 usage by A1R activation might restore the compromised balance between energy supply and demand under pathophysiological conditions, such as renal ischemia, which results in high adenosine production. © 2015 Wiley Periodicals, Inc.

  20. CD73+ regulatory T cells contribute to adenosine-mediated resolution of acute lung injury

    PubMed Central

    Ehrentraut, Heidi; Clambey, Eric T.; McNamee, Eoin N.; Brodsky, Kelley S.; Ehrentraut, Stefan F.; Poth, Jens M.; Riegel, Ann K.; Westrich, Joseph A.; Colgan, Sean P.; Eltzschig, Holger K.

    2013-01-01

    Acute lung injury (ALI) is characterized by alveolar injury and uncontrolled inflammation. Since most cases of ALI resolve spontaneously, understanding the endogenous mechanisms that promote ALI resolution is important to developing effective therapies. Previous studies have implicated extracellular adenosine signaling in tissue adaptation and wound healing. Therefore, we hypothesized a functional contribution for the endogenous production of adenosine during ALI resolution. As a model, we administered intratracheal LPS and observed peak lung injury at 3 d, with resolution by d 14. Treatment with pegylated adenosine-deaminase to enhance extracellular adenosine breakdown revealed impaired ALI resolution. Similarly, genetic deletion of cd73, the pacemaker for extracellular adenosine generation, was associated with increased mortality (0% wild-type and 40% in cd73−/− mice; P<0.05) and failure to resolve ALI adequately. Studies of inflammatory cell trafficking into the lungs during ALI resolution revealed that regulatory T cells (Tregs) express the highest levels of CD73. While Treg numbers in cd73−/− mice were similar to controls, cd73-deficient Tregs had attenuated immunosuppressive functions. Moreover, adoptive transfer of cd73-deficient Tregs into Rag−/− mice emulated the observed phenotype in cd73−/− mice, while transfer of wild-type Tregs was associated with normal ALI resolution. Together, these studies implicate CD73-dependent adenosine generation in Tregs in promoting ALI resolution.—Ehrentraut, H., Clambey, E. T., McNamee, E. N., Brodsky, K. S., Ehrentraut, S. F., Poth, J. M., Riegel, A. K., Westrich, J. A., Colgan, S. P., Eltzschig, H. K. CD73+ regulatory T cells contribute to adenosine-mediated resolution of acute lung injury. PMID:23413361

  1. The adenosine kinase hypothesis of epileptogenesis

    PubMed Central

    Boison, Detlev

    2008-01-01

    Current therapies for epilepsy are largely symptomatic and do not affect the underlying mechanisms of disease progression, i.e. epileptogenesis. Given the large percentage of pharmacoresistant chronic epilepsies, novel approaches are needed to understand and modify the underlying pathogenetic mechanisms. Although different types of brain injury (e.g. status epilepticus, traumatic brain injury, stroke) can trigger epileptogenesis, astrogliosis appears to be a homotypic response and hallmark of epilepsy. Indeed, recent findings indicate that epilepsy might be a disease of astrocyte dysfunction. This review focuses on the inhibitory neuromodulator and endogenous anticonvulsant adenosine, which is largely regulated by astrocytes and its key metabolic enzyme adenosine kinase (ADK). Recent findings support the “ADK hypothesis of epileptogenesis”: (i) Mouse models of epileptogenesis suggest a sequence of events leading from initial downregulation of ADK and elevation of ambient adenosine as an acute protective response, to changes in astrocytic adenosine receptor expression, to astrocyte proliferation and hypertrophy (i.e. astrogliosis), to consequential overexpression of ADK, reduced adenosine and – finally – to spontaneous focal seizure activity restricted to regions of astrogliotic overexpression of ADK. (ii) Transgenic mice overexpressing ADK display increased sensitivity to brain injury and seizures. (iii) Inhibition of ADK prevents seizures in a mouse model of pharmacoresistant epilepsy. (iv) Intrahippocampal implants of stem cells engineered to lack ADK prevent epileptogenesis. Thus, ADK emerges both as a diagnostic marker to predict, as well as a prime therapeutic target to prevent, epileptogenesis. PMID:18249058

  2. MOLECULAR PROBES FOR EXTRACELLULAR ADENOSINE RECEPTORS

    PubMed Central

    Jacobson, Kenneth A.; Ukena, Dieter; Padgett, William; Kirk, Kenneth L.; Daly, John W.

    2012-01-01

    Derivatives of adenosine receptor agonists (N6-phenyladenosines) and antagonists (1,3-dialkyl-8-phenylxanthines) bearing functionalized chains suitable for attachment to other molecules have been reported [Jacobson et al., J. med. Chem. 28, 1334 and 1341 (1985)]. The “functionalized congener” approach has been extended to the synthesis of spectroscopic and other probes for adenosine receptors that retain high affinity (Ki ~ 10−9 −10−8 M) in A1-receptor binding. The probes have been synthesized from an antagonist xanthine amine congener (XAC) and an adenosine amine congener (ADAC). [3H]ADAC has been synthesized and found to bind highly specifically to A1-adenosine receptors of rat and calf cerebral cortical membranes with KD values of 1.4 and 0.34 nM respectively. The higher affinity in the bovine brain, seen also with many of the probes derived from ADAC and XAC, is associated with phenyl substituents. The spectroscopic probes contain a reporter group attached at a distal site of the functionalized chain. These bifunctional ligands may contain a spin label (e.g. the nitroxyl radical TEMPO) for electron spin resonance spectroscopy, or a fluorescent dye, including fluorescein and 4-nitrobenz-2-oxa-1,3-diazole (NBD), or labels for 19F nuclear magnetic resonance spectroscopy. Potential applications of the spectroscopic probes in characterization of adenosine receptors are discussed. PMID:3036153

  3. Radioimmunochemical quantitation of human adenosine deaminase.

    PubMed Central

    Daddona, P E; Frohman, M A; Kelley, W N

    1979-01-01

    Markedly reduced or absent adenosine deaminase activity in man is associated with an autosomal recesive form of severe conbined immunodeficiency disease. To further define the genetic nature of this enzyme defect, we have quantitated immunologically active adenosine deaminase (CRM) in the hemolysate of homozygous deficient patients and their heterozygous parents. A highly specific radioimmunoassay was developed capable of detecting 0.05% of normal erythrocyte adenosine deaminase. Hemolysates from nine heterozygotes (five families) showed a wide range in CRM (32--100% of normal) and variable absolute specific activities with several being at least 1 SD BELOW THE NORMAL MEAN. Hemolysates from four unrelated patients showed less than 0.09% adenosine deaminase activity with CRM ranging from less than 0.06 to 5.6% of the normal mean. In conclusion, heterozygote and homozygote hemolysates from five of the eight families analyzed revealed variable levels of CRM suggesting heterogeneous genetic alteration or expression of the silent or defective allele(s) of adenosine deaminase. PMID:468994

  4. Adenosine receptor targeting in health and disease.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Fazzi, Debora; Stefanelli, Angela; Varani, Katia; Borea, Pier Andrea

    2011-12-01

    The adenosine receptors A(1), A(2A), A(2B) and A(3) are important and ubiquitous mediators of cellular signaling that play vital roles in protecting tissues and organs from damage. In particular, adenosine triggers tissue protection and repair by different receptor-mediated mechanisms, including increasing the oxygen supply:demand ratio, pre-conditioning, anti-inflammatory effects and the stimulation of angiogenesis. The state of the art of the role of adenosine receptors which have been proposed as targets for drug design and discovery, in health and disease, and an overview of the ligands for these receptors in clinical development. Selective ligands of A(1), A(2A), A(2B) and A(3) adenosine receptors are likely to find applications in the treatment of pain, ischemic conditions, glaucoma, asthma, arthritis, cancer and other disorders in which inflammation is a feature. The aim of this review is to provide an overview of the present knowledge regarding the role of these adenosine receptors in health and disease.

  5. Caffeine, adenosine receptors, and synaptic plasticity.

    PubMed

    Costenla, Ana Rita; Cunha, Rodrigo A; de Mendonça, Alexandre

    2010-01-01

    Few studies to date have looked at the effects of caffeine on synaptic plasticity, and those that did used very high concentrations of caffeine, whereas the brain concentrations attained by regular coffee consumption in humans should be in the low micromolar range, where caffeine exerts pharmacological actions mainly by antagonizing adenosine receptors. Accordingly, rats drinking caffeine (1 g/L) for 3 weeks, displayed a concentration of caffeine of circa 22 microM in the hippocampus. It is known that selective adenosine A1 receptor antagonists facilitate, whereas selective adenosine A2A receptor antagonists attenuate, long term potentiation (LTP) in the hippocampus. Although caffeine is a non-selective antagonist of adenosine receptors, it attenuates frequency-induced LTP in hippocampal slices in a manner similar to selective adenosine A2A receptor antagonists. These effects of low micromolar concentration of caffeine (30 microM) are maintained in aged animals, which is important when a possible beneficial effect for caffeine in age-related cognitive decline is proposed. Future studies will still be required to confirm and detail the involvement of A1 and A2A receptors in the effects of caffeine on hippocampal synaptic plasticity, using both pharmacological and genetic approaches.

  6. Differential A1 adenosine receptor reserve for two actions of adenosine on guinea pig atrial myocytes.

    PubMed

    Srinivas, M; Shryock, J C; Dennis, D M; Baker, S P; Belardinelli, L

    1997-10-01

    Adenosine activates adenosine-induced inwardly rectifying K+ current (IKAdo) and inhibits isoproterenol (100 nM)-stimulated L-type Ca2+ current (beta-ICa,L) of guinea pig atrial myocytes with EC50 values of 2.17 and 0.20 microM, respectively. We determined whether this 11-fold difference in potency of adenosine is due to the existence of a greater A1 adenosine receptor reserve for the inhibition of beta-ICa,L than for the activation of IKAdo. Atrial myocytes were pretreated with vehicle (control) or the irreversible A1 adenosine receptor antagonist 8-cyclopentyl-3-[3-[[4-(fluorosulfonyl)benzoyl]oxy]propyl]-1-propylxa nthine (FSCPX) (10 and 50 nM) for 30 min, and after a 60-min washout period, concentration-response curves were determined for the adenosine-induced activation of IKAdo and inhibition of beta-ICa,L. Pretreatment of atrial myocytes with 10 nM FSCPX reduced the maximal activation of IKAdo by 60% (7.9 +/- 0.2 to 3.2 +/- 0.1 pA/pF). In contrast, a higher concentration of FSCPX (50 nM) was required to reduce the maximal inhibition of beta-ICa,L by 39% (95 +/- 4% to 58. 7 +/- 5.6%) and caused a 15-fold increase in the EC50 value of adenosine. Values of the equilibrium dissociation constant (KA) for adenosine to activate IKAdo and inhibit beta-ICa,L, estimated according to the method of Furchgott, were 2.7 and 5.6 microM, respectively. These values were used to determine the relationship between adenosine receptor occupancy and response. Half-maximal and maximal activations of IKAdo required occupancies of 40% and 98% of A1 adenosine receptors, respectively. In contrast, occupancies of only 4% and 70%, respectively, of A1 adenosine receptors were sufficient to cause half-maximal and maximal inhibitions of beta-ICa, L. Consistent with this result, a partial agonist of the A1 adenosine receptor SHA040 inhibited beta-ICa,L by 60 +/- 3.5% but activated IKAdo by only 18.1 +/- 2.5%. The results indicate that the A1 adenosine receptor is coupled more efficiently to

  7. Role of adenosine signaling in penile erection and erectile disorders.

    PubMed

    Phatarpekar, Prasad V; Wen, Jiaming; Xia, Yang

    2010-11-01

    Penile erection is a hemodynamic process, which results from increased flow and retention of blood in the penile organ due to the relaxation of smooth muscle cells. Adenosine, a physiological vasorelaxant, has been shown to be a modulator of penile erection. To summarize the research on the role of adenosine signaling in normal penile erection and erectile disorders. Evidence in the literature on the association between adenosine signaling and normal and abnormal penile erection, i.e., erectile dysfunction (ED) and priapism. The article reviews the literature on the role of endogenous and exogenous adenosine in normal penile erection, as well as in erectile disorders namely, ED and priapism. Adenosine has been shown to relax corpus cavernosum from various species including human in both in vivo and in vitro studies. Neuromodulatory role of adenosine in corpus cavernosum has also been demonstrated. Impaired adenosine signaling through A(2B) receptor causes partial resistance of corpus cavernosum, from men with organic ED, to adenosine-mediated relaxation. Increased level of adenosine has been shown to be a causative factor for priapism. Overall, the research reviewed here suggests a general role of exogenous and endogenous adenosine signaling in normal penile erection. From this perspective, it is not surprising that impaired adenosine signaling is associated with ED, and excessive adenosine signaling is associated with priapism. Adenosine signaling represents a potentially important diagnostic and therapeutic target for the treatment of ED and priapism. © 2009 International Society for Sexual Medicine.

  8. Role of Adenosine Signaling in Penile Erection and Erectile Disorders

    PubMed Central

    Phatarpekar, Prasad V.; Wen, Jiaming; Xia, Yang

    2010-01-01

    Introduction Penile erection is a hemodynamic process, which results from increased flow and retention of blood in the penile organ due to the relaxation of smooth muscle cells. Adenosine, a physiological vasorelaxant, has been shown to be a modulator of penile erection. Aim To summarize the research on the role of adenosine signaling in normal penile erection and erectile disorders. Main Outcome Measures Evidence in the literature on the association between adenosine signaling and normal and abnormal penile erection, i.e., erectile dysfunction (ED) and priapism. Methods The article reviews the literature on the role of endogenous and exogenous adenosine in normal penile erection, as well as in erectile disorders namely, ED and priapism. Results Adenosine has been shown to relax corpus cavernosum from various species including human in both in vivo and in vitro studies. Neuromodulatory role of adenosine in corpus cavernosum has also been demonstrated. Impaired adenosine signaling through A2B receptor causes partial resistance of corpus cavernosum, from men with organic ED, to adenosine-mediated relaxation. Increased level of adenosine has been shown to be a causative factor for priapism. Conclusion Overall, the research reviewed here suggests a general role of exogenous and endogenous adenosine signaling in normal penile erection. From this perspective, it is not surprising that impaired adenosine signaling is associated with ED, and excessive adenosine signaling is associated with priapism. Adenosine signaling represents a potentially important diagnostic and therapeutic target for the treatment of ED and priapism. PMID:19889148

  9. Feasibility and safety of adenosine cardiovascular magnetic resonance in patients with MR conditional pacemaker systems at 1.5 Tesla.

    PubMed

    Klein-Wiele, Oliver; Garmer, Marietta; Urbien, Rhyan; Busch, Martin; Kara, Kaffer; Mateiescu, Serban; Grönemeyer, Dietrich; Schulte-Hermes, Michael; Garbrecht, Marc; Hailer, Birgit

    2015-12-22

    Cardiovascular Magnetic Resonance (CMR) with adenosine stress is a valuable diagnostic tool in coronary artery disease (CAD). However, despite the development of MR conditional pacemakers CMR is not yet established in clinical routine for pacemaker patients with known or suspected CAD. A possible reason is that adenosine stress perfusion for ischemia detection in CMR has not been studied in patients with cardiac conduction disease requiring pacemaker therapy. Other than under resting conditions it is unclear whether MR safe pacing modes (paused pacing or asynchronous mode) can be applied safely because the effect of adenosine on heart rate is not precisely known in this entity of patients. We investigate for the first time feasibility and safety of adenosine stress CMR in pacemaker patients in clinical routine and evaluate a pacing protocol that considers heart rate changes under adenosine. We retrospectively analyzed CMR scans of 24 consecutive patients with MR conditional pacemakers (mean age 72.1 ± 11.0 years) who underwent CMR in clinical routine for the evaluation of known or suspected CAD. MR protocol included cine imaging, adenosine stress perfusion and late gadolinium enhancement. Pacemaker indications were sinus node dysfunction (n = 18) and second or third degree AV block (n = 6). Under a pacing protocol intended to avoid competitive pacing on the one hand and bradycardia due to AV block on the other no arrhythmia occurred. Pacemaker stimulation was paused to prevent competitive pacing in sinus node dysfunction with resting heart rate >45 bpm. Sympatho-excitatory effect of adenosine led to a significant acceleration of heart rate by 12.3 ± 8.3 bpm (p < 0.001), no bradycardia occurred. On the contrary in AV block heart rate remained constant; asynchronous pacing above resting heart rate did not interfere with intrinsic rhythm. Adenosine stress CMR appears to be feasible and safe in patients with MR conditional pacemakers. Heart rate response to adenosine

  10. Binding induced colocalization activated hybridization chain reaction on the surface of magnetic nanobead for sensitive detection of adenosine.

    PubMed

    Feng, Chunjing; Hou, Zhun; Jiang, Wei; Sang, Lihong; Wang, Lei

    2016-12-15

    Herein, a sensitive and enzyme-free assay for adenosine detection has been developed on the basis of binding induced colocalization activated hybridization chain reaction (HCR) strategy on the surface of magnetic nanobead. First, the recognition probe was fabricated and divided into two parts: the Apt-1 that composed a part of adenosine aptamer and toehold domain, and the Apt-2 that consisted of another part of adenosine aptamer and branch migration domain. The Apt-1 was immobilized on a streptavidin-magnetic nanobead (streptavidin-MNBs) that played the roles of enrichment and separation. Then the recognition event of adenosine could bring the two parts of aptamer together and induce the colocalization of toehold domain and branch migration domain, which could serve as an integrated initiator to trigger the HCR, producing a long nicked double-stranded polymer. Finally, the intercalating dye SYBR Green I was inserted into the polymer, generating an enhanced fluorescence signal. In this strategy, the initiator was divided into two parts and could be suppressed effectively in the absence of adenosine. Utilizing the separated function, the spontaneous hybridization of H1 and H2 could be avoided, and a low background could be acquired. Moreover, through the double amplification of HCR and multimolecules binding of SYBR Green I, highly sensitive and enzyme-free detection were achieved. The detection limit for adenosine detection was 2.0×10(-7)mol/L, which was comparable or superior to the previous aptasensors. Importantly, adenosine analysis in human urines has been performed, and this strategy could significantly distinguish the adenosine content in normal human urines and cancer patient urines, suggesting that this proposed assay will become a reliable and sensitive adenosine detection method in early clinical diagnosis and medical research.

  11. Adenosine augments IL-10-induced STAT3 signaling in M2c macrophages.

    PubMed

    Koscsó, Balázs; Csóka, Balázs; Kókai, Endre; Németh, Zoltán H; Pacher, Pál; Virág, László; Leibovich, S Joseph; Haskó, György

    2013-12-01

    The alternatively activated macrophage phenotype induced by IL-10 is called M2c. Adenosine is an endogenous purine nucleoside that accumulates in the extracellular space in response to metabolic disturbances, hypoxia, inflammation, physical damage, or apoptosis. As adenosine is known to regulate classically activated M1 and IL4- and IL-13-activated M2a macrophages, the goal of the present study was to explore its effects on M2c macrophages. We found that adenosine augmented the IL-10-induced expression of TIMP-1 and arginase-1 by the mouse macrophage cell line RAW 264.7 and by mouse BMDMs. The effects of AR stimulation on IL-10-induced TIMP-1 or arginase-1 expression were lacking in A2BAR KO macrophages. The role of A2BAR on TIMP-1 production of RAW 264.7 cells was confirmed with specific agonist BAY606583 and antagonist PSB0788. AR stimulation augmented IL-10-induced STAT3 phosphorylation in macrophages, and pharmacological inhibition or silencing of STAT3 using siRNA reduced the stimulatory effect of AR stimulation on TIMP-1 production. In contrast to its stimulatory effect on IL-10-induced STAT3 activation, adenosine inhibited IL-6-induced STAT3 phosphorylation and SAA3 expression. In conclusion, adenosine enhances IL-10-induced STAT3 signaling and M2c macrophage activation.

  12. The A2a adenosine receptor modulates the reinforcement efficacy and neurotoxicity of MDMA.

    PubMed

    Ruiz-Medina, Jessica; Ledent, Catherine; Carretón, Olga; Valverde, Olga

    2011-04-01

    Adenosine is an endogenous purine nucleoside that plays a neuromodulatory role in the central nervous system. A2a adenosine receptors have been involved in reward-related processes, inflammatory phenomena and neurotoxicity reactions. In the present study, we investigated the role of A2a adenosine receptors on the acute pharmacological effects, reinforcement and neuroinflammation induced by MDMA administration. First, the acute effects of MDMA on body temperature, locomotor activity and anxiety-like responses were measured in A2a knockout mice and wild-type littermates. Second, MDMA reinforcing properties were evaluated using the intravenous self-administration paradigm. Finally, we assessed striatal astrogliosis and microgliosis as markers of MDMA neurotoxicity. Our results showed that acute MDMA produced a biphasic effect on body temperature and increased locomotor activity and anxiogenic-like responses in both genotypes. However, MDMA reinforcing properties were dramatically affected by the lack of A2a adenosine receptors. Thus, wild-type mice maintained MDMA self-administration under a fixed ratio 1 reinforcement schedule, whereas the operant response appeared completely abolished in A2a knockout mice. In addition, the MDMA neurotoxic regime produced an enhanced inflammatory response in striatum of wild-type mice, revealed by a significant increase in glial expression, whereas such activation was attenuated in mutant mice. This is the first report indicating that A2a adenosine receptors play a key role in reinforcement and neuroinflammation induced by the widely used psychostimulant.

  13. CD73-adenosine: a next-generation target in immuno-oncology.

    PubMed

    Allard, David; Allard, Bertrand; Gaudreau, Pierre-Olivier; Chrobak, Pavel; Stagg, John

    2016-02-01

    Cancer immunotherapy has entered in a new era with the development of first-generation immune checkpoint inhibitors targeting the PD1/PD-L1 and CTLA-4 pathways. In this context, considerable research effort is being deployed to find the next generation of cancer immunotherapeutics. The CD73-adenosine axis constitutes one of the most promising pathways in immuno-oncology. We and others have demonstrated the immunosuppressive role of CD73-adenosine in cancer and established proof-of-concept that the targeted blockade of CD73 or adenosine receptors could effectively promote anti-tumor immunity and enhance the activity of first-generation immune checkpoint blockers. With Phase I clinical trials now underway evaluating anti-CD73 or anti-A2A therapies in cancer patients, we here discuss the fundamental, preclinical and clinical findings related to the role of the CD73-adenosinergic pathway in tumor immunity.

  14. SERS detection of low-concentration adenosine by silver nanoparticles on silicon nanoporous pyramid arrays structure

    NASA Astrophysics Data System (ADS)

    Zhang, C.; Man, B. Y.; Jiang, S. Z.; Yang, C.; Liu, M.; Chen, C. S.; Xu, S. C.; Qiu, H. W.; Li, Z.

    2015-08-01

    A novel surface-enhanced Raman spectroscopy (SERS) substrate based on uniform silver nanoparticles/silicon nanoporous pyramid arrays (Ag/PS) is prepared and SERS behaviors to adenosine are discussed and compared. With a low concentration of 10-7 M, the characteristic Raman bands of adenosine demonstrate the significantly high SERS sensitivity of the prepared Ag/PS substrate. A reasonable linear correlation is obtained between the intensity of SERS signal and the adenosine concentration from 10-2 to 10-7M in log scale. These results imply that the Ag/PS with regular pyramids array might be an effective substrate for performing label-free sensitive SERS detections of biomolecule.

  15. Adenine and adenosine salvage in Leishmania donovani.

    PubMed

    Boitz, Jan M; Ullman, Buddy

    2013-08-01

    6-aminopurine metabolism in Leishmania is unique among trypanosomatid pathogens since this genus expresses two distinct routes for adenine salvage: adenine phosphoribosyltransferase (APRT) and adenine deaminase (AAH). To evaluate the relative contributions of APRT and AAH, adenine salvage was evaluated in Δaprt, Δaah, and Δaprt/Δaah null mutants of L. donovani. The data confirm that AAH plays the dominant role in adenine metabolism in L. donovani, although either enzyme alone is sufficient for salvage. Adenosine salvage was also evaluated in a cohort of null mutants. Adenosine is also primarily converted to hypoxanthine, either intracellularly or extracellularly, but can also be phosphorylated to the nucleotide level by adenosine kinase when the predominant pathways are genetically or pharmacologically blocked. These data provide genetic verification for the relative contributions of 6-aminopurine metabolizing pathways in L. donovani and demonstrate that all of the pathways can function under appropriate conditions of genetic or pharmacologic perturbation.

  16. Human adenosine deaminase. Distribution and properties.

    PubMed

    Van der Weyden, M B; Kelley, W N

    1976-09-25

    Adenosine deaminase exists in multiple molecular forms in human tissue. One form of the enzyme appears to be "particulate". Three forms of the enzyme are soluble and interconvertible with apparent molecular weights of approximately 36,000, 114,000, and 298,000 (designated small, intermediate, and large, respectively). The small form of adenosine deaminase is convertible to the large form only in the presence of a protein, which has an apparent molecular weight of 200,000 and has no adenosine deaminase activity. This conversion of the small form of the enzyme to the large form occurs at 4 degrees, exhibits a pH optimum of 5.0 to 8.0, and is associated with a loss of conversion activity. The small form of the enzyme predominates in tissue preparations exhibiting the higher enzyme-specific activities and no detectable conversion activity. The large form of adenosine deaminase predominates in tissue extracts exhibiting the lower enzyme specific activities and abundant conversion activity. The small form of adenosine deaminase shows several electrophoretic variants by isoelectric focusing. The electrophoretic heterogeneity observed with the large form of the enzyme is similar to that observed with the small form, with the exception that several additional electrophoretic variants are uniformly identified. No organ specificity is demonstrable for the different electrophoretic forms. The kinetic characteristics of the three soluble molecular species of adenosine deaminase are identical except for pH optimum, which is 5.5 for the intermediate species and 7.0 to 7.4 for the large and small forms.

  17. A new class of adenosine receptors in brain: Characterization by 2-chloro( sup 3 H)adenosine binding

    SciTech Connect

    Chin, Jerome Hsicheng.

    1988-01-01

    Considerable evidence has accumulated in recent years to support a role for adenosine as an important physiological modulator in many mammalian tissues. In brain, adenosine is a potent depressant of neuronal firing and synaptic transmission. The exact mechanisms by which adenosine analogs depress nerve cell activity in the brain are not clear. Despite considerable investigation, neither the A1 nor the A2 adenosine receptors associated with adenylate cyclase have been able to account adequately for the actions of adenosine in brain. It has been proposed that additional adenosine receptors, possibly linked to calcium channels, are present in the central nervous system and are responsible for the physiological actions of adenosine. In this thesis, evidence is provided for the existence of a novel class of adenosine receptors in rat brain. The methods used to identify this new class of receptors involved radioligand binding techniques which have been successfully employed to characterize the properties of many neurotransmitter and drug receptors. 2-Chloro({sup 3}H)adenosine (Cl({sup 3}H)Ado) was selected as the ligand for these experiments since is a water-soluble, metabolically-stable analog of adenosine and a potent depressant of synaptic transmission in brain. The results demonstrate the presence of a distinct class of 2-chloro({sup 3}H)adenosine binding sites in rat forebrain membranes with an apparent K{sub D} of about 10 {mu}M and a B{sub max} of about 60 pmol per mg of protein. Specific 2-chloro ({sup 3}H)adenosine binding is highly specific for adenosine agonists and antagonists. Inhibition of binding by adenosine agonists exhibits an order of potency 2-chloroadenosine > 5{prime}-N-ethylcarboxamide adenosine > ({minus})-N{sup 6}-(R-phenylisopropyl)adenosine, which differs from that of both A1 and A2 adenosine receptors.

  18. Pain-relieving prospects for adenosine receptors and ectonucleotidases

    PubMed Central

    Zylka, Mark J.

    2010-01-01

    Adenosine receptor agonists have potent antinociceptive effects in diverse preclinical models of chronic pain. In contrast, the efficacy of adenosine or adenosine receptor agonists at treating pain in humans is unclear. Two ectonucleotidases that generate adenosine in nociceptive neurons were recently identified. When injected spinally, these enzymes have long-lasting adenosine A1 receptor (A1R)-dependent antinociceptive effects in inflammatory and neuropathic pain models. Furthermore, recent findings indicate that spinal adenosine A2A receptor activation can enduringly inhibit neuropathic pain symptoms. Collectively, these studies suggest the possibility of treating chronic pain in humans by targeting specific adenosine receptor subtypes in anatomically defined regions with agonists or with ectonucleotidases that generate adenosine. PMID:21236731

  19. Regulation of adenosine transport by acute and chronic ethanol exposure

    SciTech Connect

    Nagy, L.E.; Casso, D.; Diamond, I.; Gordon, A.S. )

    1989-02-09

    Chronic exposure to ethanol results in a desensitization of adenosine receptor-stimulated cAMP production. Since adenosine is released by cells and is known to desensitize its own as well as other receptors, it may be involved in ethanol-induced desensitization of adenosine receptor function. Therefore, we have examine the acute and chronic effects of ethanol on the transport of adenosine via the nucleoside transport. Acute exposure to ethanol caused an inhibition of adenosine uptake in S49 lymphoma cells. This decrease in uptake resulted in accumulation of extracellular adenosine after ethanol exposure. The effect of ethanol was specific to nucleoside transport. Uptake of uridine, also transported by the nucleoside transporter, was inhibited by ethanol to the same degree as adenosine uptake, while neither isoleucine nor deoxyglucose uptake was altered by ethanol treatment. Inhibition of adenosine uptake by ethanol was non-competitive and dependent on the concentration of ethanol. After chronic exposure to ethanol, cells became tolerant to the acute effects of ethanol. There was no longer an acute inhibition of adenosine uptake, nor was these accumulation of extracellular adenosine. Chronic ethanol exposure also resulted in a decrease in the absolute rate of adenosine uptake. Binding studies using a high affinity lignad for the nucleoside transporter, nitrobenzylthioinosine (NBMPR), indicate that this decreased uptake was due to a decrease in the maximal number of binding sites. These ethanol-induced changes in adenosine transport may be important for the acute and chronic effects of ethanol.

  20. The role of adenosine in Alzheimer's disease.

    PubMed

    Rahman, Anisur

    2009-09-01

    Alzheimer's disease (AD) is a neurodegenerative disorder of the central nervous system manifested by cognitive and memory deterioration, a variety of neuropsychiatric symptoms, behavioral disturbances, and progressive impairment of daily life activities. Current pharmacotherapies are restricted to symptomatic interventions but do not prevent progressive neuronal degeneration. Therefore, new therapeutic strategies are needed to intervene with these progressive pathological processes. In the past several years adenosine, a ubiquitously released purine ribonucleoside, has become important for its neuromodulating capability and its emerging positive experimental effects in neurodegenerative diseases. Recent research suggests that adenosine receptors play important roles in the modulation of cognitive function. The present paper attempts to review published reports and data from different studies showing the evidence of a relationship between adenosinergic function and AD-related cognitive deficits. Epidemiological studies have found an association between coffee (a nonselective adenosine receptor antagonist) consumption and improved cognitive function in AD patients and in the elderly. Long-term administration of caffeine in transgenic animal models showed a reduced amyloid burden in brain with better cognitive performance. Antagonists of adenosine A2A receptors mimic these beneficial effects of caffeine on cognitive function. Neuronal cell cultures with amyloid beta in the presence of an A2A receptor antagonist completely prevented amyloid beta-induced neurotoxicity. These findings suggest that the adenosinergic system constitutes a new therapeutic target for AD, and caffeine and A2A receptor antagonists may have promise to manage cognitive dysfunction in AD.

  1. A Novel Method for Screening Adenosine Receptor Specific Agonists for Use in Adenosine Drug Development

    PubMed Central

    Jones, Karlie R.; Choi, Uimook; Gao, Ji-Liang; Thompson, Robert D.; Rodman, Larry E.; Malech, Harry L.; Kang, Elizabeth M.

    2017-01-01

    Agonists that target the A1, A2A, A2B and A3 adenosine receptors have potential to be potent treatment options for a number of diseases, including autoimmune diseases, cardiovascular disease and cancer. Because each of these adenosine receptors plays a distinct role throughout the body, obtaining highly specific receptor agonists is essential. Of these receptors, the adenosine A2AR and A2BR share many sequence and structural similarities but highly differ in their responses to inflammatory stimuli. Our laboratory, using a combination of specially developed cell lines and calcium release analysis hardware, has created a new and faster method for determining specificity of synthetic adenosine agonist compounds for the A2A and A2B receptors in human cells. A2A receptor expression was effectively removed from K562 cells, resulting in the development of a distinct null line. Using HIV-lentivector and plasmid DNA transfection, we also developed A2A and A2B receptor over-expressing lines. As adenosine is known to cause changes in intracellular calcium levels upon addition to cell culture, calcium release can be determined in these cell lines upon compound addition, providing a functional readout of receptor activation and allowing us to isolate the most specific adenosine agonist compounds. PMID:28317879

  2. Adenosine receptors and the central nervous system.

    PubMed

    Sebastião, Ana M; Ribeiro, Joaquim A

    2009-01-01

    The adenosine receptors (ARs) in the nervous system act as a kind of "go-between" to regulate the release of neurotransmitters (this includes all known neurotransmitters) and the action of neuromodulators (e.g., neuropeptides, neurotrophic factors). Receptor-receptor interactions and AR-transporter interplay occur as part of the adenosine's attempt to control synaptic transmission. A(2A)ARs are more abundant in the striatum and A(1)ARs in the hippocampus, but both receptors interfere with the efficiency and plasticity-regulated synaptic transmission in most brain areas. The omnipresence of adenosine and A(2A) and A(1) ARs in all nervous system cells (neurons and glia), together with the intensive release of adenosine following insults, makes adenosine a kind of "maestro" of the tripartite synapse in the homeostatic coordination of the brain function. Under physiological conditions, both A(2A) and A(1) ARs play an important role in sleep and arousal, cognition, memory and learning, whereas under pathological conditions (e.g., Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, stroke, epilepsy, drug addiction, pain, schizophrenia, depression), ARs operate a time/circumstance window where in some circumstances A(1)AR agonists may predominate as early neuroprotectors, and in other circumstances A(2A)AR antagonists may alter the outcomes of some of the pathological deficiencies. In some circumstances, and depending on the therapeutic window, the use of A(2A)AR agonists may be initially beneficial; however, at later time points, the use of A(2A)AR antagonists proved beneficial in several pathologies. Since selective ligands for A(1) and A(2A) ARs are now entering clinical trials, the time has come to determine the role of these receptors in neurological and psychiatric diseases and identify therapies that will alter the outcomes of these diseases, therefore providing a hopeful future for the patients who suffer from these diseases.

  3. Erythrocytes retain hypoxic adenosine response for faster acclimatization upon re-ascent

    PubMed Central

    Song, Anren; Zhang, Yujin; Han, Leng; Yegutkin, Gennady G.; Liu, Hong; Sun, Kaiqi; D'Alessandro, Angelo; Li, Jessica; Karmouty-Quintana, Harry; Iriyama, Takayuki; Weng, Tingting; Zhao, Shushan; Wang, Wei; Wu, Hongyu; Nemkov, Travis; Subudhi, Andrew W.; Jameson-Van Houten, Sonja; Julian, Colleen G.; Lovering, Andrew T.; Hansen, Kirk C.; Zhang, Hong; Bogdanov, Mikhail; Dowhan, William; Jin, Jianping; Kellems, Rodney E.; Eltzschig, Holger K.; Blackburn, Michael; Roach, Robert C.; Xia, Yang

    2017-01-01

    Faster acclimatization to high altitude upon re-ascent is seen in humans; however, the molecular basis for this enhanced adaptive response is unknown. We report that in healthy lowlanders, plasma adenosine levels are rapidly induced by initial ascent to high altitude and achieved even higher levels upon re-ascent, a feature that is positively associated with quicker acclimatization. Erythrocyte equilibrative nucleoside transporter 1 (eENT1) levels are reduced in humans at high altitude and in mice under hypoxia. eENT1 deletion allows rapid accumulation of plasma adenosine to counteract hypoxic tissue damage in mice. Adenosine signalling via erythrocyte ADORA2B induces PKA phosphorylation, ubiquitination and proteasomal degradation of eENT1. Reduced eENT1 resulting from initial hypoxia is maintained upon re-ascent in humans or re-exposure to hypoxia in mice and accounts for erythrocyte hypoxic memory and faster acclimatization. Our findings suggest that targeting identified purinergic-signalling network would enhance the hypoxia adenosine response to counteract hypoxia-induced maladaptation. PMID:28169986

  4. Multiple pathways for elevating extracellular adenosine in the rat hippocampal CA1 region characterized by adenosine sensor cells.

    PubMed

    Yamashiro, Kunihiko; Fujii, Yuki; Maekawa, Shohei; Morita, Mitsuhiro

    2017-01-01

    Extracellular adenosine in the brain, which modulates various physiological and pathological processes, fluctuates in a complicated manner that reflects the circadian cycle, neuronal activity, metabolism, and disease states. The dynamics of extracellular adenosine in the brain are not fully understood, largely because of the lack of simple and reliable methods of measuring time-dependent changes in tissue adenosine distribution. This study describes the development of a biosensor, designated an adenosine sensor cell, expressing adenosine A1 receptor, and a genetically modified G protein. This biosensor was used to characterize extracellular adenosine elevation in brain tissue by measuring intracellular calcium elevation in response to adenosine. Placement of adenosine sensor cells below hippocampal slices successfully detected adenosine releases from these slices in response to neuronal activity and astrocyte swelling by conventional calcium imaging. Pharmacological analyses indicated that high-frequency electrical stimulation-induced post-synaptic adenosine release in a manner dependent on L-type calcium channels and calcium-induced calcium release. Adenosine release following treatments that cause astrocyte swelling is independent of calcium channels, but dependent on aquaporin 4, an astrocyte-specific water channel subtype. The ability of ectonucleotidase inhibitors to inhibit adenosine release following astrocyte swelling, but not electrical stimulation, suggests that the former reflects astrocytic ATP release and subsequent enzymatic breakdown, whereas the latter reflects direct adenosine release from neurons. These results suggest that distinct mechanisms are responsible for extracellular adenosine elevations by neurons and astrocytes, allowing exquisite regulation of extracellular adenosine in the brain. © 2016 International Society for Neurochemistry.

  5. Effects of adenosine infusion into renal interstitium on renal hemodynamics

    SciTech Connect

    Pawlowska, D.; Granger, J.P.; Knox, F.G.

    1987-04-01

    This study was designed to investigate the hemodynamic effects of exogenous adenosine in the interstitium of the rat kidney. Adenosine or its analogues were infused into the renal interstitium by means of chronically implanted capsules. In fusion of adenosine decreased glomerular filtration rate (GFR) from 0.81 +/- 0.06 to 0.37 +/- 0.06 ml/min while having no effect on renal blood flow (RBF). The metabolically stable analogue, 2-chloradenosine (2-ClAdo), decreased GFR from 0.73 +/- 0.07 to 021 +/- 0.06 ml/min. Interstitial infusion of theophylline, an adenosine receptor antagonist, completely abolished the effects of adenosine and 2-ClAdo on GFR. The distribution of adenosine, when infused into the renal interstitium, was determined using radiolabeled 5'-(N-ethyl)-carboxamidoadenosine (NECA), a metabolically stable adenosine agonist. After continuous infusion, (/sup 3/H)NECA was distributed throughout the kidney. The effects of NECA to reduce GFR were similar to those of adenosine and 2-ClAdo. They conclude that increased levels of adenosine in the renal interstitium markedly decrease GFR without affecting RBF in steady-state conditions. The marked effects of adenosine agonists during their infusion into the renal interstitium and the complete blockade of these effects by theophylline suggest an extracellular action of adenosine.

  6. Neuroprotective effects of adenosine deaminase in the striatum

    PubMed Central

    Tamura, Risa; Satoh, Yasushi; Nonoyama, Shigeaki; Nishida, Yasuhiro; Nibuya, Masashi

    2016-01-01

    Adenosine deaminase (ADA) is a ubiquitous enzyme that catabolizes adenosine and deoxyadenosine. During cerebral ischemia, extracellular adenosine levels increase acutely and adenosine deaminase catabolizes the increased levels of adenosine. Since adenosine is a known neuroprotective agent, adenosine deaminase was thought to have a negative effect during ischemia. In this study, however, we demonstrate that adenosine deaminase has substantial neuroprotective effects in the striatum, which is especially vulnerable during cerebral ischemia. We used temporary oxygen/glucose deprivation (OGD) to simulate ischemia in rat corticostriatal brain slices. We used field potentials as the primary measure of neuronal damage. For stable and efficient electrophysiological assessment, we used transgenic rats expressing channelrhodopsin-2, which depolarizes neurons in response to blue light. Time courses of electrically evoked striatal field potential (eFP) and optogenetically evoked striatal field potential (optFP) were recorded during and after oxygen/glucose deprivation. The levels of both eFP and optFP decreased after 10 min of oxygen/glucose deprivation. Bath-application of 10 µg/ml adenosine deaminase during oxygen/glucose deprivation significantly attenuated the oxygen/glucose deprivation-induced reduction in levels of eFP and optFP. The number of injured cells decreased significantly, and western blot analysis indicated a significant decrease of autophagic signaling in the adenosine deaminase-treated oxygen/glucose deprivation slices. These results indicate that adenosine deaminase has protective effects in the striatum. PMID:26746865

  7. Ecto-enzyme activity of human erythrocyte adenosine deaminase.

    PubMed

    Bielat, K; Tritsch, G L

    1989-04-11

    Adenosine deaminase is found primarily in the cytoplasm of many cell types. In the human erythrocyte, about 30 per cent of the total adenosine deaminase activity is membrane associated, and about two-thirds of this is inactivated by treatment of intact erythrocytes with the nonpenetrating reagent diazotized sulfanilic acid, without affecting lactate dehydrogenase, a soluble cytoplasmic enzyme. This indicates that within the cell membranes, the catalytic site of about two-thirds of the adenosine deaminase faces the external medium, i.e., ecto adenosine deaminase. Localization of adenosine deaminase activity at the cell membrane is demonstrated directly by electron microscopy by use of the substrate 6-Chloropurine ribonucleoside, which is dechlorinated by adenosine deaminase to produce Cl-, which is precipitated at its locus of formation by added Ag+, and the precipitated AgCl converted into the electron dense Ag0 upon exposure to light. From the Hydropathic Profile of the amino acid sequence of adenosine deaminase it is evident that there are two hydrophobic domains of sufficient length to span a biological membrane, and it is proposed that these domains could function to anchor the enzyme to the membrane. The importance of adenosine deaminase is indicated by the fatal immuno-deficiency which results from untreated genetic adenosine deaminase deficiency. It may be important to determine whether the amount of ecto adenosine deaminase activity is better suited to assess the clinical status of adenosine deaminase deficient patients that the currently used total cellular enzyme activity.

  8. An enzyme-free strategy for ultrasensitive detection of adenosine using a multipurpose aptamer probe and malachite green.

    PubMed

    Zhao, Hui; Wang, Yong-Sheng; Tang, Xian; Zhou, Bin; Xue, Jin-Hua; Liu, Hui; Liu, Shan-Du; Cao, Jin-Xiu; Li, Ming-Hui; Chen, Si-Han

    2015-08-05

    We report on an enzyme-free and label-free strategy for the ultrasensitive determination of adenosine. A novel multipurpose adenosine aptamer (MAAP) is designed, which serves as an effective target recognition probe and a capture probe for malachite green. In the presence of adenosine, the conformation of the MAAP is converted from a hairpin structure to a G-quadruplex. Upon addition of malachite green into this solution, a noticeable enhancement of resonance light scattering was observed. The signal response is directly proportional to the concentration of adenosine ranging from 75 pM to 2.2 nM with a detection limit of 23 pM, which was 100-10,000 folds lower than those obtained by previous reported methods. Moreover, this strategy has been applied successfully for detecting adenosine in human urine and blood samples, further proving its reliability. The mechanism of adenosine inducing MAAP to form a G-quadruplex was demonstrated by a series of control experiments. Such a MAAP probe can also be used to other strategies such as fluorescence or spectrophotometric ones. We suppose that this strategy can be expanded to develop a universal analytical platform for various target molecules in the biomedical field and clinical diagnosis.

  9. Regulation of photoreceptor gap junction phosphorylation by adenosine in zebrafish retina

    PubMed Central

    Li, Hongyan; Chuang, Alice Z.; O’Brien, John

    2014-01-01

    Electrical coupling of photoreceptors through gap junctions suppresses voltage noise, routes rod signals into cone pathways, expands the dynamic range of rod photoreceptors in high scotopic and mesopic illumination, and improves detection of contrast and small stimuli. In essentially all vertebrates, connexin 35/36 (gene homologues Cx36 in mammals, Cx35 in other vertebrates) is the major gap junction protein observed in photoreceptors, mediating rod-cone, cone-cone, and possibly rod-rod communication. Photoreceptor coupling is dynamically controlled by the day/night cycle and light/dark adaptation, and is directly correlated with phosphorylation of Cx35/36 at two sites, serine110 and serine 276/293 (homologous sites in teleost fish and mammals respectively). Activity of protein kinase A (PKA) plays a key role during this process. Previous studies have shown that activation of dopamine D4 receptors on photoreceptors inhibits adenylyl cyclase, down-regulates cAMP and PKA activity, and leads to photoreceptor uncoupling, imposing the daytime/light condition. In this study we explored the role of adenosine, a nighttime signal with a high extracellular concentration at night and a low concentration in the day, in regulating photoreceptor coupling by examining photoreceptor Cx35 phosphorylation in zebrafish retina. Adenosine enhanced photoreceptor Cx35 phosphorylation in daytime, but with a complex dose-response curve. Selective pharmacological manipulations revealed that adenosine A2a receptors provide a potent positive drive to phosphorylate photoreceptor Cx35 under the influence of endogenous adenosine at night. A2a receptors can be activated in the daytime as well by micromolar exogenous adenosine. However, the higher affinity adenosine A1 receptors are also present and have an antagonistic though less potent effect. Thus the nighttime/darkness signal adenosine provides a net positive drive on Cx35 phosphorylation at night, working in opposition to dopamine to

  10. Theophylline and adenosine modulate the inflammatory functions of the human neutrophil by exerting an opposing influence on the stimulus-induced increase in intracellular calcium

    SciTech Connect

    Schmeichel Morley, C.J.

    1988-01-01

    Based on evidence that endogenously-produced adenosine inhibited neutrophil responses, the influence of methylxanthine bronchodilators on neutrophil responses stimulated in vitro by n-formyl-methionyl-leucyl-phenylalanine (fMLP) was examined. At concentrations between 10/sup /minus/5/ M and 10/sup /minus/4/ M, theophylline potentiated lysosomal enzyme release by 30 to 50%, superoxide anion formation by 30 to 60%, and neutrophil aggregation. Theophylline at concentrations >10/sup /minus/4/ M inhibited the same responses by >90%. Adenosine deaminase mimicked, whereas adenosine reversed the theophylline potentiation. A potential role for calcium in the modulation of the neutrophil responses by theophylline and adenosine was explored. Theophylline enhanced by >150% the fMLP-stimulated increase in cytoplasmic calcium concentration ((Ca/sup 2 +/)/sub i/) at time points between 5 and 90 sec as measured by Fura-2. Adenosine deaminase induced a comparable enhancement, whereas 3 /times/ 10/sup /minus/7/ M adenosine and 10/sup /minus/7/ M N-ethylcarboxamideadenosine decreased the (Ca/sup 2 +/)/sub i/ in fMLP-stimulated neutrophils. Extracellular calcium was not required for the opposing influences of theophylline and adenosine and neither compound altered fMLP-stimulated /sup 45/Ca uptake at the early time points.

  11. Time- and dose-related interactions between glucocorticoid and cyclic adenosine 3',5'-monophosphate on CCAAT/enhancer-binding protein-dependent insulin-like growth factor I expression by osteoblasts

    NASA Technical Reports Server (NTRS)

    McCarthy, T. L.; Ji, C.; Chen, Y.; Kim, K.; Centrella, M.

    2000-01-01

    Glucocorticoid has complex effects on osteoblasts. Several of these changes appear to be related to steroid concentration, duration of exposure, or specific effects on growth factor expression or activity within bone. One important bone growth factor, insulin-like growth factor I (IGF-I), is induced in osteoblasts by hormones such as PGE2 that increase intracellular cAMP levels. In this way, PGE2 activates transcription factor CCAAT/enhancer-binding protein-delta (C/EBPdelta) and enhances its binding to a specific control element found in exon 1 in the IGF-I gene. Our current studies show that preexposure to glucocorticoid enhanced C/EBPdelta and C/EBPbeta expression by osteoblasts and thereby potentiated IGF-I gene promoter activation in response to PGE2. Importantly, this directly contrasts with inhibitory effects on IGF-I expression that result from sustained or pharmacologically high levels of glucocorticoid exposure. Consistent with the stimulatory effect of IGF-I on bone protein synthesis, pretreatment with glucocorticoid sensitized osteoblasts to PGE2, and in this context significantly enhanced new collagen and noncollagen protein synthesis. Therefore, pharmacological levels of glucocorticoid may reduce IGF-I expression by osteoblasts and cause osteopenic disease, whereas physiological transient increases in glucocorticoid may permit or amplify the effectiveness of hormones that regulate skeletal tissue integrity. These events appear to converge on the important role of C/EBPdelta and C/EBPbeta on IGF-I expression by osteoblasts.

  12. Time- and dose-related interactions between glucocorticoid and cyclic adenosine 3',5'-monophosphate on CCAAT/enhancer-binding protein-dependent insulin-like growth factor I expression by osteoblasts

    NASA Technical Reports Server (NTRS)

    McCarthy, T. L.; Ji, C.; Chen, Y.; Kim, K.; Centrella, M.

    2000-01-01

    Glucocorticoid has complex effects on osteoblasts. Several of these changes appear to be related to steroid concentration, duration of exposure, or specific effects on growth factor expression or activity within bone. One important bone growth factor, insulin-like growth factor I (IGF-I), is induced in osteoblasts by hormones such as PGE2 that increase intracellular cAMP levels. In this way, PGE2 activates transcription factor CCAAT/enhancer-binding protein-delta (C/EBPdelta) and enhances its binding to a specific control element found in exon 1 in the IGF-I gene. Our current studies show that preexposure to glucocorticoid enhanced C/EBPdelta and C/EBPbeta expression by osteoblasts and thereby potentiated IGF-I gene promoter activation in response to PGE2. Importantly, this directly contrasts with inhibitory effects on IGF-I expression that result from sustained or pharmacologically high levels of glucocorticoid exposure. Consistent with the stimulatory effect of IGF-I on bone protein synthesis, pretreatment with glucocorticoid sensitized osteoblasts to PGE2, and in this context significantly enhanced new collagen and noncollagen protein synthesis. Therefore, pharmacological levels of glucocorticoid may reduce IGF-I expression by osteoblasts and cause osteopenic disease, whereas physiological transient increases in glucocorticoid may permit or amplify the effectiveness of hormones that regulate skeletal tissue integrity. These events appear to converge on the important role of C/EBPdelta and C/EBPbeta on IGF-I expression by osteoblasts.

  13. Phosphorylation of Cytokinin by Adenosine Kinase from Wheat Germ 1

    PubMed Central

    Chen, Chong-Maw; Eckert, Richard L.

    1977-01-01

    Adenosine kinase was partially purified from wheat germ. This enzyme preparation, which was devoid of adenine phosphoribosyltransferase and nearly free of adenosine deaminase but contained adenylate kinase, rapidly phosphorylated adenosine and a cytokinin, N6-(δ2-isopentenyl)adenosine. Electrophoretic analysis indicated that only N6-(δ2-isopentenyl)adenosine-monophosphate was formed from the cytokinin while about 55% AMP, 45% ADP, and a trace of ATP were formed from adenosine. The biosynthesized nucleoside monophosphates were quantitatively hydrolyzed to the corresponding nucleosides by 5′-nucleotidase and the isopentenyl side chain of the phosphorylated cytokinin was not cleaved. The enzyme did not catalyze phosphorylation of inosine. The phosphorylation of the cytokinin and adenosine required ATP and Mg2+. The pH optimum was from 6.8 to 7.2 for both the cytokinin and adenosine. At pH 7 and 37 C the Km and Vmax for the cytokinin were 31 μm and 8.3 nmoles per mg protein per minute, and the values for adenosine were 8.7 μm and 46 nmoles per mg protein per minute. Crude enzyme preparations from tobacco callus tissue and wheat germ phosphorylated N6-(δ2-isopentenyl)adenosine. These preparations also phosphorylated N6-(δ2-isopentenyl)adenine when 5-phosphorylribose-1-pyrophosphate was present. PMID:16659870

  14. Therapeutic epilepsy research: from pharmacological rationale to focal adenosine augmentation

    PubMed Central

    Boison, Detlev; Stewart, Kerry-Ann

    2009-01-01

    Epilepsy is a common seizure disorder affecting approximately 70 million people worldwide. Current pharmacotherapy is neuron-centered, frequently accompanied by intolerable side-effects, and fails to be effective in about one third of patients. Therefore, new therapeutic concepts are needed. Recent research suggests an astrocytic basis of epilepsy, presenting the possibility of novel therapeutic targets. In particular, dysfunction of the astrocyte-controlled, endogenous, adenosine-based seizure control system of the brain is implicated in seizure generation. Thus, astrogliosis – a pathological hallmark of the epileptic brain – is associated with upregulation of the adenosine-removing enzyme adenosine kinase (ADK), resulting in focal adenosine deficiency. Both astrogliotic upregulation of ADK in epilepsy and transgenic overexpression of ADK are associated with seizures, and inhibition of ADK prevents seizures in a mouse model of pharmacoresistant epilepsy. These findings link adenosine deficiency with seizures and predict that adenosine augmentation therapies (AATs) will likely be effective in preventing seizures. Given the widespread systemic and central side effects of systemically administered AATs, focal AATs (i.e., limited to the astrogliotic lesion) are a necessity. This Commentary will discuss the pharmacological rationale for the development of focal AATs. Additionally, several AAT strategies will be discussed: (1) adenosine released from silk-based brain implants; (2) adenosine released from locally implanted encapsulated cells; (3) adenosine released from stem cell-derived brain implants; and (4) adenosine augmenting gene therapies. Finally, new developments and therapeutic challenges in using focal AATs for epilepsy therapy will critically be evaluated. PMID:19682439

  15. Role of A2B adenosine receptor signaling in adenosine-dependent pulmonary inflammation and injury

    PubMed Central

    Sun, Chun-Xiao; Zhong, Hongyan; Mohsenin, Amir; Morschl, Eva; Chunn, Janci L.; Molina, Jose G.; Belardinelli, Luiz; Zeng, Dewan; Blackburn, Michael R.

    2006-01-01

    Adenosine has been implicated in the pathogenesis of chronic lung diseases such as asthma and chronic obstructive pulmonary disease. In vitro studies suggest that activation of the A2B adenosine receptor (A2BAR) results in proinflammatory and profibrotic effects relevant to the progression of lung diseases; however, in vivo data supporting these observations are lacking. Adenosine deaminase–deficient (ADA-deficient) mice develop pulmonary inflammation and injury that are dependent on increased lung adenosine levels. To investigate the role of the A2BAR in vivo, ADA-deficient mice were treated with the selective A2BAR antagonist CVT-6883, and pulmonary inflammation, fibrosis, and airspace integrity were assessed. Untreated and vehicle-treated ADA-deficient mice developed pulmonary inflammation, fibrosis, and enlargement of alveolar airspaces; conversely, CVT-6883–treated ADA-deficient mice showed less pulmonary inflammation, fibrosis, and alveolar airspace enlargement. A2BAR antagonism significantly reduced elevations in proinflammatory cytokines and chemokines as well as mediators of fibrosis and airway destruction. In addition, treatment with CVT-6883 attenuated pulmonary inflammation and fibrosis in wild-type mice subjected to bleomycin-induced lung injury. These findings suggest that A2BAR signaling influences pathways critical for pulmonary inflammation and injury in vivo. Thus in chronic lung diseases associated with increased adenosine, antagonism of A2BAR-mediated responses may prove to be a beneficial therapy. PMID:16841096

  16. The role of plasma adenosine deaminase in chemoattractant-stimulated oxygen radical production in neutrophils.

    PubMed

    Kälvegren, Hanna; Fridfeldt, Jonna; Bengtsson, Torbjörn

    2010-06-01

    Adenosine deaminase (ADA) has a role in many immunity mediated disorders, such as asthma, tuberculosis and coronary artery disease. This study aims to investigate the ability of plasma ADA to modulate reactive oxygen species (ROS) production in neutrophils, and examine the involvement of adenosine and the cyclic AMP signaling pathway in this process. Neutrophils were stimulated, in the absence or presence of plasma, with the chemotactic peptide fMLP (formyl-methionyl-leucyl-phenylalanine), and the ROS production was determined with luminol-enhanced chemiluminescence. Activity of ADA was measured spectrophotometrically. Plasma dose-dependently amplified the ROS generation in fMLP-stimulated neutrophils. In parallel, incubation of neutrophils in plasma elevated the total ADA-activity approximately 10 times from 1.3 U/ml to 12 U/ml. Inhibition of ADA, or type IV phosphodiesterases, significantly lowered the plasma-mediated ROS production. Furthermore, the high-affinity adenosine A(1) receptor antagonists DPCPX and 8-phenyltheophylline markedly inhibited the plasma-induced respiratory burst in neutrophils, suggesting an A(1) receptor-mediated mechanism. This study suggests that plasma ADA amplifies the release of toxic oxygen radicals from neutrophils through a downregulation of the inhibitory adenosine/cAMP-system and an enhanced activation of the stimulatory adenosine A(1)-receptor. This mechanism has probably a crucial role in regulating neutrophil function and in the defence against microbial infections. However, a sustained neutrophil activation could also contribute to inflammatory disorders such as atherosclerosis. 2010 Elsevier GmbH. All rights reserved.

  17. Chemoelectrical energy conversion of adenosine triphosphate

    NASA Astrophysics Data System (ADS)

    Sundaresan, Vishnu Baba; Sarles, Stephen Andrew; Leo, Donald J.

    2007-04-01

    Plant and animal cell membranes transport charged species, neutral molecules and water through ion pumps and channels. The energy required for moving species against established concentration and charge gradients is provided by the biological fuel - adenosine triphosphate (ATP) -synthesized within the cell. The adenosine triphosphatase (ATPases) in a plant cell membrane hydrolyze ATP in the cell cytoplasm to pump protons across the cell membrane. This establishes a proton gradient across the membrane from the cell exterior into the cell cytoplasm. This proton motive force stimulates ion channels that transport nutrients and other species into the cell. This article discusses a device that converts the chemical energy stored in adenosine triphosphate into electrical power using a transporter protein, ATPase. The V-type ATPase proteins used in our prototype are extracted from red beet(Beta vulgaris) tonoplast membranes and reconstituted in a bilayer lipid membrane or BLM formed from POPC and POPS lipids. A pH7 medium that can support ATP hydrolysis is provided on both sides of the membrane and ATP is dissolved in the pH7 buffer on one side of the membrane. Hydrolysis of ATP results in the formation of a phosphate ion and adenosine diphosphate. The energy from the reaction activates ATPase in the BLM and moves a proton across the membrane. The charge gradient established across the BLM due to the reaction and ion transport is converted into electrical current by half-cell reference electrodes. The prototype ATPase cell with an effective BLM area of 4.15 mm2 carrying 15 μl of ATPase proteins was observed to develop a steady state peak power output of 70 nW, which corresponds to a specific power of 1.69 μW/cm2 and a current density of 43.4 μA/cm2 of membrane area.

  18. The Janus face of adenosine: antiarrhythmic and proarrhythmic actions.

    PubMed

    Szentmiklosi, A József; Galajda, Zoltán; Cseppento, Ágnes; Gesztelyi, Rudolf; Susán, Zsolt; Hegyi, Bence; Nánási, Péter P

    2015-01-01

    Adenosine is a ubiquitous, endogenous purine involved in a variety of physiological and pathophysiological regulatory mechanisms. Adenosine has been proposed as an endogenous antiarrhythmic substance to prevent hypoxia/ischemia-induced arrhythmias. Adenosine (and its precursor, ATP) has been used in the therapy of various cardiac arrhythmias over the past six decades. Its primary indication is treatment of paroxysmal supraventricular tachycardia, but it can be effective in other forms of supraventricular and ventricular arrhythmias, like sinus node reentry based tachycardia, triggered atrial tachycardia, atrioventricular nodal reentry tachycardia, or ventricular tachycardia based on a cAMP-mediated triggered activity. The main advantage is the rapid onset and the short half life (1- 10 sec). Adenosine exerts its antiarrhythmic actions by activation of A1 adenosine receptors located in the sinoatrial and atrioventricular nodes, as well as in activated ventricular myocardium. However, adenosine can also elicit A2A, A2B and A3 adenosine receptor-mediated global side reactions (flushing, dyspnea, chest discomfort), but it may display also proarrhythmic actions mediated by primarily A1 adenosine receptors (e.g. bradyarrhythmia or atrial fibrillation). To avoid the non-specific global adverse reactions, A1 adenosine receptor- selective full agonists (tecadenoson, selodenoson, trabodenoson) have been developed, which agents are currently under clinical trial. During long-term administration with orthosteric agonists, adenosine receptors can be internalized and desensitized. To avoid desensitization, proarrhythmic actions, or global adverse reactions, partial A1 adenosine receptor agonists, like CVT-2759, were developed. In addition, the pharmacologically "silent" site- and event specific adenosinergic drugs, such as adenosine regulating agents and allosteric modulators, might provide attractive opportunity to increase the effectiveness of beneficial actions of adenosine

  19. Role of adenosine in oligodendrocyte precursor maturation

    PubMed Central

    Coppi, Elisabetta; Cellai, Lucrezia; Maraula, Giovanna; Dettori, Ilaria; Melani, Alessia; Pugliese, Anna Maria; Pedata, Felicita

    2015-01-01

    Differentiation and maturation of oligodendroglial cells are postnatal processes that involve specific morphological changes correlated with the expression of stage-specific surface antigens and functional voltage-gated ion channels. A small fraction of oligodendrocyte progenitor cells (OPCs) generated during development are maintained in an immature and slowly proliferative or quiescent state in the adult central nervous system (CNS) representing an endogenous reservoir of immature cells. Adenosine receptors are expressed by OPCs and a key role of adenosine in oligodendrocyte maturation has been recently recognized. As evaluated on OPC cultures, adenosine, by stimulating A1 receptors, promotes oligodendrocyte maturation and inhibits their proliferation; on the contrary, by stimulating A2A receptors, it inhibits oligodendrocyte maturation. A1 and A2A receptor-mediated effects are related to opposite modifications of outward delayed rectifying membrane K+ currents (IK) that are involved in the regulation of oligodendrocyte differentiation. Brain A1 and A2A receptors might represent new molecular targets for drugs useful in demyelinating pathologies, such as multiple sclerosis (MS), stroke and brain trauma. PMID:25964740

  20. Effects of adenosine agonists on consumptive behaviour and body temperature.

    PubMed

    Coupar, Ian M; Tran, Binh L T

    2002-02-01

    This study was designed to determine the effects of the A1-receptor selective agonist N6-cyclopentyladenosine (CPA), and the A2-selective agonist, 2-p-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine-hydrochloride (CGS-21680) on consumptive behaviour and body temperature in rats in relation to the non-selective A1/A2 adenosine agonist, N-ethylcarboxamidoadenosine (NECA), and to morphine. It was shown that two subcutaneous injections of 0.1 and 0.3 mg kg(-1) CPA caused a similar decrease in food consumption to NECA (2 x 0.03 mg kg(-1)) and morphine (2 x 10 mg kg(-1)). However, two doses of 0.03 mg kg(-1) CPA and 0.1 and 0.3 mg kg(-1)CGS-21680 enhanced feeding. These effects were not directly correlated to faecal output at all doses of the selective agonists, as NECA and morphine induced constipation. The doses of CPA and 0.1 and 0.3 mg kg(-1) of CGS-21680 enhanced water consumption, as did NECA, but not morphine. The stimulation of drinking by CPA was not absolutely associated with diuresis. Instead, urine output was reduced by 0.03 and 0.1 mg kg(-1) and increased by 0.3 mg kg(-1). CGS-21680 at 0.1 and 0.3 mg kg(-1) and NECA also induced diuresis, which was opposite to the effect of morphine. CPA and CGS-21680 both caused significant dose-dependent decreases in body temperature after the two-injection treatment, but their effects were significantly less after 36 h when four doses had been administered. The study indicates that highly selective A1 and A2A adenosine agonists might have the ability to interfere with consumptive behaviour, induce constipation, affect renal function and to lower body temperature.

  1. A Functionalized Congener Approach to Adenosine Receptor Antagonists: Amino Acid Conjugates of 1,3-Dipropylxanthine

    PubMed Central

    JACOBSON, KENNETH A.; KIRK, KENNETH L.; PADGETT, WILLIAM L.; DALY, JOHN W.

    2012-01-01

    SUMMARY 1,3-Dipropyl-8-phenylxanthine, a synthetic analog of theophylline and a potent antagonist of adenosine at A1 and A2-adenosine receptors, has been attached covalently through a functionalized chain to amino acids and oligopeptides. The xanthine conjugates have been studied as competitive inhibitors of the specific binding of [3H]N6-cyclohexyladenosine to A1-receptors of rat cerebral cortical membranes and for inhibition of cyclic AMP accumulation elicited by 2-chloroadenosine in guinea pig brain slices through A2-receptors. A free amino group on the extended chain generally resulted in high potency at A1-receptors. The potency (in some cases extending into the subnanomolar range) and selectivity for A1-receptors (up to 200-fold) suggest that this approach can yield a versatile class of “functionalized congeners” of adenosine receptor antagonists in which distal modifications of the attached moiety (“carrier”) can serve also to improve pharmacodynamic and pharmacokinetic parameters. The water solubility in many of the more potent analogs has been enhanced by two orders of magnitude over that of simple, uncharged 8-phenyl xanthine derivatives. Analogs in which the carrier contains d-tyrosine have potential for development of iodinated radioligands for adenosine receptors. The functionalized congener approach is potentially applicable to other drugs and for development of prodrugs. PMID:3005825

  2. Altered Hypoxic-Adenosine Axis and Metabolism in Group III Pulmonary Hypertension.

    PubMed

    Garcia-Morales, Luis J; Chen, Ning-Yuan; Weng, Tingting; Luo, Fayong; Davies, Jonathan; Philip, Kemly; Volcik, Kelly A; Melicoff, Ernestina; Amione-Guerra, Javier; Bunge, Raquel R; Bruckner, Brian A; Loebe, Matthias; Eltzschig, Holger K; Pandit, Lavannya M; Blackburn, Michael R; Karmouty-Quintana, Harry

    2016-04-01

    Group III pulmonary hypertension (PH) is a highly prevalent and deadly lung disorder with limited treatment options other than transplantation. Group III PH affects patients with ongoing chronic lung injury, such as idiopathic pulmonary fibrosis (IPF). Between 30 and 40% of patients with IPF are diagnosed with PH. The diagnosis of PH has devastating consequences to these patients, leading to increased morbidity and mortality, yet the molecular mechanisms involved in the development of PH in patients with chronic lung disease remain elusive. Our hypothesis was that the hypoxic-adenosinergic system is enhanced in patients with group III PH compared with patients with IPF with no PH. Explanted lung tissue was analyzed for markers of the hypoxic-adenosine axis, including expression levels of hypoxia-inducible factor (HIF)-1A, adenosine A2B receptor, CD73, and equilibrative nucleotide transporter-1. In addition, we assessed whether altered mitochondrial metabolism was present in these samples. Increased expression of HIF-1A was observed in tissues from patients with group III PH. These changes were consistent with increased evidence of adenosine accumulation in group III PH. A novel observation of our study was of evidence suggesting altered mitochondrial metabolism in lung tissue from group III PH leading to increased succinate levels that are able to further stabilize HIF-1A. Our data demonstrate that the hypoxic-adenosine axis is up-regulated in group III PH and that subsequent succinate accumulation may play a part in the development of group III PH.

  3. Effects of adenosine perfusion on the metabolism and contractile activity of Rana ridibunda heart.

    PubMed

    Lazou, A; Beis, I

    1987-01-01

    The effects of adenosine were examined on the isolated perfused heart of the frog Rana ridibunda. Adenosine produced negative chronotropic and inotropic effects on frog ventricle in a concentration-dependent manner. The effects of adenosine on cardiac metabolism were also investigated by measuring the tissue content of adenine nucleotides, lactate, pyruvate, adenosine and inorganic phosphate, during adenosine perfusion. Adenosine had no effect on the tissue content of metabolites. No net synthesis of adenine nucleotides was observed during perfusion with increasing concentrations of adenosine. Lactate output from the heart decreased significantly with adenosine perfusion. Correlation of adenosine effects on cardiac muscle with the effects of hypoxia are discussed.

  4. [The involvement of adenosine and adenosine deaminase in experimental myocardial infarct].

    PubMed

    Stratone, A; Busuioc, A; Roşca, V; Bazgan, L; Popa, M; Hăulică, I

    1989-01-01

    By the ligature of the left coronary artery in the rat anesthetized with nembutal (10 mg/100 i.p.) a significant increase of the 5'-nucleotidase activity (Wooton method) was noticed 10 minutes after the left ventricle infarction (from an average value of 1038.5 +/- 187 mU/g tissue to 1537 +/- 225 mU/g fresh tissue). The adenosine desaminase levels spectrophotometrically determined by Denstedt technique, do not appear significantly modified 10 or 30 minutes after the left ventricle infarction. The chromatographically determined adenosine levels, by HPLC technique, decrease from the average value of 11.63 +/- 1.4 micrograms/mg PT to 8.60 +/- 1.0 micrograms/mg PT 30 minutes after infarction. The observed changes are explained by the conditions of hypoxia in the infarcted ventricle which lead to the raise in adenosine levels by activating the 5'-nucleotidase and their depression by a very fast metabolism of the same substance.

  5. Effect of adenosine and adenosine analogs on ( sup 14 C)aminopyrine accumulation by rabbit parietal cells

    SciTech Connect

    Ota, S.; Hiraishi, H.; Terano, A.; Mutoh, H.; Kurachi, Y.; Shimada, T.; Ivey, K.J.; Sugimoto, T. )

    1989-12-01

    Adenosine receptors that modulate adenylate cyclase activity have been identified recently in a number of tissues. Adenosine A2 receptor is stimulatory to adenylate cyclase, whereas adenosine A1 receptor is inhibitory to adenylate cyclase. We investigated the effect of adenosine and its analogs on (14C)aminopyrine accumulation by rabbit parietal cells. Rabbit gastric mucosal cells were isolated by enzyme digestion. Parietal cells were enriched by nonlinear percoll gradients. (14C)Aminopyrine accumulation was used as an indicator of acid secretion. The effect of 2-chloroadenosine on histamine-stimulated (14C)aminopyrine accumulation was studied. The effects of N-ethylcarboxamideadenosine, 2-chloroadenosine, stable analogs of adenosine, and adenosine on (14C)aminopyrine accumulation were assessed. Cyclic AMP content of parietal cells was determined by radioimmunoassay. Histamine and carbachol, known secretagogues, stimulated (14C)aminopyrine accumulation. 2-Chloroadenosine did not suppress histamine-stimulated (14C)aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine dose dependently increased (14C)aminopyrine accumulation. The order of potency was N-ethylcarboxamideadenosine greater than 2-chloroadenosine greater than adenosine. 8-Phenyltheophylline and theophylline, adenosine-receptor antagonists, or cimetidine did not have significant effects on the increase of AP uptake induced by 2-chloroadenosine. Coadministration of dipyridamole, and adenosine uptake inhibitor, augmented the effect of adenosine on (14C)aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine each induced a significant increase in cellular cyclic AMP. We conclude that there may be adenosine A2 receptors on rabbit parietal cells which modulate gastric acid secretion.

  6. N6-(2-Hydroxyethyl)-Adenosine Exhibits Insecticidal Activity against Plutella xylostella via Adenosine Receptors

    PubMed Central

    Fang, Ming; Chai, Yiqiu; Chen, Guanjv; Wang, Huidong; Huang, Bo

    2016-01-01

    The diamondback moth, Plutella xylostella, is one of the most important pests of cruciferous crops. We have earlier shown that N6-(2-hydroxyethyl)-adenosine (HEA) exhibits insecticidal activity against P. xylostella. In the present study we investigated the possible mechanism of insecticidal action of HEA on P. xylostella. HEA is a derivative of adenosine, therefore, we speculated whether it acts via P. xylostella adenosine receptor (PxAdoR). We used RNAi approach to silence PxAdoR gene and used antagonist of denosine receptor (AdoR) to study the insecticidal effect of HEA. We cloned the whole sequence of PxAdoR gene. A BLAST search using NCBI protein database showed a 61% identity with the Drosophila adenosine receptor (DmAdoR) and a 32–35% identity with human AdoR. Though the amino acids sequence of PxAdoR was different compared to other adenosine receptors, most of the amino acids that are known to be important for adenosine receptor ligand binding and signaling were present. However, only 30% binding sites key residues was similar between PxAdoR and A1R. HEA, at a dose of 1 mg/mL, was found to be lethal to the second-instar larvae of P. xylostella, and a significant reduction of mortality and growth inhibition ratio were obtained when HEA was administered to the larvae along with PxAdoR-dsRNA or antagonist of AdoR (SCH58261) for 36, 48, or 60 h. Especially at 48 h, the rate of growth inhibition of the PxAdoR knockdown group was 3.5-fold less than that of the HEA group, and the corrected mortality of SCH58261 group was reduced almost 2-fold compared with the HEA group. Our findings show that HEA may exert its insecticidal activity against P. xylostella larvae via acting on PxAdoR. PMID:27668428

  7. N6-(2-Hydroxyethyl)-Adenosine Exhibits Insecticidal Activity against Plutella xylostella via Adenosine Receptors.

    PubMed

    Fang, Ming; Chai, Yiqiu; Chen, Guanjv; Wang, Huidong; Huang, Bo

    The diamondback moth, Plutella xylostella, is one of the most important pests of cruciferous crops. We have earlier shown that N6-(2-hydroxyethyl)-adenosine (HEA) exhibits insecticidal activity against P. xylostella. In the present study we investigated the possible mechanism of insecticidal action of HEA on P. xylostella. HEA is a derivative of adenosine, therefore, we speculated whether it acts via P. xylostella adenosine receptor (PxAdoR). We used RNAi approach to silence PxAdoR gene and used antagonist of denosine receptor (AdoR) to study the insecticidal effect of HEA. We cloned the whole sequence of PxAdoR gene. A BLAST search using NCBI protein database showed a 61% identity with the Drosophila adenosine receptor (DmAdoR) and a 32-35% identity with human AdoR. Though the amino acids sequence of PxAdoR was different compared to other adenosine receptors, most of the amino acids that are known to be important for adenosine receptor ligand binding and signaling were present. However, only 30% binding sites key residues was similar between PxAdoR and A1R. HEA, at a dose of 1 mg/mL, was found to be lethal to the second-instar larvae of P. xylostella, and a significant reduction of mortality and growth inhibition ratio were obtained when HEA was administered to the larvae along with PxAdoR-dsRNA or antagonist of AdoR (SCH58261) for 36, 48, or 60 h. Especially at 48 h, the rate of growth inhibition of the PxAdoR knockdown group was 3.5-fold less than that of the HEA group, and the corrected mortality of SCH58261 group was reduced almost 2-fold compared with the HEA group. Our findings show that HEA may exert its insecticidal activity against P. xylostella larvae via acting on PxAdoR.

  8. Use of adenosine echocardiography for diagnosis of coronary artery disease

    SciTech Connect

    Zoghbi, W.A. )

    1991-07-01

    Two-dimensional echocardiography combined with exercise is sensitive and specific in the detection of coronary artery disease (CAD) by demonstrating transient abnormalities in wall motion. Frequently, however, patients cannot achieve maximal exercise because of various factors. Pharmacologic stress testing with intravenous adenosine was evaluated as a means of detecting CAD in a noninvasive manner. Patients with suspected CAD underwent echocardiographic imaging and simultaneous thallium 201 single-photon emission computed tomography during the intravenous administration of 140 micrograms/kg/min of adenosine. An increase in heart rate, decrease in blood pressure, and increase in double product were observed during adenosine administration. Initial observations revealed that wall motion abnormalities were induced by adenosine in areas of perfusion defects. The adenosine infusion was well tolerated, and symptoms disappeared within 1 to 2 minutes after termination of the infusion. Therefore preliminary observations suggest that adenosine echocardiography appears to be useful in the assessment of CAD.

  9. Adenosine and inflammation: what's new on the horizon?

    PubMed

    Antonioli, Luca; Csóka, Balázs; Fornai, Matteo; Colucci, Rocchina; Kókai, Endre; Blandizzi, Corrado; Haskó, György

    2014-08-01

    Adenosine contributes to the maintenance of tissue integrity by modulating the immune system. Encouraging results have emerged with adenosine receptor ligands for the management of several inflammatory conditions in preclinical and clinical settings. However, therapeutic applications of these drugs are sometimes complicated by the occurrence of serious adverse effects. The scientific community is making intensive efforts to design novel adenosine receptor ligands endowed with greater selectivity or to develop innovative compounds acting as allosteric receptor modulators. In parallel, research is focusing on novel pharmacological entities (designated as adenosine-regulating agents) that can increase, in a site- and event-specific manner, adenosine concentrations at the inflammatory site, thereby minimizing the adverse systemic effects of adenosine.

  10. Rhodium Complex and Enzyme Couple Mediated Electrochemical Detection of Adenosine.

    PubMed

    Han, Dawoon; Kim, Hyeong-Mook; Chand, Rohit; Kim, Gyumin; Shin, Ik-Soo; Kim, Yong-Sang

    2015-10-01

    Adenosine is one of the nucleoside which plays an important role in signal transduction and neuromodulation. This work proposes a simple electrochemical assay, comprising two enzymes and rhodium complex based electron transfer mediator, for the detection of adenosine. Sequential reaction of adenosine deaminase and L-glutamic dehydrogenase and the supporting cycle between β-NADH and mediator enable quantitative analysis of adenosine. Role of electron transfer mediator is the conveyance of proton from electrode to β-NAD(+) for regeneration of β-NADH. The electrochemical characteristics of electron transfer mediator were also studied. Real-time adenosine detection was carried out using this multiple enzyme based chronoamperometric assay. The analysis results show a low limit of detection (140 μM) and good correspondence between current signal and the adenosine concentration (R (2) = 0.997).

  11. c-Jun enhancement of cyclic adenosine 3',5'-monophosphate response element-dependent transcription induced by transforming growth factor-beta is independent of c-Jun binding to DNA.

    PubMed

    Hu, P P; Harvat, B L; Hook, S S; Shen, X; Wang, X F; Means, A R

    1999-12-01

    Transforming growth factor-beta (TGFbeta) enhances transcription from reporter genes regulated by a single consensus cAMP-response element (CRE) upon transfection into the immortalized human keratinocyte cell line, HaCaT. Whereas both CRE-binding protein (CREB) and c-Jun present in extracts of unstimulated cells can complex with a CRE in gel-shift experiments, TGFbeta treatment increases the amount of c-Jun found in the complex. Overexpression of c-Jun is sufficient to increase CRE and GAL4-CREB-dependent transcription and mimics the stimulatory effects of TGFbeta on transcription from either reporter gene. Surprisingly, although a portion of CREB in unstimulated cells is phosphorylated on the activating serine residue, Ser-133, this level of phospho-CREB is not altered by TGFbeta treatment. In fact, the CREB-dependent transcriptional effects of TGFbeta or c-Jun do not require phosphorylation of Ser-133, although CREB-binding protein (CBP) is required as evidenced by the observation that the adenoviral oncoprotein E1A can block the effects of both agents. c-Jun enhancement of CRE or GAL4-CREB-dependent transcription neither requires the DNA-binding nor N-terminal domains of c-Jun. Collectively, these results are consistent with a model in which signaling pathways initiated by TGFbeta can stimulate CREB-dependent transcription by increasing the cellular concentration of c-Jun, which participates in activation of the CBP-containing transcription complex.

  12. Turnover of adenosine in plasma of human and dog blood

    SciTech Connect

    Moeser, G.H.S.; Schrader, J.; Deussen, A.

    1989-04-01

    To determine half-life and turnover of plasma adenosine, heparinized blood from healthy volunteers was incubated with radiolabeled adenosine in the physiological concentration range of 0.1-1 microM. Plasma levels of adenosine in vitro were 82 +/- 14 nM and were similar to those determined immediately after blood collection with a ''stopping solution.'' Dipyridamole (83 microM) and erythro-9(2-hydroxynon-3yl)-adenine (EHNA) (8 microM) did not measurably alter basal adenosine levels but completely blocked the uptake of added adenosine. Inhibition of ecto-5'-nucleotidase with 100 microM alpha, beta-methyleneadenosine 5'-diphosphate (AOPCP) reduced plasma adenosine to 22 +/- 6 nM. For the determination of adenosine turnover, the decrease in specific radioactivity of added (/sup 3/H)adenosine was measured using a dipyridamole-containing stopping solution. Without altering basal adenosine levels, the half-life was estimated to be 0.6 s. Similar experiments were carried out with washed erythrocytes or in the presence of AOPCP, yielding half-lives of 0.7 and 0.9 s, respectively. When the initial adenosine concentration was 1 microM, its specific activity decreased by only 11% within 5 s, whereas total plasma adenosine exponentially decreased with a half-life of 1.5 s. Venous plasma concentrations were measured after relief of a 3-min forearm ischemia. Changes in plasma adenosine did not correlate well with changes in blood flow but were augmented in the presence of dipyridamole.

  13. Drugs elevating extracellular adenosine promote regeneration of haematopoietic progenitor cells in severely myelosuppressed mice: their comparison and joint effects with the granulocyte colony-stimulating factor.

    PubMed

    Hofer, Michal; Pospísil, Milan; Znojil, Vladimír; Vacek, Antonín; Weiterova, Lenka; Holá, Jirina; Vácha, Jirí

    2002-01-01

    We tested capabilities of drugs elevating extracellular adenosine and of granulocyte colony-stimulating factor (G-CSF) given alone or in combination to modulate regeneration from severe myelosuppression resulting from combined exposure of mice to ionizing radiation and carboplatin. Elevation of extracellular adenosine was induced by joint administration of dipyridamole (DP), a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate (AMP), serving as an adenosine prodrug. DP+AMP, G-CSF or all these drugs in combination were administered in a 4-d treatment regimen starting on day 3 after induction of myelosuppression. Comparable enhancements of haematopoietic regeneration due to elevation of extracellular adenosine or to action of G-CSF were demonstrated as shown by elevated numbers of haematopoietic progenitor cells for granulocytes/macrophages (GM-CFC) and erythrocytes (BFU-E) in the bone marrow and spleen in early time intervals after termination of the drug treatment, i.e. on days 7 and 10 after induction of myelosuppression. Coadministration of all the drugs further potentiated the restoration of progenitor cell pools in the haematopoietic organs. The effects of the drug treatments on progenitor cells were reflected in the peripheral blood in later time intervals of days 15 and 20 after induction of myelosuppression, especially as significantly elevated numbers of granulocytes and less pronounced elevation of lymphocytes and erythrocytes. The results substantiate the potential of drugs elevating extracellular adenosine for clinical utilization in myelosuppressive states, e.g. those accompanying oncological radio- and chemotherapy.

  14. The Role of Adenosine Signaling in Headache: A Review

    PubMed Central

    Fried, Nathan T.; Elliott, Melanie B.; Oshinsky, Michael L.

    2017-01-01

    Migraine is the third most prevalent disease on the planet, yet our understanding of its mechanisms and pathophysiology is surprisingly incomplete. Recent studies have built upon decades of evidence that adenosine, a purine nucleoside that can act as a neuromodulator, is involved in pain transmission and sensitization. Clinical evidence and rodent studies have suggested that adenosine signaling also plays a critical role in migraine headache. This is further supported by the widespread use of caffeine, an adenosine receptor antagonist, in several headache treatments. In this review, we highlight evidence that supports the involvement of adenosine signaling in different forms of headache, headache triggers, and basic headache physiology. This evidence supports adenosine A2A receptors as a critical adenosine receptor subtype involved in headache pain. Adenosine A2A receptor signaling may contribute to headache via the modulation of intracellular Cyclic adenosine monophosphate (cAMP) production or 5' AMP-activated protein kinase (AMPK) activity in neurons and glia to affect glutamatergic synaptic transmission within the brainstem. This evidence supports the further study of adenosine signaling in headache and potentially illuminates it as a novel therapeutic target for migraine. PMID:28335379

  15. Adenosine signaling: good or bad in erectile function?

    PubMed

    Wen, Jiaming; Xia, Yang

    2012-04-01

    The erectile status of penile tissue is governed largely by the tone of cavernosal smooth muscle cells, which is determined by the balance of vascular relaxants and constrictors. Vascular relaxants play a key role in regulating the tone of cavernosal smooth muscle and thus the initiation and maintenance of penile erection. Early studies drew attention to the potential role of adenosine signaling in this process. However, the serendipitous discovery of the effect of sildenafil on erectile physiology drew more attention toward nitric oxide (NO) as a vasodilator in the process of penile erection, and a recently discovered, unexpected erectile phenotype of adenosine deaminase-deficient mice reemphasizes the importance of adenosine as a key regulatory of erectile status. Adenosine, like NO, is a potent and short-lived vasorelaxant that functions via cyclic nucleotide second messenger signaling to promote smooth muscle relaxation. Recent studies reviewed here show that adenosine functions to relax the corpus cavernosum and promote penile erection. Excess adenosine in penile tissue contributes to the disorder called priapism, and impaired adenosine signaling is associated with erectile dysfunction. More recent research summarized in this review reveals that adenosine functions as a key endogenous vasodilator in the initiation and maintenance of normal penile erection. This new insight highlights adenosine signaling pathways operating in penile tissue as significant therapeutic targets for the treatment of erectile disorders.

  16. A Metabolic Immune Checkpoint: Adenosine in Tumor Microenvironment

    PubMed Central

    Ohta, Akio

    2016-01-01

    Within tumors, some areas are less oxygenated than others. Since their home ground is under chronic hypoxia, tumor cells adapt to this condition by activating aerobic glycolysis; however, this hypoxic environment is very harsh for incoming immune cells. Deprivation of oxygen limits availability of energy sources and induces accumulation of extracellular adenosine in tumors. Extracellular adenosine, upon binding with adenosine receptors on the surface of various immune cells, suppresses pro-inflammatory activities. In addition, signaling through adenosine receptors upregulates a number of anti-inflammatory molecules and immunoregulatory cells, leading to the establishment of a long-lasting immunosuppressive environment. Thus, due to hypoxia and adenosine, tumors can discourage antitumor immune responses no matter how the response was induced, whether it was spontaneous or artificially introduced with a therapeutic intention. Preclinical studies have shown the significance of adenosine in tumor survival strategy by demonstrating tumor regression after inactivation of adenosine receptors, inhibition of adenosine-producing enzymes, or reversal of tissue hypoxia. These promising results indicate a potential use of the inhibitors of the hypoxia–adenosine pathway for cancer immunotherapy. PMID:27066002

  17. Adenosine modulates LPS-induced cytokine production in porcine monocytes.

    PubMed

    Ondrackova, Petra; Kovaru, Hana; Kovaru, Frantisek; Leva, Lenka; Faldyna, Martin

    2013-03-01

    Adenosine plays an important role during inflammation, particularly through modulation of monocyte function. The objective of the present study was to evaluate the effect of synthetic adenosine analogs on cytokine production by porcine monocytes. The LPS-stimulated cytokine production was measured by flow cytometry and quantitative real-time PCR. Adenosine receptor expression was measured by quantitative real-time PCR. The present study demonstrates that adenosine analog N-ethylcarboxyamidoadenosine (NECA) down-regulates TNF-α production and up-regulates IL-8 production by LPS-stimulated porcine monocytes. The effect was more pronounced in CD163(-) subset of monocytes compared to the CD163(+) subset. Although both monocyte subsets express mRNA for A1, A2A, A2B and A3 adenosine receptors, the treatment of monocytes with various adenosine receptor agonists and antagonists proved that the effect of adenosine is mediated preferentially via A2A adenosine receptor. Moreover, the study suggests that the effect of NECA on porcine monocytes alters the levels of the cytokines which could play a role in the differentiation of naive T cells into Th17 cells. The results suggest that adenosine plays an important role in modulation of cytokine production by porcine monocytes.

  18. Cardioprotection with adenosine: 'a riddle wrapped in a mystery'.

    PubMed

    Przyklenk, Karin; Whittaker, Peter

    2005-07-01

    Review of the published literature on adenosine and cardioprotection could lead one to paraphrase the famous words of Sir Winston Churchill (Radio broadcast, 1 October 1939 (in reference to Russia)) and conclude: 'I cannot forecast to you the action of adenosine. It is a riddle wrapped in a mystery inside an enigma'. That is, although it is well-established that adenosine can render cardiomyocytes resistant to lethal ischemia/reperfusion-induced injury, new and intriguing insights continue to emerge as to the mechanisms by which adenosine might limit myocardial infarct size.

  19. An essential role for adenosine signaling in alcohol abuse.

    PubMed

    Ruby, Christina L; Adams, Chelsea A; Knight, Emily J; Nam, Hyung Wook; Choi, Doo-Sup

    2010-09-01

    In the central nervous system (CNS), adenosine plays an important role in regulating neuronal activity and modulates signaling by other neurotransmitters, including GABA, glutamate, and dopamine. Adenosine suppresses neurotransmitter release, reduces neuronal excitability, and regulates ion channel function through activation of four classes of G protein-coupled receptors, A(1), A(2A), A(2B), and A(3). Central adenosine are largely controlled by nucleoside transporters, which transport adenosine levels across the plasma membrane. Adenosine has been shown to modulate cortical glutamate signaling and ventral-tegmental dopaminergic signaling, which are involved in several aspects of alcohol use disorders. Acute ethanol elevates extracellular adenosine levels by selectively inhibiting the type 1 equilibrative nucleoside transporter, ENT1. Raised adenosine levels mediate the ataxic and sedative/hypnotic effects of ethanol through activation of A(1) receptors in the cerebellum, striatum, and cerebral cortex. Recently, we have shown that pharmacological inhibition or genetic deletion of ENT1 reduces the expression of excitatory amino acid transporter 2 (EAAT2), the primary regulator of extracellular glutamate, in astrocytes. These lines of evidence support a central role for adenosine-mediated glutamate signaling and the involvement of astrocytes in regulating ethanol intoxication and preference. In this paper, we discuss recent findings on the implication of adenosine signaling in alcohol use disorders.

  20. An Essential Role for Adenosine Signaling in Alcohol Abuse

    PubMed Central

    Ruby, Christina L.; Adams, Chelsea; Knight, Emily J.; Nam, Hyung Wook; Choi, Doo-Sup

    2014-01-01

    In the central nervous system (CNS), adenosine plays an important role in regulating neuronal activity and modulates signaling by other neurotransmitters, including GABA, glutamate, and dopamine. Adenosine suppresses neurotransmitter release, reduces neuronal excitability, and regulates ion channel function through activation of four classes of G protein-coupled receptors, A1, A2A, A2B, and A3. Central adenosine levels are largely controlled by nucleoside transporters, which regulate adenosine levels across the plasma membrane. Adenosine has been shown to modulate cortical glutamate signaling and ventral-tegmental dopaminergic signaling, which are involved in several aspects of alcohol use disorders. Acute ethanol elevates extracellular adenosine levels by selectively inhibiting the type 1 equilibrative nucleoside transporter, ENT1. Raised adenosine levels mediate the ataxic and sedative/hypnotic effects of ethanol through activation of A1 receptors in the cerebellum, striatum, and cerebral cortex. Recently, we have shown that pharmacological inhibition or genetic deletion of ENT1 reduces the expression of excitatory amino acid transporter 2 (EAAT2), the primary regulator of extracellular glutamate, in astrocytes. These lines of evidence support a central role for adenosine-mediated glutamate signaling and the involvement of astrocytes in regulating ethanol intoxication and preference. In this paper, we discuss recent findings on the implication of adenosine signaling in alcohol use disorders. PMID:21054262

  1. The effect of cannabidiol on ischemia/reperfusion-induced ventricular arrhythmias: the role of adenosine A1 receptors.

    PubMed

    Gonca, Ersöz; Darıcı, Faruk

    2015-01-01

    Cannabidiol (CBD) is a nonpsychoactive phytocannabinoid with anti-inflammatory activity mediated by enhancing adenosine signaling. As the adenosine A1 receptor activation confers protection against ischemia/reperfusion (I/R)-induced ventricular arrhythmias, we hypothesized that CBD may have antiarrhythmic effect through the activation of adenosine A1 receptor. Cannabidiol has recently been shown to suppress ischemia-induced ventricular arrhythmias. We aimed to research the effect of CBD on the incidence and the duration of I/R-induced ventricular arrhythmias and to investigate the role of adenosine A1 receptor activation in the possible antiarrhythmic effect of CBD. Myocardial ischemia and reperfusion was induced in anesthetized male rats by ligating the left anterior descending coronary artery for 6 minutes and by loosening the bond at the coronary artery, respectively. Cannabidiol alone was given in a dose of 50 µg/kg, 10 minutes prior to coronary artery occlusion and coadministrated with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) in a dose of 100 µg/kg, 15 minutes prior to coronary artery occlusion to investigate whether the antiarrhythmic effect of CBD is modified by the activation of adenosine A1 receptors. The experimental groups were as follows: (1) vehicle control (n = 10), (2) CBD (n = 9), (3) DPCPX (n = 7), and (4) CBD + DPCPX group (n = 7). Cannabidiol treatment significantly decreased the incidence and the duration of ventricular tachycardia, total length of arrhythmias, and the arrhythmia scores compared to control during the reperfusion period. The DPCPX treatment alone did not affect the incidence and the duration of any type of arrhythmias. However, DPCPX aborted the antiarrhythmic effect of CBD when it was combined with it. The present results demonstrated that CBD has an antiarrhythmic effect against I/R-induced arrhythmias, and the antiarrhythmic effect of CBD may be mediated through the activation of adenosine

  2. A1 and A2a receptors mediate inhibitory effects of adenosine on the motor activity of human colon.

    PubMed

    Fornai, M; Antonioli, L; Colucci, R; Ghisu, N; Buccianti, P; Marioni, A; Chiarugi, M; Tuccori, M; Blandizzi, C; Del Tacca, M

    2009-04-01

    Experimental evidence in animal models suggests that adenosine is involved in the regulation of digestive functions. This study examines the influence of adenosine on the contractile activity of human colon. Reverse transcription-polymerase chain reaction revealed A(1) and A(2a) receptor expression in colonic neuromuscular layers. Circular muscle preparations were connected to isotonic transducers to determine the effects of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; A(1) receptor antagonist), ZM 241385 (A(2a) receptor antagonist), CCPA (A(1) receptor agonist) and 2-[(p-2-carboxyethyl)-phenethylamino]-5'-N-ethyl-carboxamide-adenosine (CGS 21680; A(2a) receptor agonist) on motor responses evoked by electrical stimulation or carbachol. Electrically evoked contractions were enhanced by DPCPX and ZM 241385, and reduced by CCPA and CGS 21680. Similar effects were observed when colonic preparations were incubated with guanethidine (noradrenergic blocker), L-732,138, GR-159897 and SB-218795 (NK receptor antagonists). However, in the presence of guanethidine, NK receptor antagonists and N(omega)-propyl-L-arginine (NPA; neuronal nitric oxide synthase inhibitor), the effects of DPCPX and CCPA were still evident, while those of ZM 241385 and CGS 21680 no longer occurred. Carbachol-induced contractions were unaffected by A(2a) receptor ligands, but they were enhanced or reduced by DPCPX and CCPA, respectively. When colonic preparations were incubated with guanethidine, NK antagonists and atropine, electrically induced relaxations were partly reduced by ZM 241385 or NPA, but unaffected by DPCPX. Dipyridamole or application of exogenous adenosine reduced electrically and carbachol-evoked contractions, whereas adenosine deaminase enhanced such motor responses. In conclusion, adenosine exerts an inhibitory control on human colonic motility. A(1) receptors mediate direct modulating actions on smooth muscle, whereas A(2a) receptors operate through inhibitory nitrergic nerve pathways.

  3. Mechanisms of the adenosine A2A receptor-induced sensitization of esophageal C fibers

    PubMed Central

    Brozmanova, M.; Mazurova, L.; Ru, F.; Tatar, M.; Hu, Y.; Yu, S.

    2015-01-01

    Clinical studies indicate that adenosine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the adenosine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selective agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10–60 mmHg) in a concentration-dependent fashion (1–100 nM). At the half-maximally effective concentration (EC50: ≈3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mimicked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist α,β-methylene-ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demonstrated that CGS21682 potentiated TRPA1 currents evoked by allylisothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involvement of TRPA1. PMID:26564719

  4. Involvement of A(1) adenosine receptors in osmotic volume regulation of retinal glial cells in mice.

    PubMed

    Wurm, Antje; Lipp, Stephan; Pannicke, Thomas; Linnertz, Regina; Färber, Katrin; Wiedemann, Peter; Reichenbach, Andreas; Bringmann, Andreas

    2009-09-12

    Osmotic swelling of Müller glial cells has been suggested to contribute to retinal edema. We determined the role of adenosine signaling in the inhibition of Müller cell swelling in the murine retina. The size of Müller cell somata was recorded before and during perfusion of retinal sections and isolated Müller cells with a hypoosmolar solution. Retinal tissues were freshly isolated from wild-type mice and mice deficient in A(1) adenosine receptors (A(1)AR(-/-)), or cultured as whole-mounts for three days. The potassium conductance of Müller cells was recorded in isolated cells, and retinal slices were immunostained against Kir4.1. Hypotonic exposure for 4 min induced a swelling of Müller cell bodies in retinal slices from A(1)AR(-/-) mice but not wild-type mice. Pharmacological inhibition of A(1) receptors or of the ecto-5'-nucleotidase induced hypoosmotic swelling of Müller cells from wild-type mice. Exogenous adenosine prevented the swelling of Müller cells from wild-type but not A(1)AR(-/-) mice. The antiinflammatory corticosteroid, triamcinolone acetonide, inhibited the swelling of Müller cells from wild-type mice; this effect was blocked by an antagonist of A(1) receptors. The potassium conductance of Müller cells and the Kir4.1 immunolabeling of retinal slices were not different between A(1)AR(-/-) and wild-type mice, both in freshly isolated tissues and retinal organ cultures. The data suggest that autocrine activation of A(1) receptors by extracellularly generated adenosine mediates the volume homeostasis of Müller cells in the murine retina. The swelling-inhibitory effect of triamcinolone is mediated by enhancement of endogenous adenosine signaling.

  5. Involvement of A1 adenosine receptors in osmotic volume regulation of retinal glial cells in mice

    PubMed Central

    Wurm, Antje; Lipp, Stephan; Pannicke, Thomas; Linnertz, Regina; Färber, Katrin; Wiedemann, Peter; Reichenbach, Andreas

    2009-01-01

    Purpose Osmotic swelling of Müller glial cells has been suggested to contribute to retinal edema. We determined the role of adenosine signaling in the inhibition of Müller cell swelling in the murine retina. Methods The size of Müller cell somata was recorded before and during perfusion of retinal sections and isolated Müller cells with a hypoosmolar solution. Retinal tissues were freshly isolated from wild-type mice and mice deficient in A1 adenosine receptors (A1AR−/−), or cultured as whole-mounts for three days. The potassium conductance of Müller cells was recorded in isolated cells, and retinal slices were immunostained against Kir4.1. Results Hypotonic exposure for 4 min induced a swelling of Müller cell bodies in retinal slices from A1AR−/− mice but not wild-type mice. Pharmacological inhibition of A1 receptors or of the ecto-5′-nucleotidase induced hypoosmotic swelling of Müller cells from wild-type mice. Exogenous adenosine prevented the swelling of Müller cells from wild-type but not A1AR−/− mice. The antiinflammatory corticosteroid, triamcinolone acetonide, inhibited the swelling of Müller cells from wild-type mice; this effect was blocked by an antagonist of A1 receptors. The potassium conductance of Müller cells and the Kir4.1 immunolabeling of retinal slices were not different between A1AR−/− and wild-type mice, both in freshly isolated tissues and retinal organ cultures. Conclusions The data suggest that autocrine activation of A1 receptors by extracellularly generated adenosine mediates the volume homeostasis of Müller cells in the murine retina. The swelling-inhibitory effect of triamcinolone is mediated by enhancement of endogenous adenosine signaling. PMID:19756184

  6. Mechanisms of the adenosine A2A receptor-induced sensitization of esophageal C fibers.

    PubMed

    Brozmanova, M; Mazurova, L; Ru, F; Tatar, M; Hu, Y; Yu, S; Kollarik, M

    2016-02-01

    Clinical studies indicate that adenosine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the adenosine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selective agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10-60 mmHg) in a concentration-dependent fashion (1-100 nM). At the half-maximally effective concentration (EC50: ≈3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mimicked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist α,β-methylene-ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demonstrated that CGS21682 potentiated TRPA1 currents evoked by allylisothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involvement of TRPA1.

  7. Cloning of two adenosine receptor subtypes from mouse bone marrow-derived mast cells.

    PubMed

    Marquardt, D L; Walker, L L; Heinemann, S

    1994-05-01

    Adenosine potentiates the stimulated release of mast cell mediators. Pharmacologic studies suggest the presence of two adenosine receptors, one positively coupled to adenylate cyclase and the other coupled to phospholipase C activation. To identify mast cell adenosine receptor subtypes, cDNAs for the A1 and A2a adenosine receptors were obtained by screening a mouse brain cDNA library with the use of PCR-derived probes. Mouse bone marrow-derived mast cell cDNA libraries were constructed and screened with the use of A1 and A2a cDNA probes, which revealed the presence of A2a, but not A1, receptor clones. A putative A2b receptor was identified by using low stringency mast cell library screening. Northern blotting of mast cell poly(A)+ RNA with the use of receptor subtype probes labeled single mRNA bands of 2.4 kb and 1.8 kb for the A2a and A2b receptors, respectively. In situ cells. An A2a receptor-specific agonist failed to enhance mast cell mediator release, which suggests that the secretory process is modulated through the A2b and/or another receptor subtype. By using RNase protection assays, we found that mast cells that had been cultured in the presence of N-ethylcarboxamidoadenosine for 24 h exhibited a decrease in both A2a and A2b receptor RNA levels. Cells that had been cultured for 1 to 2 days in the presence of dexamethasone demonstrated increased amounts of A2a receptor mRNA, but no identifiable change in A2b receptor mRNA. Mast cells possess at least two adenosine receptor subtypes that may be differentially regulated.

  8. Phentolamine prevents the adverse effects of adenosine on glycolysis and mechanical function in isolated working rat hearts subjected to antecedent ischemia.

    PubMed

    Finegan, B A; Gandhi, M; Clanachan, A S

    2000-06-01

    Adenosine inhibits glycolysis from exogenous glucose, reduces proton production and enhances post-ischemic left ventricular minute work (LV work) following ischemia in isolated working rat hearts perfused with glucose and fatty acids. In hearts partially depleted of glycogen by antecedent ischemic stress (AIS)--two cycles of ischemia (10 min) and reperfusion (5 min)--adenosine stimulates rather than inhibits glycolysis, increases proton production and worsens recovery of post-ischemic LV work. We determined if the switch in adenosine effect on glycolysis and recovery of LV work following ischemia in hearts subject to AIS was due to the reduction in glycogen content per se or because of alpha-adrenoceptor stimulation. One series of hearts underwent a 35-min period of substrate-free Langendorff perfusion (substrate-free glycogen depletion; SFGD) and a second series of hearts was subjected to AIS. Both series of hearts had a similar glycogen content (approximately 70 micromol/g dry wt) prior to drug treatment. In SFGD hearts perfused aerobically, adenosine (500 microM) inhibited glycolysis from exogenous glucose and reduced proton production. In SFGD hearts reperfused after prolonged ischemia, adenosine exerted similar effects on glucose metabolism and enhanced recovery of post-ischemic LV work (87.2 +/- 2.2% of preischemic values) relative to untreated hearts (25.9 +/- 13.3% of preischemic values). In AIS hearts perfused aerobically or subject to ischemia and reperfusion, phentolamine (1 microM) given in combination with adenosine, prevented adenosine-induced stimulation of glycolysis from exogenous glucose and reduced calculated proton production from glucose. Recoveries of post-ischemic LV work in AIS hearts for untreated, adenosine, phentolamine and adenosine/phentolamine groups were 34.4 +/- 11.4%, 8.6 +/- 3.9%, 16.3 +/- 13.5% and 73.2 +/- 13.1% respectively, of preischemic values. Glycogen depletion in the absence of ischemia does not switch the effect of

  9. Probing the Binding Site of the A1 Adenosine Receptor Reengineered for Orthogonal Recognition by Tailored Nucleosides

    PubMed Central

    Palaniappan, Krishnan K.; Gao, Zhan-Guo; Ivanov, Andrei A.; Greaves, Rebecca; Adachi, Hayamitsu; Besada, Pedro; Kim, Hea Ok; Kim, Ae Yil; Choe, Seung Ah; Jeong, Lak Shin; Jacobson, Kenneth A.

    2011-01-01

    His272 (7.43) in the seventh transmembrane domain (TM7) of the human A3 adenosine receptor (AR) interacts with the 3′ position of nucleosides, based on selective affinity enhancement at a H272E mutant A3 AR (neoceptor) of 3′-ureido, but not 3′-OH, adenosine analogues. Here, mutation of the analogous H278 of the human A1 AR to Ala, Asp, Glu, or Leu enhanced the affinity of novel 2′- and 3′-ureido adenosine analogues, such as 10 (N6-cyclopentyl-3′-ureido-3′-deoxyadenosine), by >100-fold, while decreasing the affinity or potency of adenosine and other 3′-OH adenosine analogues. His278 mutant receptors produced a similar enhancement regardless of the charge character of the substituted residue, implicating steric rather than electrostatic factors in the gain of function, a hypothesis supported by rhodopsin-based molecular modeling. It was also demonstrated that this interaction was orientationally specific; i.e., mutations at the neighboring Thr277 did not enhance the affinity for a series of 2′- and 3′-ureido nucleosides. Additionally, H-bonding groups placed on substituents at the N6 or 5′ position demonstrated no enhancement in the mutant receptors. These reengineered human A1 ARs revealed orthogonality similar to that of the A3 but not the A2A AR, in which mutation of the corresponding residue, His278, to Asp did not enhance nucleoside affinity. Functionally, the H278D A1 AR was detectable only in a measure of membrane potential and not in calcium mobilization. This neoceptor approach should be useful for the validation of molecular modeling and the dissection of promiscuous GPCR signaling. PMID:17542617

  10. Elevated adenosine signaling via adenosine A2B receptor induces normal and sickle erythrocyte sphingosine kinase 1 activity.

    PubMed

    Sun, Kaiqi; Zhang, Yujin; Bogdanov, Mikhail V; Wu, Hongyu; Song, Anren; Li, Jessica; Dowhan, William; Idowu, Modupe; Juneja, Harinder S; Molina, Jose G; Blackburn, Michael R; Kellems, Rodney E; Xia, Yang

    2015-03-05

    Erythrocyte possesses high sphingosine kinase 1 (SphK1) activity and is the major cell type supplying plasma sphingosine-1-phosphate, a signaling lipid regulating multiple physiological and pathological functions. Recent studies revealed that erythrocyte SphK1 activity is upregulated in sickle cell disease (SCD) and contributes to sickling and disease progression. However, how erythrocyte SphK1 activity is regulated remains unknown. Here we report that adenosine induces SphK1 activity in human and mouse sickle and normal erythrocytes in vitro. Next, using 4 adenosine receptor-deficient mice and pharmacological approaches, we determined that the A2B adenosine receptor (ADORA2B) is essential for adenosine-induced SphK1 activity in human and mouse normal and sickle erythrocytes in vitro. Subsequently, we provide in vivo genetic evidence that adenosine deaminase (ADA) deficiency leads to excess plasma adenosine and elevated erythrocyte SphK1 activity. Lowering adenosine by ADA enzyme therapy or genetic deletion of ADORA2B significantly reduced excess adenosine-induced erythrocyte SphK1 activity in ADA-deficient mice. Finally, we revealed that protein kinase A-mediated extracellular signal-regulated kinase 1/2 activation functioning downstream of ADORA2B underlies adenosine-induced erythrocyte SphK1 activity. Overall, our findings reveal a novel signaling network regulating erythrocyte SphK1 and highlight innovative mechanisms regulating SphK1 activity in normal and SCD.

  11. Separation of adenosine diphosphate--adenosine triphosphate-exchange activity from the cerebral microsomal sodium-plus-potassium ion-stimulated adenosine triphosphatase.

    PubMed

    Stahl, W L; Sattin, A; McIlwain, H

    1966-05-01

    1. A microsomal fraction from ox cerebral cortex catalysed [(14)C]ADP-ATP exchange at a speed similar to that at which it liberated P(i) from ATP in the presence of Na(+), K(+) and Mg(2+). 2. Repeated washing the fraction with MgATP solutions solubilized most of the exchange activity and left the adenosine triphosphatase insoluble and little changed in activity. The exchange activity was accompanied by negligible adenosine-triphosphatase activity and was enriched by precipitation at chosen pH and by DEAE-Sephadex. At no stage was its activity affected by Na(+), K(+) or ouabain. 3. The washed microsomal fraction was exposed to a variety of reagents; a sodium iodide-cysteine treatment increased both adenosine-triphosphatase and exchange activities, as also did a synthetic zeolite. Preparations were obtained with exchange activities less than 3% of their Na(+)-plus-K(+)-stimulated adenosine-triphosphatase activity. Some contribution to the residual exchange activity was made by an adenylate kinase. 4. Thus over 95% of the microsomal ADP-ATP-exchange activity does not take part in the Na(+)-plus-K(+)-stimulated adenosine-triphosphatase reaction. Participation of some of the residual 3% of the ADP-ATP-exchange activity has not been excluded, but there appears no firm evidence for its participation in the adenosine triphosphatase; the bearing of this conclusion on mechanisms proposed for the Na(+)-plus-K(+)-stimulated adenosine triphosphatase is indicated.

  12. Separation of adenosine diphosphate-adenosine triphosphate–exchange activity from the cerebral microsomal sodium-plus-potassium ion-stimulated adenosine triphosphatase

    PubMed Central

    Stahl, W. L.; Sattin, A.; McIlwain, H.

    1966-01-01

    1. A microsomal fraction from ox cerebral cortex catalysed [14C]ADP–ATP exchange at a speed similar to that at which it liberated Pi from ATP in the presence of Na+, K+ and Mg2+. 2. Repeated washing the fraction with MgATP solutions solubilized most of the exchange activity and left the adenosine triphosphatase insoluble and little changed in activity. The exchange activity was accompanied by negligible adenosine-triphosphatase activity and was enriched by precipitation at chosen pH and by DEAE-Sephadex. At no stage was its activity affected by Na+, K+ or ouabain. 3. The washed microsomal fraction was exposed to a variety of reagents; a sodium iodide–cysteine treatment increased both adenosine-triphosphatase and exchange activities, as also did a synthetic zeolite. Preparations were obtained with exchange activities less than 3% of their Na+-plus-K+-stimulated adenosine-triphosphatase activity. Some contribution to the residual exchange activity was made by an adenylate kinase. 4. Thus over 95% of the microsomal ADP–ATP-exchange activity does not take part in the Na+-plus-K+-stimulated adenosine-triphosphatase reaction. Participation of some of the residual 3% of the ADP–ATP-exchange activity has not been excluded, but there appears no firm evidence for its participation in the adenosine triphosphatase; the bearing of this conclusion on mechanisms proposed for the Na+-plus-K+-stimulated adenosine triphosphatase is indicated. PMID:4223577

  13. Adenosine: Tipping the balance towards hepatic steatosis and fibrosis

    PubMed Central

    Robson, Simon C.; Schuppan, Detlef

    2010-01-01

    Fatty liver is commonly associated with alcohol ingestion and abuse. While the molecular pathogenesis of these fatty changes is well understood, the histochemical and pharmacological mechanisms by which ethanol stimulates these molecular changes remain unknown. During ethanol metabolism, adenosine is generated by the enzyme ecto-5′-nucleotidase, and adenosine production and adenosine receptor activation are known to play critical roles in the development of hepatic fibrosis. We therefore investigated whether adenosine and its receptors play a role in the development of alcohol-induced fatty liver. WT mice fed ethanol on the Lieber-DeCarli diet developed hepatic steatosis, including increased hepatic triglyceride content, while mice lacking ecto-5-nucleotidase or adenosine A1 or A2B receptors were protected from developing fatty liver. Similar protection was also seen in WT mice treated with either an adenosine A1 or A2B receptor antagonist. Steatotic livers demonstrated increased expression of genes involved in fatty acid synthesis, which was prevented by blockade of adenosine A1 receptors, and decreased expression of genes involved in fatty acid metabolism, which was prevented by blockade of adenosine A2B receptors. In vitro studies supported roles for adenosine A1 receptors in promoting fatty acid synthesis and for A2B receptors in decreasing fatty acid metabolism. These results indicate that adenosine generated by ethanol metabolism plays an important role in ethanol-induced hepatic steatosis via both A1 and A2B receptors and suggest that targeting adenosine receptors may be effective in the prevention of alcohol-induced fatty liver. PMID:20395005

  14. Characterization of adenosine binding proteins in human placental membranes

    SciTech Connect

    Hutchison, K.A.

    1989-01-01

    We have characterized two adenosine binding proteins in human placenta. In membranes, one site is detected with ({sup 3}H) -N-ethylcarboxamidoadenosine (({sup 3}H)NECA). This site is similar to the adenosine A{sub 2} receptor. We call this site the adenosine A{sub 2}-like binding site. In detergent extracts, the second site is detected and has the characteristics of an adenosine A{sub 1} receptor. The soluble adenosine A{sub 2}-like binding site cannot be detected without a rapid assay. Binding to the adenosine A{sub 1} receptor with ({sup 3}H)-2-chloroadenosine and ({sup 3}H)NECA is time dependent, saturable, and reversible. Equilibrium displacement analysis with adenosine agonists reveals an A{sub 1} specificity: 2-chloroadenosine > R-phenylisopropyladenosine > 5{prime}-N-ethylcarboxamidoadenosine. The antagonist potency order is 1,3-diethyl-8-phenylxanthine > isobutylmethylxanthine > theophylline. Competition analysis of membranes with the A,-selective ligands ({sup 3}H)-cyclohexyladenosine ({sup 3}H) cylopentylxanthine revealed adenosine A{sub 1} agonist and antagonist potency orders. We have purified the adenosine A{sub 2}-like binding site. The adenosine A{sub 2}-like binding site is an ubiquitous major cellular protein. It is glycosylated, highly asymmetric, and acidic. The native protein is an homodimer with a subunit molecular mass of 98 kDa. The sedimentation coefficient and partial specific volume of the binding complex are 6.9 s and 0.698 ml/g, respectively. The Stokes' radius is 70 {Angstrom}. The native molecular mass of the detergent-protein complex is 230 kDa. The adenosine A{sub 2}-like binding site has an agonist potency order of 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine >> R-phenylisopropyladenosine and an antagonist potency order of isobutylmethylxanthine > theophylline >> 1,3-diethyl-8-phenylxanthine.

  15. Repetitive systemic morphine alters activity-dependent plasticity of Schaffer-collateral-CA1 pyramidal cell synapses: involvement of adenosine A1 receptors and adenosine deaminase.

    PubMed

    Sadegh, Mehdi; Fathollahi, Yaghoub

    2014-10-01

    The effectiveness of O-pulse stimulation (TPS) for the reversal of O-pattern primed bursts (PB)-induced long-term potentiation (LTP) were examined at the Schaffer-collateral-CA1 pyramidal cell synapses of hippocampal slices derived from rats chronically treated with morphine (M-T). The results showed that slices derived from both control and M-T rats had normal field excitatory postsynaptic potential (fEPSP)-LTP, whereas PS-LTP in slices from M-T rats was significantly greater than that from control slices. When morphine was applied in vitro to slices derived from rats chronically treated with morphine, the augmentation of PS-LTP was not seen. TPS given 30 min after LTP induction failed to reverse the fEPSP- or PS-LTP in both groups of slices. However, TPS delivered in the presence of long-term in vitro morphine caused the PS-LTP reversal. This effect was blocked by the adenosine A1 receptor (A1R) antagonist CPX (200 nM) and furthermore was enhanced by the adenosine deaminase (ADA) inhibitor EHNA (10 μM). Interestingly, TPS given 30 min after LTP induction in the presence of EHNA (10 μM) can reverse LTP in morphine-exposed control slices in vitro. These results suggest adaptive changes in the hippocampus area CA1 in particular in adenosine system following repetitive systemic morphine. Chronic in vivo morphine increases A1R and reduces ADA activity in the hippocampus. Consequently, adenosine can accumulate because of a stimulus train-induced activity pattern in CA1 area and takes the opportunity to work as an inhibitory neuromodulator and also to enable CA1 to cope with chronic morphine. In addition, adaptive mechanisms are differentially working in the dendrite layer rather than the somatic layer of hippocampal CA1.

  16. Adenosine produced from adenine nucleotides through an interaction between apoptotic cells and engulfing macrophages contributes to the appearance of transglutaminase 2 in dying thymocytes.

    PubMed

    Sándor, Katalin; Pallai, Anna; Duró, Edina; Legendre, Pascal; Couillin, Isabelle; Sághy, Tibor; Szondy, Zsuzsa

    2017-03-01

    Transglutaminase 2 (TG2) has been known for a long time to be associated with the in vivo apoptosis program of various cell types, including T cells. Though the expression of the enzyme is strongly induced in mouse thymocytes following apoptosis induction in vivo, no significant induction of TG2 can be detected, when thymocytes are induced to die by the same stimuli in vitro indicating that signals arriving from the tissue environment are required for the proper in vivo induction of the enzyme. Previous studies from our laboratory have demonstrated that two of these signals, transforming growth factor-β (TGF-β) and retinoids, are produced by macrophages engulfing apoptotic cells. However, in addition to TGF-β and retinoids, engulfing macrophages produce adenosine as well. Here, we show that in vitro adenosine, adenosine, and retinoic acid or adenosine, TGF-β and retinoic acids together can significantly enhance the TG2 mRNA expression in dying thymocytes. The effect of adenosine is mediated via adenosine A2A receptors (A2ARs) and the A2AR-triggered adenylate cyclase signaling pathway. In accordance, loss of A2ARs in A2AR null mice significantly attenuates the in vivo induction of TG2 following apoptosis induction in the thymus indicating that adenosine indeed contributes in vivo to the apoptosis-related appearance of the enzyme. We also demonstrate that adenosine is produced extracellularly during engulfment of apoptotic thymocytes, partly from adenine nucleotides released via thymocyte pannexin-1 channels. Our data reveal a novel crosstalk between macrophages and apoptotic cells, in which apoptotic cell uptake-related adenosine production contributes to the appearance of TG2 in the dying thymocytes.

  17. Modulation of dopamine-mediated facilitation at the neuromuscular junction of Wistar rats: A role for adenosine A1/A2A receptors and P2 purinoceptors.

    PubMed

    Elnozahi, Neveen A; AlQot, Hadir E; Mohy El-Din, Mahmoud M; Bistawroos, Azza E; Abou Zeit-Har, Mohamed S

    2016-06-21

    This study aims to understand how dopamine and the neuromodulators, adenosine and adenosine triphosphate (ATP) modulate neuromuscular transmission. Adenosine and ATP are well-recognized for their regulatory effects on dopamine in the central nervous system. However, if similar interactions occur at the neuromuscular junction is unknown. We hypothesize that the activation of adenosine A1/A2A and/or P2 purinoceptors may influence the action of dopamine on neuromuscular transmission. Using the rat phrenic nerve hemi-diaphragm, we assessed the influence of dopamine, adenosine and ATP on the height of nerve-evoked muscle twitches. We investigated how the selective blockade of adenosine A1 receptors (2.5nM DPCPX), adenosine A2A receptors (50nM CSC) and P2 purinoceptors (100μM suramin) modified the effects of dopamine. Dopamine alone increased indirect muscle contractions while adenosine and ATP either enhanced or depressed nerve-evoked muscle twitches in a concentration-dependent manner. The facilitatory effects of 256μM dopamine were significantly reduced to 29.62±2.79% or 53.69±5.45% in the presence of DPCPX or CSC, respectively, relative to 70.03±1.57% with dopamine alone. Alternatively, the action of 256μM dopamine was potentiated from 70.03±1.57, in the absence of suramin, to 86.83±4.36%, in the presence of suramin. It can be concluded that the activation of adenosine A1 and A2A receptors and P2 purinoceptors potentially play a central role in the regulation of dopamine effects at the neuromuscular junction. Clinically this study offers new insights for the indirect manipulation of neuromuscular transmission for the treatment of disorders characterized by motor dysfunction. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  18. High salt diet exacerbates vascular contraction in the absence of adenosine A2A receptor

    PubMed Central

    Pradhan, Isha; Zeldin, Darryl C.; Ledent, Catherine; Mustafa, S. Jamal; Falck, John R.; Nayeem, Mohammed A

    2014-01-01

    High salt (4%NaCl, HS) diet modulates adenosine-induced vascular response through adenosine A2A-receptor (A2AAR). Evidence suggests A2AAR stimulates cyp450-epoxygenases, leading to epoxyeicosatrienoic acids (EETs) generation. The aim of this study was to understand the vascular reactivity to HS and underlying signaling mechanism in the presence or absence of A2AAR. Therefore, we hypothesized that HS enhances adenosine-induced relaxation through EETs in A2AAR+/+, but exaggerates contraction in A2AAR−/−. Organ-bath and Western-blot experiments were conducted in HS and normal salt (NS, 0.18% NaCl)-fed A2AAR+/+ and A2AAR−/− mice aortae. HS produced concentration-dependent relaxation to non-selective adenosine analog, NECA in A2AAR+/+, whereas contraction was observed in A2AAR−/− mice and this was attenuated by A1AR antagonist (DPCPX). CGS-21680 (selective A2AAR-agonist) enhanced relaxation in HS-A2AAR+/+ vs. NS-A2AAR+/+, that was blocked by EETs antagonist (14,15-EEZE). Compared to NS, HS significantly upregulated expression of vasodilators A2AAR and cyp2c29, while vasoconstrictors A1AR and cyp4a in A2AAR+/+ were downregulated. In A2AAR−/− mice, however, HS significantly downregulated the expression of cyp2c29, while A1AR and cyp4a were upregulated compared to A2AAR+/+ mice. Hence, our data suggest that in A2AAR+/+, HS enhances A2AAR-induced relaxation through increased cyp-expoxygenases-derived EETs and decreased A1AR levels, whereas in A2AAR−/−, HS exaggerates contraction through decreased cyp-epoxygenases and increased A1AR levels. PMID:24390173

  19. Magnetite nanoparticle-induced fluorescence quenching of adenosine triphosphate-BODIPY Conjugates: application to adenosine triphosphate and pyrophosphate sensing.

    PubMed

    Yu, Cheng-Ju; Wu, Su-Mei; Tseng, Wei-Lung

    2013-09-17

    We report that magnetite nanoparticles (Fe3O4 NPs) act as an efficient quencher for boron dipyrromethene-conjugated adenosine 5'-triphosphate (BODIPY-ATP) that is highly fluorescent in bulk solution. BODIPY-ATP molecules attached to the surface of Fe3O4 NPs through the coordination between the triphosphate group of BODIPY-ATP and Fe(3+)/Fe(2+) on the NP surface. The formed complexes induced an apparent reduction in the BODIPY-ATP fluorescence resulting from an oxidative-photoinduced electron transfer (PET) from the BODIPY-ATP excited state to an unfilled d shell of Fe(3+)/Fe(2+) on the NP surface. A comparison of the Stern-Volmer quenching constant between Fe(3+) and Fe(2+) suggests that Fe(3+) on the NP surface dominantly controls this quenching process. The efficiency for Fe3O4 NP-induced fluorescence quenching of the BODIPY-ATP was enhanced by increasing the concentration of Fe3O4 NPs and lowering the pH of the solution to below 6.0. We found that pyrophosphate and ATP compete with BODIPY-ATP for binding to Fe3O4 NPs. Thus, we amplified BODIPY-ATP fluorescence in the presence of increasing the pyrophosphate and ATP concentration; the detection limits at a signal-to-noise ratio of 3 for pyrophosphate and ATP were determined to be 7 and 30 nM, respectively. The Fe3O4 NP-based competitive binding assay detected ATP and pyrophosphate in only 5 min. The selectivity of this assay for ATP over metal ions, amino acids, and adenosine analogues is particularly high. The practicality of using the developed method to determine ATP in a single drop of blood is also validated.

  20. Modulation of adenylyl cyclase by FPP and adenosine involves stimulatory and inhibitory adenosine receptors and g proteins.

    PubMed

    Fraser, L R; Adeoya-Osiguwa, S

    1999-08-01

    FPP and adenosine modulate the adenylyl cyclase (AC)/cAMP signal transduction pathway in mammalian spermatozoa to elicit a biphasic response, initially stimulating capacitation and then inhibiting spontaneous acrosome loss. This study addressed the hypothesis that responses to FPP involve interactions between receptors for FPP and adenosine, the biphasic responses involving stimulatory and inhibitory adenosine receptors. Gln-FPP, a competitive inhibitor of FPP, significantly inhibited binding of an adenosine analogue and responses to adenosine, especially in capacitated suspensions, consistent with interaction between FPP and adenosine receptors. CGS-21680 (1 microM), a stimulatory A2a adenosine receptor agonist, significantly stimulated capacitation and cAMP in uncapacitated cells, while cyclopentyl adenosine (1 microM), an inhibitory A1 adenosine receptor agonist only affected capacitated cells, inhibiting spontaneous acrosome loss. Responses to FPP and adenosine were inhibited in uncapacitated cells by a selective A2a antagonist and in capacitated cells by a selective A1 antagonist; subsequent investigations indicated possible involvement of G proteins. Like FPP, cholera toxin stimulated capacitation and cAMP production in uncapacitated cells, suggesting involvement of a G protein with a Galphas subunit. In contrast, pertussis toxin prevented FPP's inhibition of both spontaneous acrosome loss and cAMP production, suggesting involvement of a Galphai/o subunit. Immunoblotting evidence revealed the presence of proteins of the appropriate molecular weights for Galphas, Galphai2, Galpha i3, and Galphao subunits. This study provides the first direct evidence suggesting the involvement of two different types of adenosine receptors and both Galphas and Galphai/o subunits in the regulation of capacitation, resulting in modulation of AC activity and availability of cAMP. Copyright 1999 Wiley-Liss, Inc.

  1. THE TIME COURSE OF ADENOSINE, NITRIC OXIDE (NO) AND INDUCIBLE NO SYNTHASE CHANGES IN THE BRAIN WITH SLEEP LOSS AND THEIR ROLE IN THE NREM SLEEP HOMEOSTATIC CASCADE

    PubMed Central

    Kalinchuk, Anna V.; McCarley, Robert W.; Porkka-Heiskanen, Tarja; Basheer, Radhika

    2011-01-01

    Both adenosine and nitric oxide (NO) are known for their role in sleep homeostasis, with the basal forebrain (BF) wakefulness center as an important site of action. Previously we reported a cascade of homeostatic events, wherein sleep deprivation (SD) induces the production of inducible nitric oxide synthase (iNOS)-dependent NO in BF, leading to enhanced release of extracellular adenosine. In turn, increased BF adenosine leads to enhanced sleep intensity, as measured by increased non-rapid eye movement (NREM) EEG delta activity. However, the presence and time course of similar events in cortex has not been studied, although a frontal cortical role for the increase in NREM recovery sleep EEG delta power is known. Accordingly, we performed simultaneous hourly microdialysis sample collection from BF and frontal cortex (FC) during 11h SD. We observed that both areas showed sequential increases in iNOS and NO, followed by increases in adenosine. BF increases began at 1h SD, while FC increases began at 5h SD. iNOS and Fos-double labeling indicated that iNOS induction occurred in BF and FC wake-active neurons. These data support the role of BF adenosine and NO in sleep homeostasis and indicate the temporal and spatial sequence of sleep homeostatic cascade for NO and adenosine. PMID:21062286

  2. Stabilizing effects of G protein on the active conformation of adenosine A1 receptor differ depending on G protein type.

    PubMed

    Tateyama, Michihiro; Kubo, Yoshihiro

    2016-10-05

    G protein coupled receptors (GPCRs) trigger various cellular and physiological responses upon the ligand binding. The ligand binding induces conformational change in GPCRs which allows G protein to interact with the receptor. The interaction of G protein also affects the active conformation of GPCRs. In this study, we have investigated the effects of Gαi1, Gαo and chimeric Gαqi5 on the active conformation of the adenosine A1 receptor, as each Gα showed difference in the interaction with adenosine A1 receptor. The conformational changes in the adenosine A1 receptor were detected as the agonist-induced decreases in efficiency of Förster resonance energy transfer (FRET) between fluorescent proteins (FPs) fused at the two intracellular domains of the adenosine A1 receptor. Amplitudes of the agonist-induced FRET decreases were subtle when the FP-tagged adenosine A1 receptor was expressed alone, whereas they were significantly enhanced when co-expressed with Gαi1Gβ1Gγ22 (Gi1) or Gαqi5Gβ1Gγ22 (Gqi5) but not with GαοGβ1Gγ22 (Go). The enhancement of the agonist-induced FRET decrease in the presence of Gqi5 was significantly larger than that of Gi1. Furthermore, the FRET recovery upon the agonist removal in the presence of Gqi5 was significantly slower than that of Gi1. From these results it was revealed that the agonist-bound active conformation of adenosine A1 receptor is unstable without the binding of G protein and that the stabilizing effects of G protein differ depending on the types of G protein.

  3. Comorbidities in Neurology: Is Adenosine the Common Link?

    PubMed Central

    Boison, Detlev; Aronica, Eleonora

    2015-01-01

    Comorbidities in Neurology represent a major conceptual and therapeutic challenge. For example, temporal lobe epilepsy (TLE) is a syndrome comprised of epileptic seizures and comorbid symptoms including memory and psychiatric impairment, depression, and sleep dysfunction. Similarly, Alzheimer’s disease (AD), Parkinson’s disease (PD), and Amyotrophic Lateral Sclerosis (ALS) are accompanied by various degrees of memory dysfunction. Patients with AD have an increased likelihood for seizures, whereas all four conditions share certain aspects of psychosis, depression, and sleep dysfunction. This remarkable overlap suggests common pathophysiological mechanisms, which include synaptic dysfunction and synaptotoxicity, as well as glial activation and astrogliosis. Astrogliosis is linked to synapse function via the tripartite synapse, but astrocytes also control the availability of gliotransmitters and adenosine. Here we will specifically focus on the ‘adenosine hypothesis of comorbidities’ implying that astrocyte activation, via overexpression of adenosine kinase (ADK), induces a deficiency in the homeostatic tone of adenosine. We present evidence from patient-derived samples showing astrogliosis and overexpression of ADK as common pathological hallmark of epilepsy, AD, PD, and ALS. We discuss a transgenic ‘comorbidity model’, in which brain-wide overexpression of ADK and resulting adenosine deficiency produces a comorbid spectrum of seizures, altered dopaminergic function, attentional impairment, and deficits in cognitive domains and sleep regulation. We conclude that dysfunction of adenosine signaling is common in neurological conditions, that adenosine dysfunction can explain comorbid phenotypes, and that therapeutic adenosine augmentation might be effective for the treatment of comorbid symptoms in multiple neurological conditions. PMID:25979489

  4. Elevated placental adenosine signaling contributes to the pathogenesis of preeclampsia.

    PubMed

    Iriyama, Takayuki; Sun, Kaiqi; Parchim, Nicholas F; Li, Jessica; Zhao, Cheng; Song, Anren; Hart, Laura A; Blackwell, Sean C; Sibai, Baha M; Chan, Lee-Nien L; Chan, Teh-Sheng; Hicks, M John; Blackburn, Michael R; Kellems, Rodney E; Xia, Yang

    2015-02-24

    Preeclampsia is a prevalent hypertensive disorder of pregnancy and a leading cause of maternal and neonatal morbidity and mortality worldwide. This pathogenic condition is speculated to be caused by placental abnormalities that contribute to the maternal syndrome. However, the specific factors and signaling pathways that lead to impaired placentas and maternal disease development remain elusive. Using 2 independent animal models of preeclampsia (genetically engineered pregnant mice with elevated adenosine exclusively in placentas and a pathogenic autoantibody-induced preeclampsia mouse model), we demonstrated that chronically elevated placental adenosine was sufficient to induce hallmark features of preeclampsia, including hypertension, proteinuria, small fetuses, and impaired placental vasculature. Genetic and pharmacological approaches revealed that elevated placental adenosine coupled with excessive A₂B adenosine receptor (ADORA2B) signaling contributed to the development of these features of preeclampsia. Mechanistically, we provided both human and mouse evidence that elevated placental CD73 is a key enzyme causing increased placental adenosine, thereby contributing to preeclampsia. We determined that elevated placental adenosine signaling is a previously unrecognized pathogenic factor for preeclampsia. Moreover, our findings revealed the molecular basis underlying the elevation of placental adenosine and the detrimental role of excess placental adenosine in the pathophysiology of preeclampsia, and thereby, we highlight novel therapeutic targets. © 2014 American Heart Association, Inc.

  5. Adenosine receptor modulation: potential implications in veterinary medicine.

    PubMed

    Dip, Ramiro G

    2009-01-01

    Adenosine is a purine nucleoside whose concentration increases during inflammation and hypoxia and the many roles of this molecule are becoming better understood. Increased reactivity to adenosine of the airways of asthmatic but not of normal subjects underlines the role of adenosine in airway inflammation. The identification and pharmacological characterisation of different adenosine receptors have stimulated the search for subtype-specific ligands able to modulate the effects of this molecule in a directed way. Several compounds of different chemical classes have been identified as having potential drawbacks, including side effects resulting from the broad distribution of the receptors across the organism, have prevented clinical application. In this article, the effects of adenosine's different receptors and the intracellular signalling pathways are reviewed. The potential of adenosine receptor modulation as a therapeutic target for chronic airway inflammation is considered, taking equine recurrent airway disease and feline asthma as examples of naturally occurring airway obstructive diseases. Other potential applications for adenosine receptor modulation are also discussed. As the intrinsic molecular events of adenosine's mechanism of action become uncovered, new concrete therapeutic approaches will become available for the treatment of various conditions in veterinary medicine.

  6. Silk fibroin encapsulated powder reservoirs for sustained release of adenosine.

    PubMed

    Pritchard, Eleanor M; Szybala, Cory; Boison, Detlev; Kaplan, David L

    2010-06-01

    Due to its unique properties, silk fibroin was studied as a biodegradable polymer vehicle for sustained, local delivery of the anticonvulsant adenosine from encapsulated reservoirs. Silk is a biologically derived protein polymer that is biocompatible, mechanically strong and degrades to non-toxic products in vivo. To achieve local, sustained, controlled adenosine release from fully degradable implants, solid adenosine powder reservoirs were coated with silk fibroin. Material properties of the silk coating including thickness, crystallinity and morphology were investigated to assess the relationships between silk coating biomaterial features and adenosine release from silk encapsulated reservoirs. Reservoir coating thickness was varied through manipulation of the silk coating solution concentration and number of coatings applied. Release studies were also performed in proteinase type XIV to model the effects of degradation. Increasing the barrier to diffusion, either by increasing coating thickness or crystallinity was found to delay adenosine burst, decrease average daily release rate, and increase duration of release. In the case of encapsulated reservoirs coated with eight layers of 8% (w/v) silk, a linear release profile was observed and adenosine release was sustained for 14days. The ability to achieve nearly constant release for 2weeks for adenosine via control of the silk coating suggests these encapsulated reservoirs represent a novel system for delivering adenosine. We anticipate that this approach could also be extended to other implant needs and small-molecule drugs to treat a range of clinical needs.

  7. Adenosine strongly potentiates pressor responses to nicotine in rats.

    PubMed Central

    von Borstel, R W; Renshaw, A A; Wurtman, R J

    1984-01-01

    Intravenous infusion of subhypotensive doses of adenosine strongly potentiates the pressor response of anesthetized rats to nicotine. A dose of nicotine (40 micrograms/kg, i.v.), which, given alone, elicits a peak increase in diastolic pressure of approximately equal to 15 mm Hg, increases pressure by approximately equal to 70 mm Hg when arterial plasma adenosine levels have been increased to 2 microM from a basal concentration of approximately equal to 1 microM. The pressor response to cigarette smoke applied to the lungs is also strongly potentiated during infusion of adenosine. Slightly higher adenosine concentrations (approximately equal to 4 microM) attenuate pressor responses to electrical stimulation of preganglionic sympathetic nerves, or to injections of the alpha-adrenergic agonist phenylephrine, but continue to potentiate pressor responses to nicotine. Low doses (0.25-5 micrograms/kg) of the synthetic adenosine receptor agonists 5'-N-cyclopropylcarboxamidoadenosine, 2-chloroadenosine, and N6-L-phenylisopropyladenosine also potentiate pressor responses to nicotine. Caffeine and theophylline (10 mg/kg) block the potentiating effect of adenosine, and also decrease basal responses to nicotine, suggesting that endogenous adenosine might normally potentiate some nicotine responses. The synergism between nicotine and adenosine appears to take place within sympathetic ganglia. PMID:6591207

  8. Effect of theophylline on adenosine production in the canine myocardium

    SciTech Connect

    McKenzie, J.E.; Steffen, R.P.; Haddy, F.J.

    1987-01-01

    Adenosine is thought to participate in local regulation of coronary blood flow. However, competitive antagonists of adenosine fail to block myocardial active hyperemia. The authors examined the effect of locally administered theophylline on active hyperemia and myocardial adenosine production during intracoronary isoproterenol infusion in the dog heart. Isoproterenol decreased coronary resistance and increased myocardial adenosine production. Infusion of theophylline at a rate that attenuated the vasodilator response to exogenously administered adenosine failed to attenuate the increase in coronary blood flow produced by isoproterenol. However, theophylline plus isoproterenol production greater increases in myocardial adensine production than isoproterenol alone. The curves relating resistance and adenosine in the presence of theophylline fell to the right of those in the absence of theophylline. These findings suggest that the failure of theophylline to attenuate isoproterenol hyperemia in the dog heart results at least in part from an increase in adenosine concentration at the arteriole to a level beyond that blocked by this competitive antagonist and that adenosine may in fact play a role in isoproterenol-induced active hyperemia.

  9. Norepinephrines effect on adenosine transport in the proximal straight tubule

    SciTech Connect

    Barfuss, D.W.; McCann, W.P.; Katholi, R.E.

    1986-03-01

    The effect of norepinephrine on C/sup 14/-adenosine transport in the rabbit proximal tubule (S/sub 2/) was studied. The transepithelial transport of adenosine (0.02 mM0 from lumin to bathing solution was measured by its rate of appearance (J/sub A/) in the bathing solution and by its disappearances (J/sub D/) from the luminal fluid. Norepinephrine (0.24 ..mu..M) was added to the bathing solution after a control flux period. After three samples from the experiment period the tubules were quickly harvested and the cellular concentration of C/sup 14/-adenosine was determined. The high cellular adenosine concentration and th marked difference in adenosine appearance rate in the bathing solution compared to the luminal disappearance rate indicates the absorbed adenosine is trapped in the cells. This trapping may be due to adenosine metabolism or difficulty of crossing the basolateral membrane. Whichever is the case, norepinephrine appears to stimulate movement of adenosine or its metabolites into the bathing solution across the basolateral membrane.

  10. Adenosine signaling promotes hematopoietic stem and progenitor cell emergence.

    PubMed

    Jing, Lili; Tamplin, Owen J; Chen, Michael J; Deng, Qing; Patterson, Shenia; Kim, Peter G; Durand, Ellen M; McNeil, Ashley; Green, Julie M; Matsuura, Shinobu; Ablain, Julien; Brandt, Margot K; Schlaeger, Thorsten M; Huttenlocher, Anna; Daley, George Q; Ravid, Katya; Zon, Leonard I

    2015-05-04

    Hematopoietic stem cells (HSCs) emerge from aortic endothelium via the endothelial-to-hematopoietic transition (EHT). The molecular mechanisms that initiate and regulate EHT remain poorly understood. Here, we show that adenosine signaling regulates hematopoietic stem and progenitor cell (HSPC) development in zebrafish embryos. The adenosine receptor A2b is expressed in the vascular endothelium before HSPC emergence. Elevated adenosine levels increased runx1(+)/cmyb(+) HSPCs in the dorsal aorta, whereas blocking the adenosine pathway decreased HSPCs. Knockdown of A2b adenosine receptor disrupted scl(+) hemogenic vascular endothelium and the subsequent EHT process. A2b adenosine receptor activation induced CXCL8 via cAMP-protein kinase A (PKA) and mediated hematopoiesis. We further show that adenosine increased multipotent progenitors in a mouse embryonic stem cell colony-forming assay and in embryonic day 10.5 aorta-gonad-mesonephros explants. Our results demonstrate that adenosine signaling plays an evolutionary conserved role in the first steps of HSPC formation in vertebrates.

  11. Comorbidities in Neurology: Is adenosine the common link?

    PubMed

    Boison, Detlev; Aronica, Eleonora

    2015-10-01

    Comorbidities in Neurology represent a major conceptual and therapeutic challenge. For example, temporal lobe epilepsy (TLE) is a syndrome comprised of epileptic seizures and comorbid symptoms including memory and psychiatric impairment, depression, and sleep dysfunction. Similarly, Alzheimer's disease (AD), Parkinson's disease (PD), and Amyotrophic Lateral Sclerosis (ALS) are accompanied by various degrees of memory dysfunction. Patients with AD have an increased likelihood for seizures, whereas all four conditions share certain aspects of psychosis, depression, and sleep dysfunction. This remarkable overlap suggests common pathophysiological mechanisms, which include synaptic dysfunction and synaptotoxicity, as well as glial activation and astrogliosis. Astrogliosis is linked to synapse function via the tripartite synapse, but astrocytes also control the availability of gliotransmitters and adenosine. Here we will specifically focus on the 'adenosine hypothesis of comorbidities' implying that astrocyte activation, via overexpression of adenosine kinase (ADK), induces a deficiency in the homeostatic tone of adenosine. We present evidence from patient-derived samples showing astrogliosis and overexpression of ADK as common pathological hallmark of epilepsy, AD, PD, and ALS. We discuss a transgenic 'comorbidity model', in which brain-wide overexpression of ADK and resulting adenosine deficiency produces a comorbid spectrum of seizures, altered dopaminergic function, attentional impairment, and deficits in cognitive domains and sleep regulation. We conclude that dysfunction of adenosine signaling is common in neurological conditions, that adenosine dysfunction can explain co-morbid phenotypes, and that therapeutic adenosine augmentation might be effective for the treatment of comorbid symptoms in multiple neurological conditions.

  12. Adenosine receptors and dyskinesia in pathophysiology.

    PubMed

    Tomiyama, Masahiko

    2014-01-01

    First, the recent progress in the pathogenesis of levodopa-induced dyskinesia was described. Serotonin neurons play an important role in conversion from levodopa to dopamine and in the release of converted dopamine into the striatum in the Parkinsonian state. Since serotonin neurons lack buffering effects on synaptic dopamine concentration, the synaptic dopamine markedly fluctuates depending on the fluctuating levodopa concentration in the serum after taking levodopa. The resultant pulsatile stimulation makes the striatal direct-pathway neurons get potential that releases excessive GABA into the output nuclei of the basal ganglia. When levodopa is administered, the stored GABA is released, the output nuclei become hypoactive, and then dyskinesias emerge. Second, effects of adenosine A2A receptor antagonists on dyskinesia were described. It has been demonstrated that the expression of adenosine A2A receptors is increased in Parkinson's disease (PD) patients with dyskinesias, suggesting that blockade of A2A receptors is beneficial for dyskinesias. Preclinical studies have shown that A2A receptor antagonists reduce liability of dyskinesias in PD models. Clinical trials have demonstrated that A2A antagonists increase functional ON-time (ON without troublesome dyskinesia) in PD patients suffering from wearing-off phenomenon, although they may increase dyskinesia in patients with advanced PD.

  13. Regulation of muscarinic acetylcholine receptor-mediated synaptic responses by adenosine receptors in the rat hippocampus.

    PubMed Central

    Morton, R A; Davies, C H

    1997-01-01

    1. Intracellular current clamp recordings were made from CA1 pyramidal neurones in rat hippocampal slices. Experiments were performed in the presence of ionotropic glutamate receptor antagonists and gamma-aminobutyric acid (GABA) receptor antagonists to block all fast excitatory and inhibitory synaptic transmission. A single stimulus, delivered extracellularly in the stratum oriens, caused a reduction in spike frequency adaptation in response to a depolarizing current step delivered 2 s after the stimulus. A 2- to 10-fold increase in stimulus intensity evoked a slow excitatory postsynaptic potential (EPSP) which was associated with a small increase in input resistance. The peak amplitude of the EPSP occurred approximately 2.5 s after the stimulus and its magnitude (up to 30 mV) and duration (10-50 s) increased with increasing stimulus intensity. 2. The slow EPSP was unaffected by the metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG; 1000 microM) but was greatly enhanced by the acetylcholinesterase inhibitor physostigmine (1-5 microM). Both the slow EPSP and the stimulus-evoked reduction in spike frequency adaptation were inhibited by the muscarinic acetylcholine receptor (mAChR) antagonist atropine (1-5 microM). These results are consistent with these effects being mediated by mAChRs. 3. Both the mAChR-mediated EPSP (EPSPm) and the associated reduction in spike frequency adaptation were reversibly depressed (up to 97%) by either adenosine (100 microM) or its non-hydrolysable analogue 2-chloroadenosine (CADO; 0.1-5.0 microM). These effects were often accompanied by postsynaptic hyperpolarization (up to 8 mV) and a reduction in input resistance (up to 11%). The selective adenosine A1 receptor agonists 2-chloro-N6-cyclopentyladenosine (CCPA; 0.1-0.4 microM) and R(-)N6-(2-phenylisopropyl)-adenosine (R-PIA; 1 microM) both depressed the EPSPm. In contrast, the adenosine A2A receptor agonist 2-p-(2-carboxyethyl)-phenethylamino-5

  14. Extracellular Adenosine: A Safety Signal That Dampens Hypoxia-Induced Inflammation During Ischemia

    PubMed Central

    Grenz, Almut; Homann, Dirk

    2011-01-01

    Abstract Traditionally, the single most unique feature of the immune system has been attributed to its capability to discriminate between self (e.g., host proteins) and nonself (e.g., pathogens). More recently, an emerging immunologic concept involves the notion that the immune system responds via a complex system for sensing signals of danger, such as pathogens or host-derived signals of cellular distress (e.g., ischemia), while remaining unresponsive to nondangerous motifs. Experimental studies have provided strong evidence that the production and signaling effects of extracellular adenosine are dramatically enhanced during conditions of limited oxygen availability as occurs during ischemia. As such, adenosine would fit the bill of signaling molecules that are enhanced during situations of cellular distress. In contrast to a danger signal, we propose here that extracellular adenosine operates as a countermeasure, in fact as a safety signal, to both restrain potentially harmful immune responses and to maintain and promote general tissue integrity during conditions of limited oxygen availability. Antioxid. Redox Signal. 15, 2221–2234. PMID:21126189

  15. Conformational basis for the activation of adenylate cyclase by adenosine

    PubMed Central

    Miles, D. L.; Miles, D. W.; Eyring, H.

    1977-01-01

    The ability of adenosine to stimulate adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] and increase adenosine 3′:5′-cyclic monophosphate (cAMP) levels has important biochemical consequences. These include the suppression of immune responses and cardiovascular effects. Recent investigations involving the ability of adenosine and adenosine analogs to stimulate adenylate cyclase provided experimental data that appear to be correlated with the ability of adenosine and analogs of adenosine to exist in the glycosidic high anti conformation. 9-β-D-Arabinofuranosyladenine, which is not stable in the high anti conformation, is inactive as a stimulator of adenylate cyclase. 2′-Deoxyadenosine is also not stable in the high anti conformation but its instability may be significantly decreased by intramolecular adjustments promoted by receptor or active site interactions. 2′-Deoxyadenosine does not activate adenylate cyclase in lymphocytes when ATP is the substrate but is able to activate adenylate cyclase when 2-fluoro ATP is the substrate. The inability of certain analogs of adenosine, with bulky groups substituted for hydrogen at the 8 position of the adenine base, to activate adenylate cyclase and increase either lymphocyte or cardiac cell cAMP levels is consistent with the designation of the high anti conformation as being the conformation required for the activation of adenylate cyclase. An understanding of the glycosidic conformation required by the extracellular adenosine receptor of the adenosine molecule provides the basis for designing nucleoside analogs of adenosine that will exert a desired effect on cAMP levels. The avoidance of unwanted immunosuppressive or cardiotoxic effects can be arranged by structural changes that prohibit the high anti conformation. PMID:267918

  16. A High-Affinity Adenosine Kinase from Anopheles Gambiae

    SciTech Connect

    M Cassera; M Ho; E Merino; E Burgos; A Rinaldo-Matthis; S Almo; V Schramm

    2011-12-31

    Genome analysis revealed a mosquito orthologue of adenosine kinase in Anopheles gambiae (AgAK; the most important vector for the transmission of Plasmodium falciparum in Africa). P. falciparum are purine auxotrophs and do not express an adenosine kinase but rely on their hosts for purines. AgAK was kinetically characterized and found to have the highest affinity for adenosine (K{sub m} = 8.1 nM) of any known adenosine kinase. AgAK is specific for adenosine at the nucleoside site, but several nucleotide triphosphate phosphoryl donors are tolerated. The AgAK crystal structure with a bound bisubstrate analogue Ap{sub 4}A (2.0 {angstrom} resolution) reveals interactions for adenosine and ATP and the geometry for phosphoryl transfer. The polyphosphate charge is partly neutralized by a bound Mg{sup 2+} ion and an ion pair to a catalytic site Arg. The AgAK structure consists of a large catalytic core in a three-layer {alpha}/{beta}/{alpha} sandwich, and a small cap domain in contact with adenosine. The specificity and tight binding for adenosine arise from hydrogen bond interactions of Asn14, Leu16, Leu40, Leu133, Leu168, Phe168, and Thr171 and the backbone of Ile39 and Phe168 with the adenine ring as well as through hydrogen bond interactions between Asp18, Gly64, and Asn68 and the ribosyl 2'- and 3'-hydroxyl groups. The structure is more similar to that of human adenosine kinase (48% identical) than to that of AK from Toxoplasma gondii (31% identical). With this extraordinary affinity for AgAK, adenosine is efficiently captured and converted to AMP at near the diffusion limit, suggesting an important role for this enzyme in the maintenance of the adenine nucleotide pool. mRNA analysis verifies that AgAK transcripts are produced in the adult insects.

  17. Role of adenosine in tubuloglomerular feedback and acute renal failure.

    PubMed

    Osswald, H; Vallon, V; Mühlbauer, B

    1996-12-01

    1. Adenosine (ADO) can induce renal vasoconstriction and a fall in glomerular filtration rate. When the rate of ATP hydrolysis prevails over the rate of ATP synthesis the kidney generates ADO at an enhanced rate. 2. Tubuloglomerular feedback (TGF) is the vascular response to changes of the NaCl concentration in the tubular fluid at the macula densa segment, which is the result of transepithelial electrolyte reabsorption by the proximal tubule and the loop of Henle. 3. TGF can be inhibited by ADO-A1 receptor antagonists and is potentiated by substances that can elevate extracellular ADO concentrations. These observations led to the hypothesis that ADO is an element of the signal transmission processes in the juxtaglomerular apparatus. 4. Renal ischaemia and nephrotoxic substances can induce acute renal failure (ARF). ADO receptor antagonists have been shown to ameliorate renal function in several different models of ARF in laboratory animals and humans. 5. A number of factors, such as extracellular volume contraction, low NaCl diet, angiotensin II and cyclooxygenase inhibitors enhance to a similar extent: (a) the renal vascular response to endogenous and exogenous ADO; (b) the TGF response of the nephron; and (c) the severity of ARF. All three phenomena are susceptible to antagonism by ADO receptor antagonists. 6. Therefore, we conclude that ADO plays a significant role in normal and pathological states of kidney function.

  18. Adenosine A2B receptor blockade slows growth of bladder and breast tumors.

    PubMed

    Cekic, Caglar; Sag, Duygu; Li, Yuesheng; Theodorescu, Dan; Strieter, Robert M; Linden, Joel

    2012-01-01

    The accumulation of high levels of adenosine in tumors activates A(2A) and A(2B) receptors on immune cells and inhibits their ability to suppress tumor growth. Deletion of adenosine A(2A) receptors (A(2A)ARs) has been reported to activate antitumor T cells, stimulate dendritic cell (DC) function, and inhibit angiogenesis. In this study, we evaluated the effects of intermittent intratumor injection of a nonselective adenosine receptor antagonist, aminophylline (AMO; theophylline ethylenediamine) and, for the first time to our knowledge, a selective A(2B)AR antagonist, ATL801. AMO and ATL801 slowed the growth of MB49 bladder and 4T1 breast tumors in syngeneic mice and reduced by 85% metastasizes of breast cancer cells from mammary fat to lung. Based on experiments with A(2A)AR(-/-) or adenosine A(2B) receptor(-/-) mice, the effect of AMO injection was unexpectedly attributed to A(2B)AR and not to A(2A)AR blockade. AMO and ATL801 significantly increased tumor levels of IFN-γ and the IFN-inducible chemokine CXCL10, which is a ligand for CXCR3. This was associated with an increase in activated tumor-infiltrating CXCR3(+) T cells and a decrease in endothelial cell precursors within tumors. Tumor growth inhibition by AMO or ATL801 was eliminated in CXCR3(-/-) mice and RAG1(-/-) mice that lack mature T cells. In RAG1(-/-) mice, A(2B)AR deletion enhanced CD86 expression on CD11b(-) DCs. Bone marrow chimera experiments demonstrated that CXCR3 and A(2B)AR expression on bone marrow cells is required for the antitumor effects of AMO. The data suggest that blockade of A(2B)ARs enhances DC activation and CXCR3-dependent antitumor responses.

  19. Activation of neuronal adenosine A1 receptors suppresses secretory reflexes in the guinea pig colon.

    PubMed

    Cooke, H J; Wang, Y; Liu, C Y; Zhang, H; Christofi, F L

    1999-02-01

    The role of adenosine A1 receptors (A1R) in reflex-evoked short-circuit current (Isc) indicative of chloride secretion was studied in the guinea pig colon. The A1R antagonist 8-cyclopentyltheophylline (CPT) enhanced reflex-evoked Isc. Adenosine deaminase and the nucleoside transport inhibitor S-(4-nitrobenzyl)-6-thioinosine enhanced and reduced reflex-induced Isc, respectively. The A1R agonist 2-chloro-N6-cyclopentyladenosine (CCPA) inhibited reflex-evoked Isc at nanomolar concentrations, and its action was antagonized by CPT. In the presence of either N-acetyl-5-hydroxytryptophyl-5-hydroxytryptophan amide to block the 5-hydroxytryptamine (5-HT)-mediated pathway or piroxicam to block the prostaglandin-mediated pathway, CCPA reduced the residual reflex-evoked Isc. CCPA reduced the response to a 5-HT pulse without affecting the tetrodotoxin-insensitive Isc responses to carbachol or forskolin. Immunoreactivity for A1R was detected in the membrane (10% of neurons) and cytoplasm (90% of neurons) of neural protein gene product 9.5-immunoreactive (or S-100-negative) submucosal neurons, in glia, and in the muscularis mucosa. A1R immunoreactivity in a majority of neurons remained elevated in the cytoplasm despite preincubation with adenosine deaminase or CPT. A1R immunoreactivity colocalized in synaptophysin-immunoreactive presynaptic varicose nerve terminals. The results indicate that endogenous adenosine binding to high-affinity A1R on submucosal neurons acts as a physiological brake to suppress reflex-evoked Isc indicative of chloride secretion.

  20. Physical origins of remarkable thermostabilization by an octuple mutation for the adenosine A2a receptor

    NASA Astrophysics Data System (ADS)

    Kajiwara, Yuta; Ogino, Takahiro; Yasuda, Satoshi; Takamuku, Yuuki; Murata, Takeshi; Kinoshita, Masahiro

    2016-07-01

    It was experimentally showed that the thermal stability of a membrane protein, the adenosine A2a receptor, was remarkably enhanced by an octuple mutation. Here we theoretically prove that the energy decrease arising from the formation of protein intramolecular hydrogen bonds and the solvent-entropy gain upon protein folding are made substantially larger by the mutation, leading to the remarkable enhancement. The solvent is formed by hydrocarbon groups constituting nonpolar chains of the lipid bilayer within a membrane. The mutation modifies geometric characteristics of the structure so that the solvent crowding can be reduced to a larger extent when the protein folds.

  1. Adenosine receptors and diabetes: Focus on the A(2B) adenosine receptor subtype.

    PubMed

    Merighi, Stefania; Borea, Pier Andrea; Gessi, Stefania

    2015-09-01

    Over the last two decades, diabetes mellitus has become one of the most challenging health problems worldwide. Diabetes mellitus, classified as type I and II, is a pathology concerning blood glucose level in the body. The nucleoside adenosine has long been known to affect insulin secretion, glucose homeostasis and lipid metabolism, through activation of four G protein coupled adenosine receptors (ARs), named A1, A2A, A2B and A3. Currently, the novel promising subtype to develop new drugs for diabetes treatment is the A2BAR subtype. The use of selective agonists and antagonists for A2BAR subtype in various diabetic animal models allowed us to identify several effects of A2BAR signaling in cell metabolism. In particular, the focus of this review is to summarize the studies on purinergic signaling associated with diabetes through A2BARs modulation.

  2. Serum adenosine deaminase activity in cutaneous anthrax.

    PubMed

    Sunnetcioglu, Mahmut; Karadas, Sevdegul; Aslan, Mehmet; Ceylan, Mehmet Resat; Demir, Halit; Oncu, Mehmet Resit; Karahocagil, Mustafa Kasım; Sunnetcioglu, Aysel; Aypak, Cenk

    2014-07-06

    Adenosine deaminase (ADA) activity has been discovered in several inflammatory conditions; however, there are no data associated with cutaneous anthrax. The aim of this study was to investigate serum ADA activity in patients with cutaneous anthrax. Sixteen patients with cutaneous anthrax and 17 healthy controls were enrolled. We measured ADA activity; peripheral blood leukocyte, lymphocyte, neutrophil, and monocyte counts; erythrocyte sedimentation rate; and C reactive protein levels. Serum ADA activity was significantly higher in patients with cutaneous anthrax than in the controls (p<0.001). A positive correlation was observed between ADA activity and lymphocyte counts (r=0.589, p=0.021) in the patient group. This study suggests that serum ADA could be used as a biochemical marker in cutaneous anthrax.

  3. Characterization of P1 (adenosine) purinoceptors.

    PubMed

    Jarvis, Michael F

    2013-10-08

    The purine nucleoside adenosine (ADO) is an important modulator of cellular function in mammalian tissues, modulating cellular function and neuronal excitability via interactions with different cell surface receptor subtypes that are heterogeneously distributed in both the mammalian CNS and peripheral tissues. Four ADO receptor subtypes have been cloned and characterized. Described in this unit are three radioligand binding assays for pharmacological characterization of the high-affinity ADO receptor subtypes A1, A2A, and A3 receptors. Pharmacological characterization of the low-affinity A2B receptor has been enabled by the use of tritiated xanthine PSB-603. Because receptor localization is an important criterion for differentiation of receptor subtypes, a support protocol that describes the methodology for the localization of ADO receptors in rat brain tissue using autoradiography is also included.

  4. Extracellular cAMP-adenosine pathways in the mouse kidney

    PubMed Central

    Ren, Jin; Cheng, Dongmei; Mi, Zaichuan

    2011-01-01

    The renal extracellular 2′,3′-cAMP-adenosine and 3′,5′-cAMP-adenosine pathways (extracellular cAMPs→AMPs→adenosine) may contribute to renal adenosine production. Because mouse kidneys provide opportunities to investigate renal adenosine production in genetically modified kidneys, it is important to determine whether mouse kidneys express these cAMP-adenosine pathways. We administered (renal artery) 2′,3′-cAMP and 3′,5′-cAMP to isolated, perfused mouse kidneys and measured renal venous secretion rates of 2′,3′-cAMP, 3′,5′-cAMP, 2′-AMP, 3′-AMP, 5′-AMP, adenosine, and inosine. Arterial infusions of 2′,3′-cAMP increased (P < 0.0001) the mean venous secretion of 2′-AMP (390-fold), 3′-AMP (497-fold), adenosine (18-fold), and inosine (adenosine metabolite; 7-fold), but they did not alter 5′-AMP secretion. Infusions of 3′,5′-cAMP did not affect venous secretion of 2′-AMP or 3′-AMP, but they increased (P < 0.0001) secretion of 5′-AMP (5-fold), adenosine (17-fold), and inosine (6-fold). Energy depletion (metabolic inhibitors) increased the secretion of 2′,3′-cAMP (8-fold, P = 0.0081), 2′-AMP (4-fold, P = 0.0028), 3′-AMP (4-fold, P = 0.0270), 5′-AMP (3-fold, P = 0.0662), adenosine (2-fold, P = 0.0317), and inosine (7-fold, P = 0.0071), but it did not increase 3′,5′-cAMP secretion. The 2′,3′-cAMP-adenosine pathway was quantitatively similar in CD73 −/− vs. +/+ kidneys. However, 3′,5′-cAMP induced a 6.7-fold greater increase in 5′-AMP, an attenuated increase (61% reduction) in inosine and a similar increase in adenosine in CD73 −/− vs. CD73 +/+ kidneys. In mouse kidneys, 1) 2′,3′-cAMP and 3′,5′-cAMP are metabolized to their corresponding AMPs, which are subsequently metabolized to adenosine; 2) energy depletion activates the 2′,3′-cAMP-adenosine, but not the 3′,5′-cAMP-adenosine, pathway; and 3) although CD73 is involved in the 3′,5′-AMP-adenosine pathway, alternative

  5. Ethanol Tolerance Affects Endogenous Adenosine Signaling in Mouse Hippocampus

    PubMed Central

    Zhang, Dali; Xiong, Wei; Jackson, Michael F.

    2016-01-01

    Ethanol has many pharmacological effects, including increases in endogenous adenosine levels and adenosine receptor activity in brain. Ethanol consumption is associated with both positive and negative health outcomes, but tolerance to the behavioral effects of ethanol can lead to increased consumption, which increases the risk of negative health outcomes. The present study was performed to test whether a 7-day treatment with ethanol is linked to reduced adenosine signaling and whether this is a consequence of reduced ecto-5′-nucleotidase activity. Wild-type (CD73+/+) and ecto-5′-nucleotidase-deficient (CD73−/−) mice were treated with ethanol (2 g/kg) or saline for 7 days. In CD73+/+ mice, repeated ethanol treatment reduced the hypothermic and ataxic effects of acute ethanol, indicating the development of tolerance to the acute effects of ethanol. In CD73+/+ mice, this 7-day ethanol treatment led to increased hippocampal synaptic activity and reduced adenosine A1 receptor activity under both basal and low Mg2+ conditions. These effects of ethanol tolerance were associated with an 18% decrease in activity of ecto-5′-nucleotidase activity in hippocampal cell membranes. In contrast, ethanol treatment was not associated with changes in synaptic activity or adenosine signaling in hippocampus from CD73−/− mice. These data indicate that ethanol treatment is associated with a reduction in adenosine signaling through adenosine A1 receptors in hippocampus, mediated, at least in part, via reduced ecto-5′-nucleotidase activity. PMID:27189965

  6. Ethanol Tolerance Affects Endogenous Adenosine Signaling in Mouse Hippocampus.

    PubMed

    Zhang, Dali; Xiong, Wei; Jackson, Michael F; Parkinson, Fiona E

    2016-07-01

    Ethanol has many pharmacological effects, including increases in endogenous adenosine levels and adenosine receptor activity in brain. Ethanol consumption is associated with both positive and negative health outcomes, but tolerance to the behavioral effects of ethanol can lead to increased consumption, which increases the risk of negative health outcomes. The present study was performed to test whether a 7-day treatment with ethanol is linked to reduced adenosine signaling and whether this is a consequence of reduced ecto-5'-nucleotidase activity. Wild-type (CD73(+/+)) and ecto-5'-nucleotidase-deficient (CD73(-/-)) mice were treated with ethanol (2 g/kg) or saline for 7 days. In CD73(+/+) mice, repeated ethanol treatment reduced the hypothermic and ataxic effects of acute ethanol, indicating the development of tolerance to the acute effects of ethanol. In CD73(+/+) mice, this 7-day ethanol treatment led to increased hippocampal synaptic activity and reduced adenosine A1 receptor activity under both basal and low Mg(2+) conditions. These effects of ethanol tolerance were associated with an 18% decrease in activity of ecto-5'-nucleotidase activity in hippocampal cell membranes. In contrast, ethanol treatment was not associated with changes in synaptic activity or adenosine signaling in hippocampus from CD73(-/-) mice. These data indicate that ethanol treatment is associated with a reduction in adenosine signaling through adenosine A1 receptors in hippocampus, mediated, at least in part, via reduced ecto-5'-nucleotidase activity. Copyright © 2016 The Author(s).

  7. Adenosine 2A receptors in acute kidney injury.

    PubMed

    Vincent, I S; Okusa, M D

    2015-07-01

    Acute kidney injury (AKI) is an important clinical problem that may lead to death and for those who survive, the sequelae of AKI include loss of quality of life, chronic kidney disease and end-stage renal disease. The incidence of AKI continues to rise without clear successes in humans for the pharmacological prevention of AKI or treatment of established AKI. Dendritic cells and macrophages are critical early initiators of innate immunity in the kidney and orchestrate inflammation subsequent to ischaemia-reperfusion injury. These innate cells are the most abundant leucocytes present in the kidney, and they represent a heterogeneous population of cells that are capable of responding to cues from the microenvironment derived from pathogens or endogenous inflammatory mediators such as cytokines or anti-inflammatory mediators such as adenosine. Lymphocyte subsets such as natural killer T cells and Tregs also play roles in regulating ischaemic injury by promoting and suppressing inflammation respectively. Adenosine, produced in response to IR, is generally considered as a protective signalling molecule and elicits its physiological responses through four distinct adenosine receptors. However, its short half-life, lack of specificity and rapid metabolism limit the use of adenosine as a therapeutic agent. These adenosine receptors play various roles in regulating the activity of the aforementioned hematopoietic cells in elevated levels of adenosine such as during hypoxia. This review focuses on the importance of one receptor, the adenosine 2A subtype, in blocking inflammation associated with AKI.

  8. Temporal variations of adenosine metabolism in human blood.

    PubMed

    Chagoya de Sánchez, V; Hernández-Muñoz, R; Suárez, J; Vidrio, S; Yáñez, L; Aguilar-Roblero, R; Oksenberg, A; Vega-González, A; Villalobos, L; Rosenthal, L; Fernández-Cancino, F; Drucker-Colín, R; Díaz-Muñoz, M

    1996-08-01

    Eight diurnally active (06:00-23:00 h) subjects were adapted for 2 days to the room conditions where the experiments were performed. Blood sampling for adenosine metabolites and metabolizing enzymes was done hourly during the activity span and every 30 min during sleep. The results showed that adenosine and its catabolites (inosine, hypoxanthine, and uric acid), adenosine synthesizing (S-adenosylhomocysteine hydrolase and 5'-nucleotidase), degrading (adenosine deaminase) and nucleotide-forming (adenosine kinase) enzymes as well as adenine nucleotides (AMP, ADP, and ATP) undergo statistically significant fluctuations (ANOVA) during the 24 h. However, energy charge was invariable. Glucose and lactate chronograms were determined as metabolic indicators. The same data analyzed by the chi-square periodogram and Fourier series indicated ultradian oscillatory periods for all the metabolites and enzymatic activities determined, and 24-h oscillatory components for inosine, hypoxanthine, adenine nucleotides, glucose, and the activities of SAH-hydrolase, 5'-nucleotidase, and adenosine kinase. The single cosinor method showed significant oscillatory components exclusively for lactate. As a whole, these results suggest that adenosine metabolism may play a role as a biological oscillator coordinating and/or modulating the energy homeostasis and physiological status of erythrocytes in vivo and could be an important factor in the distribution of purine rings for the rest of the organism.

  9. Non-additive modulation of synaptic transmission by serotonin, adenosine, and cholinergic modulators in the sensory thalamus.

    PubMed

    Yang, Ya-Chin; Hu, Chun-Chang; Lai, Yi-Chen

    2015-01-01

    The thalamus relays sensory information to the cortex. Oscillatory activities of the thalamocortical network are modulated by monoamines, acetylcholine, and adenosine, and could be the key features characteristic of different vigilance states. Although the thalamus is almost always subject to the actions of more than just one neuromodulators, reports on the modulatory effect of coexisting neuromodulators on thalamic synaptic transmission are unexpectedly scarce. We found that, if present alone, monoamine or adenosine decreases retinothalamic synaptic strength and short-term depression, whereas cholinergic modulators generally enhance postsynaptic response to presynaptic activity. However, coexistence of different modulators tends to produce non-additive effect, not predictable based on the action of individual modulators. Acetylcholine, acting via nicotinic receptors, can interact with either serotonin or adenosine to abolish most short-term synaptic depression. Moreover, the coexistence of adenosine and monoamine, with or without acetylcholine, results in robustly decreased synaptic strength and transforms short-term synaptic depression to facilitation. These findings are consistent with a view that acetylcholine is essential for an "enriched" sensory flow through the thalamus, and the flow is trimmed down by concomitant monoamine or adenosine (presumably for the wakefulness and rapid-eye movement, or REM, sleep states, respectively). In contrast, concomitant adenosine and monoamine would lead to a markedly "deprived" (and high-pass filtered) sensory flow, and thus the dramatic decrease of monoamine may constitute the basic demarcation between non-REM and REM sleep. The collective actions of different neuromodulators on thalamic synaptic transmission thus could be indispensable for the understanding of network responsiveness in different vigilance states.

  10. Adenosine triphosphate-sensitive potassium channel blockers attenuate the antiallodynic effect of R-PIA in neuropathic rats.

    PubMed

    Song, Jun-Gol; Hahm, Kyung Don; Kim, Young Ki; Leem, Jeong Gil; Lee, Chung; Jeong, Sung Moon; Park, Pyung Hwan; Shin, Jin Woo

    2011-06-01

    Nerve injury can generate neuropathic pain. The accompanying mechanical allodynia may be reduced by the intrathecal administration of adenosine. The neuroprotective effects of adenosine are mediated by the adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channel. We assessed the relationship between the adenosine A1 receptor agonist, N⁶-(R)-phenylisopropyl adenosine (R-PIA), and K(ATP) channels to determine whether the antiallodynic effects of R-PIA are also mediated through K(ATP) channels in a rat nerve ligation injury model of neuropathic pain. Mechanical allodynia was induced by tight ligation of the left lumbar fifth and sixth spinal nerves. Mechanical allodynia in the left hindpaw was evaluated using von Frey filaments to measure withdrawal thresholds. R-PIA (0.5, 1, or 2 μg) was administered intrathecally to induce antiallodynia. We assessed whether pretreatment with the K(ATP) channel blockers glibenclamide or 5-hydroxydecanoate reversed the antiallodynic effect of R-PIA. Also, we evaluated whether diazoxide, a K(ATP) channel opener, had an antiallodynic effect and promoted the antiallodynic effect of R-PIA. Lastly, we investigated whether the voltage-activated K channel blocker 4-aminopyridine attenuated the effect of R-PIA. Intrathecal R-PIA produced maximal antiallodynia at 2 μg (P < 0.05). Intrathecal pretreatment with glibenclamide and intraperitoneal pretreatment 5-hydroxydecanoate significantly reduced the antiallodynic effect of R-PIA. Diazoxide produced an antiallodynic effect and also enhanced the antiallodynic action of R-PIA. 4-Aminopyridine had no effect on the antiallodynic action of R-PIA. The antiallodynic effects of adenosine A1 receptor stimulation may be related to K(ATP) channel activity in a rat model of nerve ligation injury.

  11. Non-additive modulation of synaptic transmission by serotonin, adenosine, and cholinergic modulators in the sensory thalamus

    PubMed Central

    Yang, Ya-Chin; Hu, Chun-Chang; Lai, Yi-Chen

    2015-01-01

    The thalamus relays sensory information to the cortex. Oscillatory activities of the thalamocortical network are modulated by monoamines, acetylcholine, and adenosine, and could be the key features characteristic of different vigilance states. Although the thalamus is almost always subject to the actions of more than just one neuromodulators, reports on the modulatory effect of coexisting neuromodulators on thalamic synaptic transmission are unexpectedly scarce. We found that, if present alone, monoamine or adenosine decreases retinothalamic synaptic strength and short-term depression, whereas cholinergic modulators generally enhance postsynaptic response to presynaptic activity. However, coexistence of different modulators tends to produce non-additive effect, not predictable based on the action of individual modulators. Acetylcholine, acting via nicotinic receptors, can interact with either serotonin or adenosine to abolish most short-term synaptic depression. Moreover, the coexistence of adenosine and monoamine, with or without acetylcholine, results in robustly decreased synaptic strength and transforms short-term synaptic depression to facilitation. These findings are consistent with a view that acetylcholine is essential for an “enriched” sensory flow through the thalamus, and the flow is trimmed down by concomitant monoamine or adenosine (presumably for the wakefulness and rapid-eye movement, or REM, sleep states, respectively). In contrast, concomitant adenosine and monoamine would lead to a markedly “deprived” (and high-pass filtered) sensory flow, and thus the dramatic decrease of monoamine may constitute the basic demarcation between non-REM and REM sleep. The collective actions of different neuromodulators on thalamic synaptic transmission thus could be indispensable for the understanding of network responsiveness in different vigilance states. PMID:25852468

  12. IFN-γ prevents adenosine receptor (A2bR) upregulation to sustain the macrophage activation response

    PubMed Central

    Cohen, Heather B.; Ward, Amanda; Hamidzadeh, Kajal; Ravid, Katya; Mosser, David M.

    2015-01-01

    The priming of macrophages with IFN-γ prior to TLR stimulation results in enhanced and prolonged inflammatory cytokine production. Here, we demonstrate that following TLR stimulation, macrophages up regulate the adenosine 2b receptor (A2bR) to enhance their sensitivity to immunosuppressive extracellular adenosine. This up-regulation of A2bR leads to the induction of a macrophage with an immunoregulatory phenotype and the down regulation of inflammation. IFN-γ priming of macrophages, selectively prevents the induction of the A2bR in macrophages to mitigate sensitivity to adenosine and prevent this regulatory transition. IFN-γ-mediated A2bR blockade leads to a prolonged production of TNFα and IL-12 in response to TLR ligation. The pharmacological inhibition or the genetic deletion of the A2bR results in a hyper-inflammatory response to TLR ligation, similar to IFN-γ treatment of macrophages. Conversely, the overexpression of A2bR on macrophages blunts the IFN-γ effects and promotes the development of immunoregulatory macrophages. Thus, we propose a novel mechanism whereby IFN-γ contributes to host defense, by desensitizing macrophages to the immunoregulatory effects of adenosine. This mechanism overcomes the transient nature of TLR activation, and prolongs the anti-microbial state of the classically activated macrophage. This study may offer promising new targets to improve the clinical outcome of inflammatory diseases in which macrophage activation is dysregulated. PMID:26355158

  13. Increased Cortical Extracellular Adenosine Correlates with Seizure Termination

    PubMed Central

    Van Gompel, Jamie J.; Bower, Mark R.; Worrell, Gregory A.; Stead, Matt; Chang, Su-Youne; Goerss, Stephan J.; Kim, Inyong; Bennet, Kevin E.; Meyer, Fredric B.; Marsh, W. Richard; Blaha, Charles D.; Lee, Kendall H.

    2014-01-01

    Objective Seizures are currently defined by their electrographic features. However, neuronal networks are intrinsically dependent upon neurotransmitters of which little is known regarding their peri-ictal dynamics. Evidence supports adenosine as having a prominent role in seizure termination, as its administration can terminate and reduce seizures in animal models. Further, microdialysis studies in humans suggest adenosine is elevated peri-ictally, but the relationship to the seizure is obscured by its temporal measurement limitations. Because electrochemical techniques can provide vastly superior temporal resolution, we test the hypothesis that extracellular adenosine concentrations rise during seizure termination in an animal model and humans using electrochemistry. Methods White farm swine (n=45) were used in an acute cortical model of epilepsy and 10 human epilepsy patients were studied during intraoperative electrocorticography (Ecog). Wireless Instantaneous Neurotransmitter Concentration Sensor (WINCS) based fast scan cyclic voltametry (FSCV) and fixed potential amperometry were obtained utilizing an adenosine specific triangular waveform or biosensors respectively. Results Simultaneous Ecog and electrochemistry demonstrated an average adenosine rise of 260% compared to baseline at 7.5 ± 16.9 seconds with amperometry (n=75 events) and 2.6 ± 11.2 seconds with FSCV (n=15 events) prior to electrographic seizure termination. In agreement with these animal data, adenosine elevation prior to seizure termination in a human patient utilizing FSCV was also seen. Significance Simultaneous Ecog and electrochemical recording supports the hypothesis that adenosine rises prior to seizure termination, suggesting that adenosine itself may be responsible for seizure termination. Future work using intraoperative WINCS based FSCV recording may help to elucidate the precise relationship between adenosine and seizure termination. PMID:24483230

  14. Gene Duplication in Pseudomonas aeruginosa Improves Growth on Adenosine.

    PubMed

    Toussaint, Jean-Paul; Farrell-Sherman, Anna; Feldman, Tamar Perla; Smalley, Nicole E; Schaefer, Amy L; Greenberg, E Peter; Dandekar, Ajai A

    2017-11-01

    The laboratory strain of Pseudomonas aeruginosa, PAO1, activates genes for catabolism of adenosine using quorum sensing (QS). However, this strain is not well-adapted for growth on adenosine, with doubling times greater than 40 h. We previously showed that when PAO1 is grown on adenosine and casein, variants emerge that grow rapidly on adenosine. To understand the mechanism by which this adaptation occurs, we performed whole-genome sequencing of five isolates evolved for rapid growth on adenosine. All five genomes had a gene duplication-amplification (GDA) event covering several genes, including the quorum-regulated nucleoside hydrolase gene, nuh, and PA0148, encoding an adenine deaminase. In addition, two of the growth variants also exhibited a nuh promoter mutation. We recapitulated the rapid growth phenotype with a plasmid containing six genes common to all the GDA events. We also showed that nuh and PA0148, the two genes at either end of the common GDA, were sufficient to confer rapid growth on adenosine. Additionally, we demonstrated that the variant nuh promoter increased basal expression of nuh but maintained its QS regulation. Therefore, GDA in P. aeruginosa confers the ability to grow efficiently on adenosine while maintaining QS regulation of nucleoside catabolism.IMPORTANCEPseudomonas aeruginosa thrives in many habitats and is an opportunistic pathogen of humans. In these diverse environments, P. aeruginosa must adapt to use myriad potential carbon sources. P. aeruginosa PAO1 cannot grow efficiently on nucleosides, including adenosine; however, it can acquire this ability through genetic adaptation. We show that the mechanism of adaptation is by amplification of a specific region of the genome and that the amplification preserves the regulation of the adenosine catabolic pathway by quorum sensing. These results demonstrate an underexplored mechanism of adaptation by P. aeruginosa, with implications for phenotypes such as development of antibiotic

  15. Impact of time to therapy and reperfusion modality on the efficacy of adenosine in acute myocardial infarction: the AMISTAD-2 trial.

    PubMed

    Kloner, Robert A; Forman, Mervyn B; Gibbons, Raymond J; Ross, Allan M; Alexander, R Wayne; Stone, Gregg W

    2006-10-01

    The purpose of this analysis was to determine whether the efficacy of adenosine vs. placebo was dependent on the timing of reperfusion therapy in the second Acute Myocardial Infarction Study of Adenosine (AMISTAD-II). Patients presenting with ST-segment elevation anterior AMI were randomized to receive placebo vs. adenosine (50 or 70 microg/kg/min) for 3 h starting within 15 min of reperfusion therapy. In the present post hoc hypothesis generating study, the results were stratified according to the timing of reperfusion, i.e. > or = or < the median 3.17 h, and by reperfusion modality. In patients receiving reperfusion < 3.17 h, adenosine compared with placebo significantly reduced 1-month mortality (5.2 vs. 9.2%, respectively, P = 0.014), 6-month mortality (7.3 vs. 11.2%, P = 0.033), and the occurrence of the primary 6-month composite clinical endpoint of death, in-hospital CHF, or rehospitalization for CHF at 6 months (12.0 vs. 17.2%, P = 0.022). Patients reperfused beyond 3 h did not benefit from adenosine. In this post hoc analysis, 3 h adenosine infusion administered as an adjunct to reperfusion therapy within the first 3.17 h onset of evolving anterior ST-segment elevation AMI enhanced early and late survival, and reduced the composite clinical endpoint of death or CHF at 6 months.

  16. Adenosine and Preexcitation Variants: Reappraisal of Electrocardiographic Changes.

    PubMed

    Ali, Hussam; Lupo, Pierpaolo; Foresti, Sara; De Ambroggi, Guido; Epicoco, Gianluca; Fundaliotis, Angelica; Cappato, Riccardo

    2016-07-01

    Intravenous adenosine is a short-acting blocker of the atrioventricular node that has been used to unmask subtle or latent preexcitation, and also to enable catheter ablation in selected patients with absent or intermittent preexcitation. Depending on the accessory pathway characteristics, intravenous adenosine may produce specific electrocardiographic changes highly suggestive of the preexcitation variant. Herein, we view different ECG responses to this pharmacological test in various preexcitation patterns that were confirmed by electrophysiological studies. Careful analysis of electrocardiographic changes during adenosine test, with emphasis on P-delta interval, preexcitation degree, and atrioventricular block, can be helpful to diagnose the preexcitation variant/pattern.

  17. A3 adenosine receptor inhibition improves the efficacy of hypertonic saline resuscitation

    PubMed Central

    Inoue, Yoshiaki; Tanaka, Hiroshi; Sumi, Yuka; Woehrle, Tobias; Chen, Yu; Hirsh, Mark I.; Junger, Wolfgang G.

    2011-01-01

    We reported previously that hypertonic saline (HS) treatment can prevent or upregulate the function of polymorphonuclear neutrophils (PMN) via A2a adenosine receptors (A2aR) or A3 adenosine receptors (A3R), respectively. A3R translocate to the cell surface upon PMN stimulation and thus HS promotes PMN responses under conditions of delayed HS treatment. Here we investigated if inhibition of A3R improves the protective effects of HS resuscitation in a mouse sepsis model. We found that HS nearly triples extracellular adenosine concentrations in whole blood and that inhibition of A3R with the selective antagonist MRS-1191 dose-dependently improves the inhibitory effect of HS. MRS-1191 at a concentration of 1 nM enhanced the inhibitory effect of HS and reduced stimulatory effects of delayed HS treatment. Using a mouse model of cecal ligation and puncture (CLP)-induced sepsis, we found that MRS-1191 reduces acute lung injury and PMN accumulation in lung tissue. While delayed HS treatment (4 ml/kg of 7.5 % NaCl) of mice 1 h after CLP aggravated PMN accumulation, lung tissue damage, and mortality 24 h after CLP, infusion of MRS-1191 (2 ng/kg body weight) combined with HS reduced these detrimental effects of delayed HS treatment. Our data thus show that A3 receptor antagonists can strengthen the beneficial effects of HS resuscitation by avoiding stimulatory side effects that result from delayed HS administration. PMID:20661181

  18. Excess adenosine in murine penile erectile tissues contributes to priapism via A2B adenosine receptor signaling

    PubMed Central

    Mi, Tiejuan; Abbasi, Shahrzad; Zhang, Hong; Uray, Karen; Chunn, Janci L.; Xia, Ling Wei; Molina, Jose G.; Weisbrodt, Norman W.; Kellems, Rodney E.; Blackburn, Michael R.; Xia, Yang

    2008-01-01

    Priapism, abnormally prolonged penile erection in the absence of sexual excitation, is associated with ischemia-mediated erectile tissue damage and subsequent erectile dysfunction. It is common among males with sickle cell disease (SCD), and SCD transgenic mice are an accepted model of the disorder. Current strategies to manage priapism suffer from a poor fundamental understanding of the molecular mechanisms underlying the disorder. Here we report that mice lacking adenosine deaminase (ADA), an enzyme necessary for the breakdown of adenosine, displayed unexpected priapic activity. ADA enzyme therapy successfully corrected the priapic activity both in vivo and in vitro, suggesting that it was dependent on elevated adenosine levels. Further genetic and pharmacologic evidence demonstrated that A2B adenosine receptor–mediated (A2BR-mediated) cAMP and cGMP induction was required for elevated adenosine–induced prolonged penile erection. Finally, priapic activity in SCD transgenic mice was also caused by elevated adenosine levels and A2BR activation. Thus, we have shown that excessive adenosine accumulation in the penis contributes to priapism through increased A2BR signaling in both Ada–/– and SCD transgenic mice. These findings provide insight regarding the molecular basis of priapism and suggest that strategies to either reduce adenosine or block A2BR activation may prove beneficial in the treatment of this disorder. PMID:18340377

  19. Adenosine through the A2A adenosine receptor increases IL-1β in the brain contributing to anxiety

    PubMed Central

    Chiu, Gabriel S.; Darmody, Patrick T.; Walsh, John P.; Moon, Morgan L.; Kwakwa, Kristin A.; Bray, Julie K.; McCusker, Robert H.; Freund, Gregory G.

    2014-01-01

    Anxiety is one of the most commonly reported psychiatric conditions, but its pathogenesis is poorly understood. Ailments associated with activation of the innate immune system, however, are increasingly linked to anxiety disorders. In adult male mice, we found that adenosine doubled caspase-1 activity in brain by a pathway reliant on ATP-sensitive potassium (KATP) channels, protein kinase A (PKA) and the A2A adenosine receptor (AR). In addition, adenosine-dependent activation of caspase-1 increased interleukin (IL)-1β in the brain by two-fold. Peripheral administration of adenosine in wild-type (WT) mice led to a 2.3-fold increase in caspase-1 activity in the amygdala and to a 33% and 42% reduction in spontaneous locomotor activity and food intake, respectively, that were not observed in caspase-1 knockout (KO), IL-1 receptor type 1 (IL-1R1) KO and A2A AR KO mice or in mice administered a caspase-1 inhibitor centrally. Finally, adenosine administration increased anxiety-like behaviors in WT mice by 28% in the open field test and by 55% in the elevated zero-maze. Caspase-1 KO mice, IL-1R1 KO mice, A2A AR KO mice and WT mice treated with the KATP channel blocker, glyburide, were resistant to adenosine-induced anxiety-like behaviors. Thus, our results indicate that adenosine can act as an anxiogenic by activating caspase-1 and increasing IL-1β in the brain. PMID:24907587

  20. High-dose parenteral ascorbate enhanced chemosensitivity of ovarian cancer and reduced toxicity of chemotherapy.

    PubMed

    Ma, Yan; Chapman, Julia; Levine, Mark; Polireddy, Kishore; Drisko, Jeanne; Chen, Qi

    2014-02-05

    Ascorbate (vitamin C) was an early, unorthodox therapy for cancer, with an outstanding safety profile and anecdotal clinical benefit. Because oral ascorbate was ineffective in two cancer clinical trials, ascorbate was abandoned by conventional oncology but continued to be used in complementary and alternative medicine. Recent studies provide rationale for reexamining ascorbate treatment. Because of marked pharmacokinetic differences, intravenous, but not oral, ascorbate produces millimolar concentrations both in blood and in tissues, killing cancer cells without harming normal tissues. In the interstitial fluid surrounding tumor cells, millimolar concentrations of ascorbate exert local pro-oxidant effects by mediating hydrogen peroxide (H(2)O(2)) formation, which kills cancer cells. We investigated downstream mechanisms of ascorbate-induced cell death. Data show that millimolar ascorbate, acting as a pro-oxidant, induced DNA damage and depleted cellular adenosine triphosphate (ATP), activated the ataxia telangiectasia mutated (ATM)/adenosine monophosphate-activated protein kinase (AMPK) pathway, and resulted in mammalian target of rapamycin (mTOR) inhibition and death in ovarian cancer cells. The combination of parenteral ascorbate with the conventional chemotherapeutic agents carboplatin and paclitaxel synergistically inhibited ovarian cancer in mouse models and reduced chemotherapy-associated toxicity in patients with ovarian cancer. On the basis of its potential benefit and minimal toxicity, examination of intravenous ascorbate in combination with standard chemotherapy is justified in larger clinical trials.

  1. Recent advances in the neurobiology of alcoholism: the role of adenosine.

    PubMed

    Mailliard, William S; Diamond, Ivan

    2004-01-01

    Neuronal responses to alcohol involve several hormone- and neurotransmitter-activated signal transduction pathways. Recent studies suggest that the adenosine A2 receptor (A2) mediates important actions of alcohol. Ethanol inhibits adenosine reuptake, increases extracellular adenosine, and promotes activation of A2. This leads to enhanced cAMP/protein kinase A (PKA) signaling ranging from increases in cAMP to stimulation of cAMP-dependent cAMP response element (CRE)-mediated gene expression. Medium spiny neurons in the striatum/nucleus accumbens (NAc) express A2 and dopamine D2 receptor (D2) on the same cells. Studies in model neuronal cell lines and primary neurons in culture expressing A2 and D2 provide evidence for synergy between ethanol/A2 and D2. Subthreshold concentrations of ethanol or a D2 agonist, without effect separately, synergistically activate cAMP/PKA signaling. Thus, neurons expressing A2 and D2 on the same cells, like in the NAc, are characterized by hypersensitivity to ethanol with a simultaneous activation of dopaminergic signaling. Synergy requires adenosine and appears to be mediated by the release of free betagamma dimers from G(i/o) via D2 activation. The release of free betagamma has pathophysiological significance in the drinking animal because specific blockade of betagamma signaling in the NAc strikingly reduces voluntary alcohol consumption. These findings suggest that signaling pathways, which regulate synergy between A2 and D2, might contain molecular targets for the prevention and treatment of alcoholism and alcohol abuse.

  2. Effects of nitroglycerine on coronary flow velocity before and during adenosine provocation.

    PubMed

    Wittfeldt, Ann; Jeppsson, Anders; Gan, Li-Ming

    2016-12-06

    Transthoracic echocardiography-assessed coronary flow velocity reserve (CFVR) evaluates coronary microvascular arterial function. Coronary flow velocity measurements at baseline and during hyperemia are used to calculate CFVR. Adenosine infusion induces hyperemia but it is not known if it causes a maximal response. We hypothesized that pre-treatment with nitroglycerine before adenosine provocation enhances hyperemia. Twenty-three healthy study subjects (mean age 27.5 ± 5.5, 35% women) underwent CFVR measurements before and after pretreatment with sublingual nitroglycerine (0.5 mg). Hyperemia was induced by adenosine infusion (140 μg/kg/min). In addition, the effect of nitroglycerin on left main coronary artery diameter was assessed. Pretreatment with nitroglycerine increased median CFVR from 3.6 (range 2.8-4.3) to 5.0 (4.1-6.0), p = 0.002. The increase was caused by a marked reduction in baseline coronary flow velocity 17 (15-24) vs 27 (19-31) cm/s, p < 0.0001) while hyperemic velocity remained unchanged (90 (68-116) vs 93 (75-105) cm/s, p = 0.48). Nitroglycerin significantly dilated the left main coronary artery (from median 3.1 (2.7-3.6) mm to 3.8 (3.1-4.3) mm, p = 0.018). Pretreatment with nitroglycerine dilates coronary arteries and increases coronary flow velocity reserve indicating that adenosine alone causes a submaximal hyperemia.

  3. Adenosine receptor ligands: differences with acute versus chronic treatment

    PubMed Central

    Jacobson, Kenneth A.; von Lubitz, Dag K. J. E.; Daly, John W.; Fredholm, Bertil B.

    2012-01-01

    Adenosine receptors have been the target of intense research with respect to potential use of selective ligands in a variety of therapeutic areas. Caffeine and theophylline are adenosine receptor antagonists, and over the past three decades a wide range of selective agonists and antagonists for adenosine receptor subtypes have been developed. A complication to the therapeutic use of adenosine receptor ligands is the observation that the effects of acute administration of a particular ligand can be diametrically opposite to the chronic effects of the same ligand. This ‘effect inversion’ is discussed here by Ken Jecobson and colleagues, and has been observed for effects on cognitive processes, seizures and ischaemic damage. PMID:8936347

  4. Extracellular Adenosine Mediates a Systemic Metabolic Switch during Immune Response

    PubMed Central

    Bajgar, Adam; Kucerova, Katerina; Jonatova, Lucie; Tomcala, Ales; Schneedorferova, Ivana; Okrouhlik, Jan; Dolezal, Tomas

    2015-01-01

    Immune defense is energetically costly, and thus an effective response requires metabolic adaptation of the organism to reallocate energy from storage, growth, and development towards the immune system. We employ the natural infection of Drosophila with a parasitoid wasp to study energy regulation during immune response. To combat the invasion, the host must produce specialized immune cells (lamellocytes) that destroy the parasitoid egg. We show that a significant portion of nutrients are allocated to differentiating lamellocytes when they would otherwise be used for development. This systemic metabolic switch is mediated by extracellular adenosine released from immune cells. The switch is crucial for an effective immune response. Preventing adenosine transport from immune cells or blocking adenosine receptor precludes the metabolic switch and the deceleration of development, dramatically reducing host resistance. Adenosine thus serves as a signal that the “selfish” immune cells send during infection to secure more energy at the expense of other tissues. PMID:25915062

  5. Adenosine promotes vascular barrier function in hyperoxic lung injury

    PubMed Central

    Davies, Jonathan; Karmouty‐Quintana, Harry; Le, Thuy T.; Chen, Ning‐Yuan; Weng, Tingting; Luo, Fayong; Molina, Jose; Moorthy, Bhagavatula; Blackburn, Michael R.

    2014-01-01

    Abstract Hyperoxic lung injury is characterized by cellular damage from high oxygen concentrations that lead to an inflammatory response in the lung with cellular infiltration and pulmonary edema. Adenosine is a signaling molecule that is generated extracellularly by CD73 in response to injury. Extracellular adenosine signals through cell surface receptors and has been found to be elevated and plays a protective role in acute injury situations. In particular, ADORA2B activation is protective in acute lung injury. However, little is known about the role of adenosine signaling in hyperoxic lung injury. We hypothesized that hyperoxia‐induced lung injury leads to CD73‐mediated increases in extracellular adenosine, which is protective through ADORA2B signaling pathways. To test this hypothesis, we exposed C57BL6, CD73−/−, and Adora2B−/− mice to 95% oxygen or room air and examined markers of pulmonary inflammation, edema, and monitored lung histology. Hyperoxic exposure caused pulmonary inflammation and edema in association with elevations in lung adenosine levels. Loss of CD73‐mediated extracellular adenosine production exacerbated pulmonary edema without affecting inflammatory cell counts. Furthermore, loss of the ADORA2B had similar results with worsening of pulmonary edema following hyperoxia exposure without affecting inflammatory cell infiltration. This loss of barrier function correlated with a decrease in occludin in pulmonary vasculature in CD73−/− and Adora2B−/− mice following hyperoxia exposure. These results demonstrate that exposure to a hyperoxic environment causes lung injury associated with an increase in adenosine concentration, and elevated adenosine levels protect vascular barrier function in hyperoxic lung injury through the ADORA2B‐dependent regulation of occludin. PMID:25263205

  6. The A3 adenosine receptor: history and perspectives.

    PubMed

    Borea, Pier Andrea; Varani, Katia; Vincenzi, Fabrizio; Baraldi, Pier Giovanni; Tabrizi, Mojgan Aghazadeh; Merighi, Stefania; Gessi, Stefania

    2015-01-01

    By general consensus, the omnipresent purine nucleoside adenosine is considered a major regulator of local tissue function, especially when energy supply fails to meet cellular energy demand. Adenosine mediation involves activation of a family of four G protein-coupled adenosine receptors (ARs): A(1), A(2)A, A(2)B, and A(3). The A(3) adenosine receptor (A(3)AR) is the only adenosine subtype to be overexpressed in inflammatory and cancer cells, thus making it a potential target for therapy. Originally isolated as an orphan receptor, A(3)AR presented a twofold nature under different pathophysiologic conditions: it appeared to be protective/harmful under ischemic conditions, pro/anti-inflammatory, and pro/antitumoral depending on the systems investigated. Until recently, the greatest and most intriguing challenge has been to understand whether, and in which cases, selective A(3) agonists or antagonists would be the best choice. Today, the choice has been made and A(3)AR agonists are now under clinical development for some disorders including rheumatoid arthritis, psoriasis, glaucoma, and hepatocellular carcinoma. More specifically, the interest and relevance of these new agents derives from clinical data demonstrating that A(3)AR agonists are both effective and safe. Thus, it will become apparent in the present review that purine scientists do seem to be getting closer to their goal: the incorporation of adenosine ligands into drugs with the ability to save lives and improve human health.

  7. Adenosine 5'-tetraphosphate and adenosine 5'-pentaphosphate are synthesized by yeast acetyl coenzyme A synthetase.

    PubMed Central

    Guranowski, A; Günther Sillero, M A; Sillero, A

    1994-01-01

    Yeast (Saccharomyces cerevisiae) acetyl coenzyme A (CoA) synthetase (EC 6.2.1.1) catalyzes the synthesis of adenosine 5'-tetraphosphate (P4A) and adenosine 5'-pentaphosphate (p5A) from ATP and tri- or tetrapolyphosphate (P3 or P4), with relative velocities of 7:1, respectively. Of 12 nucleotides tested as potential donors of nucleotidyl moiety, only ATP, adenosine-5'-O-[3-thiotriphosphate], and acetyl-AMP were substrates, with relative velocities of 100, 62, and 80, respectively. The Km values for ATP, P3, and acetyl-AMP were 0.16, 4.7, and 1.8 mM, respectively. The synthesis of p4A could proceed in the absence of exogenous acetate but was stimulated twofold by acetate, with an apparent Km value of 0.065 mM. CoA did not participate in the synthesis of p4A (p5A) and inhibited the reaction (50% inhibitory concentration of 0.015 mM). At pH 6.3, which was optimum for formation of p4A (p5A), the rate of acetyl-CoA synthesis (1.84 mumol mg-1 min-1) was 245 times faster than the rate of synthesis of p4A measured in the presence of acetate. The known formation of p4A (p5A) in yeast sporulation and the role of acetate may therefore be related to acetyl-CoA synthetase. Images PMID:7910605

  8. Adenosine deaminase 1 and concentrative nucleoside transporters 2 and 3 regulate adenosine on the apical surface of human airway epithelia: implications for inflammatory lung diseases.

    PubMed

    Hirsh, Andrew J; Stonebraker, Jaclyn R; van Heusden, Catja A; Lazarowski, Eduardo R; Boucher, Richard C; Picher, Maryse

    2007-09-11

    Adenosine is a multifaceted signaling molecule mediating key aspects of innate and immune lung defenses. However, abnormally high airway adenosine levels exacerbate inflammatory lung diseases. This study identifies the mechanisms regulating adenosine elimination from the apical surface of human airway epithelia. Experiments conducted on polarized primary cultures of nasal and bronchial epithelial cells showed that extracellular adenosine is eliminated by surface metabolism and cellular uptake. The conversion of adenosine to inosine was completely inhibited by the adenosine deaminase 1 (ADA1) inhibitor erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA). The reaction exhibited Km and Vmax values of 24 microM and 0.14 nmol x min(-1) x cm(-2). ADA1 (not ADA2) mRNA was detected in human airway epithelia. The adenosine/mannitol permeability coefficient ratio (18/1) indicated a minor contribution of paracellular absorption. Adenosine uptake was Na+-dependent and was inhibited by the concentrative nucleoside transporter (CNT) blocker phloridzin but not by the equilibrative nucleoside transporter (ENT) blocker dipyridamole. Apparent Km and Vmax values were 17 microM and 7.2 nmol x min(-1) x cm(-2), and transport selectivity was adenosine = inosine = uridine > guanosine = cytidine > thymidine. CNT3 mRNA was detected throughout the airways, while CNT2 was restricted to nasal epithelia. Inhibition of adenosine elimination by EHNA or phloridzin raised apical adenosine levels by >3-fold and stimulated IL-13 and MCP-1 secretion by 6-fold. These responses were reproduced by the adenosine receptor agonist 5'-(N-ethylcarboxamido)adenosine (NECA) and blocked by the adenosine receptor antagonist, 8-(p-sulfophenyl) theophylline (8-SPT). This study shows that adenosine elimination on human airway epithelia is mediated by ADA1, CNT2, and CNT3, which constitute important regulators of adenosine-mediated inflammation.

  9. Involvement of A1 adenosine receptors in altered vascular responses and inflammation in an allergic mouse model of asthma

    PubMed Central

    Ponnoth, Dovenia S.; Nadeem, Ahmed; Tilley, Stephen

    2010-01-01

    Poor lung function and respiratory disorders like asthma have a positive correlation with the development of adverse cardiovascular events. Increased adenosine levels are associated with lung inflammation that could lead to altered vascular responses and systemic inflammation. We hypothesized that asthmatic lung inflammation has systemic effects through A1 adenosine receptors (A1AR) and investigated the effects of aerosolized adenosine on vascular reactivity and inflammation, using A1AR knockout (A1KO) and corresponding wild-type (A1WT) mice that were divided into three experimental groups each: control (CON), allergen sensitized and challenged (SEN), and SEN + aerosolized adenosine (SEN + AD). Animals were sensitized with ragweed (200 μg ip; days 1 and 6), followed by 1% ragweed aerosol challenges (days 11 to 13). On day 14, the SEN + AD groups received one adenosine aerosol challenge (6 mg/ml) for 2 min, and aortae were collected on day 15. 5′-N-ethylcarboxamidoadenosine (NECA; nonselective adenosine analog) induced concentration-dependent aortic relaxation in the A1WT CON group, which was impaired in the A1WT SEN and SEN + AD groups. All groups of A1KO mice showed similar (no significant difference) concentration-dependent relaxation to NECA. The A1WT SEN and SEN + AD groups had a significantly higher contraction to selective A1 agonist 2-chloro-N6-cyclopentyladenosine (CCPA) compared with the CON group. Western blot data showed that aortic A1AR expression was significantly increased in WT SEN and SEN + AD mice compared with CON mice. Gene expression of ICAM-1 and IL-5 was significantly increased in allergic A1WT aorta and were undetected in the A1KO groups. A1WT allergic mice had significantly higher airway hyperresponsiveness (enhanced pause) to NECA, with adenosine aerosol further enhancing it. In conclusion, allergic A1WT mice showed altered vascular reactivity, increased airway hyperresponsiveness, and systemic inflammation. These data suggest that A1AR

  10. [60]Fullerene derivative modulates adenosine and metabotropic glutamate receptors gene expression: a possible protective effect against hypoxia

    PubMed Central

    2014-01-01

    Background Glutamate, the main excitatory neurotransmitter, is involved in learning and memory processes but at higher concentration results excitotoxic causing degeneration and neuronal death. Adenosine is a nucleoside that exhibit neuroprotective effects by modulating of glutamate release. Hypoxic and related oxidative conditions, in which adenosine and metabotropic glutamate receptors are involved, have been demonstrated to contribute to neurodegenerative processes occurring in certain human pathologies. Results Human neuroblastoma cells (SH-SY5Y) were used to evaluate the long time (24, 48 and 72 hours) effects of a [60]fullerene hydrosoluble derivative (t3ss) as potential inhibitor of hypoxic insult. Low oxygen concentration (5% O2) caused cell death, which was avoided by t3ss exposure in a concentration dependent manner. In addition, gene expression analysis by real time PCR of adenosine A1, A2A and A2B and metabotropic glutamate 1 and 5 receptors revealed that t3ss significantly increased A1 and mGlu1 expression in hypoxic conditions. Moreover, t3ss prevented the hypoxia-induced increase in A2A mRNA expression. Conclusions As t3ss causes overexpression of adenosine A1 and metabotropic glutamate receptors which have been shown to be neuroprotective, our results point to a radical scavenger protective effect of t3ss through the enhancement of these neuroprotective receptors expression. Therefore, the utility of these nanoparticles as therapeutic target to avoid degeneration and cell death of neurodegenerative diseases is suggested. PMID:25123848

  11. [Gene therapy for adenosine deaminase deficiency].

    PubMed

    Sakiyama, Yukio; Ariga, Tadashi; Ohtsu, Makoto

    2005-03-01

    A four year-old boy with adenosine deaminase (ADA-) deficient severe combined immunodeficiency(SCID) receiving PEG-ADA was treated under a gene therapy protocol targeting peripheral blood lymphocytes (PBLs) in 1995. After eleven infusions of autologous PBLs transduced with retroviral vector LASN encoding ADAcDNA, he exhibited increased levels of the CD8+ T lymphocytes, serum immunoglobulin, specific antibodies and delayed type hypersensitivity skin tests. Follow-up studies also provided evidence of long-term persistence and function of transduced PBLs with improvement in the immune function. However, the therapeutic effect of this gene therapy has been difficult to assess because of the concomitant treatment of PEG-ADA. Two ADA-SCID patients have been currently treated with autologous bone marrow CD34+ cells engineered with a retroviral vector GCsapM-ADA after discontinuation of PEG-ADA. The restoration of intracellular ADA enzymatic activity in lymphocytes and granulocytes resulted in correction of the systemic toxicity and liver function in the absence of PEG-ADA treatment. Both patients are at home where they are clinically well, and they do not experience adversed effect, with follow up being 12 months after CD34+ cells gene therapy.

  12. Detecting adenosine triphosphate in the pericellular space.

    PubMed

    Falzoni, Simonetta; Donvito, Giovanna; Di Virgilio, Francesco

    2013-06-06

    Release of adenosine triphosphate (ATP) into the extracellular space occurs in response to a multiplicity of physiological and pathological stimuli in virtually all cells and tissues. A role for extracellular ATP has been identified in processes as different as neurotransmission, endocrine and exocrine secretion, smooth muscle contraction, bone metabolism, cell proliferation, immunity and inflammation. However, ATP measurement in the extracellular space has proved a daunting task until recently. To tackle this challenge, some years ago, we designed and engineered a novel luciferase probe targeted to and expressed on the outer aspect of the plasma membrane. This novel probe was constructed by appending to firefly luciferase the N-terminal leader sequence and the C-terminal glycophosphatidylinositol anchor of the folate receptor. This chimeric protein, named plasma membrane luciferase, is targeted and localized to the outer side of the plasma membrane. With this probe, we have generated stably transfected HEK293 cell clones that act as an in vitro and in vivo sensor of the extracellular ATP concentration in several disease conditions, such as experimentally induced tumours and inflammation.

  13. Adenosine triphosphate inhibits melatonin synthesis in the rat pineal gland.

    PubMed

    Souza-Teodoro, Luis Henrique; Dargenio-Garcia, Letícia; Petrilli-Lapa, Camila Lopes; Souza, Ewerton da Silva; Fernandes, Pedro A C M; Markus, Regina P; Ferreira, Zulma S

    2016-03-01

    Adenosine triphosphate (ATP) is released onto the pinealocyte, along with noradrenaline, from sympathetic neurons and triggers P2Y1 receptors that enhance β-adrenergic-induced N-acetylserotonin (NAS) synthesis. Nevertheless, the biotransformation of NAS into melatonin, which occurs due to the subsequent methylation by acetylserotonin O-methyltransferase (ASMT; EC 2.1.1.4), has not yet been evaluated in the presence of purinergic stimulation. We therefore evaluated the effects of purinergic signaling on melatonin synthesis induced by β-adrenergic stimulation. ATP increased NAS levels, but, surprisingly, inhibited melatonin synthesis in an inverse, concentration-dependent manner. Our results demonstrate that enhanced NAS levels, which depend on phospholipase C (PLC) activity (but not the induction of gene transcription), are a post-translational effect. By contrast, melatonin reduction is related to an ASMT inhibition of expression at both the gene transcription and protein levels. These results were independent of nuclear factor-kappa B (NF-kB) translocation. Neither the P2Y1 receptor activation nor the PLC-mediated pathway was involved in the decrease in melatonin, indicating that ATP regulates pineal metabolism through different mechanisms. Taken together, our data demonstrate that purinergic signaling differentially modulates NAS and melatonin synthesis and point to a regulatory role for ATP as a cotransmitter in the control of ASMT, the rate-limiting enzyme in melatonin synthesis. The endogenous production of melatonin regulates defense responses; therefore, understanding the mechanisms involving ASMT regulation might provide novel insights into the development and progression of neurological disorders since melatonin presents anti-inflammatory, neuroprotective, and neurogenic effects.

  14. Adenosine acts as an inhibitor of lymphoma cell growth: a major role for the A3 adenosine receptor.

    PubMed

    Fishman, P; Bar-Yehuda, S; Ohana, G; Pathak, S; Wasserman, L; Barer, F; Multani, A S

    2000-07-01

    In this study, we demonstrated several mechanisms exploring the inhibitory effect of low-dose adenosine on lymphoma cell growth. Adenosine, a purine nucleoside present in plasma and other extracellular fluids, acts as a regulatory molecule, by binding to G-protein associated cell-surface receptors, A1, A2 and A3. Recently we showed that low-dose adenosine released by muscle cells, inhibits tumour cell growth and thus attributes to the rarity of muscle metastases. In the present work, a cytostatic effect of adenosine on the proliferation of the Nb2-11C rat lymphoma cell line was demonstrated. This effect was mediated through the induction of cell cycle arrest in the G0/G1 phase and by decreasing the telomeric signal in these cells. Adenosine was found to exert its antiproliferative effect mainly through binding to its A3 receptor. The cytostatic anticancer activity, mediated through the A3 adenosine receptor, turns it into a potential target for the development of anticancer therapies.

  15. Adenosine deaminase from Streptomyces coelicolor: recombinant expression, purification and characterization.

    PubMed

    Pornbanlualap, Somchai; Chalopagorn, Pornchanok

    2011-08-01

    The sequencing of the genome of Streptomyces coelicolor A3(2) identified seven putative adenine/adenosine deaminases and adenosine deaminase-like proteins, none of which have been biochemically characterized. This report describes recombinant expression, purification and characterization of SCO4901 which had been annotated in data bases as a putative adenosine deaminase. The purified putative adenosine deaminase gives a subunit Mr=48,400 on denaturing gel electrophoresis and an oligomer molecular weight of approximately 182,000 by comparative gel filtration. These values are consistent with the active enzyme being composed of four subunits with identical molecular weights. The turnover rate of adenosine is 11.5 s⁻¹ at 30 °C. Since adenine is deaminated ∼10³ slower by the enzyme when compared to that of adenosine, these data strongly show that the purified enzyme is an adenosine deaminase (ADA) and not an adenine deaminase (ADE). Other adenine nucleosides/nucleotides, including 9-β-D-arabinofuranosyl-adenine (ara-A), 5'-AMP, 5'-ADP and 5'-ATP, are not substrates for the enzyme. Coformycin and 2'-deoxycoformycin are potent competitive inhibitors of the enzyme with inhibition constants of 0.25 and 3.4 nM, respectively. Amino acid sequence alignment of ScADA with ADAs from other organisms reveals that eight of the nine highly conserved catalytic site residues in other ADAs are also conserved in ScADA. The only non-conserved residue is Asn317, which replaces Asp296 in the murine enzyme. Based on these data, it is suggested here that ADA and ADE proteins are divergently related enzymes that have evolved from a common α/β barrel scaffold to catalyze the deamination of different substrates, using a similar catalytic mechanism. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Role of A3 adenosine receptor in diabetic neuropathy.

    PubMed

    Yan, Heng; Zhang, Enshui; Feng, Chang; Zhao, Xin

    2016-10-01

    Neuropathy is the most common diabetic complication. Although the A1 and A2A adenosine receptors are important pharmacological targets in alleviating diabetic neuropathy, the role of the A3 adenosine receptor remains unknown. Because the A3 adenosine receptor regulates pain induced by chronic constriction injury or chemotherapy, its stimulation might also attenuate diabetic neuropathy. This study examines the effects of systemic treatment with the A3 adenosine receptor agonist 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-β-d-ribofuranuronamide (IB-MECA) on diabetic neuropathy and explores the putative mechanisms underlying its pharmacological effects. We show that IB-MECA alleviated mechanical hyperalgesia and thermal hypoalgesia in mice 2 weeks but not 4 weeks after streptozocin (STZ) treatment. Furthermore, IB-MECA prevented the reduction in sciatic motor nerve conduction velocity and sensory nerve conduction velocity in diabetic mice 2 weeks but not 4 weeks after STZ treatment. Similarly, IB-MECA inhibited the activation of nuclear factor-κB and decreased the generation of tumor necrosis factor-α in the spinal cord of mice 2 weeks but not 4 weeks after STZ treatment. These phenomena were associated with reduction of A3 adenosine receptor expression in the spinal cord after long-term diabetes. Our results suggest that the A3 adenosine receptor plays a critical role in regulating diabetic neuropathy and that reduction in A3 adenosine receptor expression/function might contribute to the progression of diabetic neuropathy. © 2016 Wiley Periodicals, Inc.

  17. "Dormant transisthmus conduction" revealed by adenosine after cavotricuspid isthmus ablation.

    PubMed

    Lehrmann, Heiko; Weber, Reinhold; Park, Chan-il; Allgeier, Jürgen; Schiebeling-Römer, Jochen; Arentz, Thomas; Jadidi, Amir

    2012-12-01

    Linear radiofrequency ablation at the cavotricuspid isthmus (CTI) is the treatment of choice for typical flutter. Despite a high acute success rate, reconduction through the CTI may occur in approximately 15% of patients and eventually lead to flutter recurrence. The purpose of this study was to test the hypothesis that injection of adenosine may reveal transient CTI reconduction and predict early relapse of permanent CTI conduction. Thirty-one patients with CTI-dependent flutter (mean age 65 ± 11 years, 87% male, ejection fraction 55% ± 11%) were included in the study. CTI ablation was performed using an open-irrigated ablation catheter. Bidirectional conduction block was confirmed using conventional criteria. Subsequently, transisthmus conduction was reevaluated after adenosine injection. During a 30-minute waiting period, permanent recovery of CTI conduction was monitored. During a mean follow-up of 6 ± 3 months, clinical recurrences of typical flutter were assessed. Bidirectional isthmus block was achieved in all patients. Injection of 16 ± 3 mg adenosine IV induced transient second- or third-degree AV block in all patients. An adenosine-induced brief sequence reversal at the right lateral wall occurred in 6 of 31 patients (19%) and revealed transient CTI reconduction. Among these 6 patients, 4 (67%) had permanent recovery of transisthmus conduction in the subsequent waiting period; the remaining 2 patients (33%) had clinical recurrence of common flutter. Importantly, no patient without adenosine-mediated dormant transisthmus conduction (25/31 [81%]) showed permanent recovery during the waiting period or clinical flutter recurrence during follow-up. Adenosine-induced "dormant transisthmus conduction" precedes early relapse of permanent CTI conduction. Patients without "dormant transisthmus conduction" develop no recovery of conduction during the postablation waiting period. Routine use of adenosine for assessment of ablation lines may help to reduce the

  18. Preferential activation of excitatory adenosine receptors at rat hippocampal and neuromuscular synapses by adenosine formed from released adenine nucleotides.

    PubMed Central

    Cunha, R. A.; Correia-de-Sá, P.; Sebastião, A. M.; Ribeiro, J. A.

    1996-01-01

    1. In the present work, we investigated the action of adenosine originating from extracellular catabolism of adenine nucleotides, in two preparations where synaptic transmission is modulated by both inhibitory A1 and excitatory A(2a)-adenosine receptors, the rat hippocampal Schaffer fibres/CA1 pyramid synapses and the rat innervated hemidiaphragm. 2. Endogenous adenosine tonically inhibited synaptic transmission, since 0.5-2 u ml-1 of adenosine deaminase increased both the population spike amplitude (30 +/- 4%) and field excitatory post-synaptic potential (f.e.p.s.p.) slope (27 +/- 4%) recorded from hippocampal slices and the evoked [3H]-acetylcholine ([3H]-ACh) release from the motor nerve terminals (25 +/- 2%). 3. alpha, beta-Methylene adenosine diphosphate (AOPCP) in concentrations (100-200 microM) that almost completely inhibited the formation of adenosine from the extracellular catabolism of AMP, decreased population spike amplitude by 39 +/- 5% and f.e.p.s.p. slope by 32 +/- 3% in hippocampal slices and [3H]-ACh release from motor nerve terminals by 27 +/- 3%. 4. Addition of exogenous 5'-nucleotidase (5 u ml-1) prevented the inhibitory effect of AOPCP on population spike amplitude and f.e.p.s.p. slope by 43-57%, whereas the P2 antagonist, suramin (100 microM), did not modify the effect of AOPCP. 5. In both preparations, the effect of AOPCP resulted from prevention of adenosine formation since it was no longer evident when accumulation of extracellular adenosine was hindered by adenosine deaminase (0.5-2 u ml-1). The inhibitory effect of AOPCP was still evident when A1 receptors were blocked by 1,3-dipropyl-8-cyclopentylxanthine (2.5-5 nM), but was abolished by the A2 antagonist, 3,7-dimethyl-1-propargylxanthine (10 microM). 6. These results suggest that adenosine originating from catabolism of released adenine nucleotides preferentially activates excitatory A2 receptors in hippocampal CAI pyramid synapses and in phrenic motor nerve endings. PMID:8886406

  19. Adenosine and protection from acute kidney injury

    PubMed Central

    Yap, Steven C.; Lee, H. Thomas

    2012-01-01

    Purpose of Review Acute Kidney Injury (AKI) is a major clinical problem without effective therapy. Development of AKI among hospitalized patients drastically increases mortality, and morbidity. With increases in complex surgical procedures together with a growing elderly population, the incidence of AKI is rising. Renal adenosine receptor (AR) manipulation may have great therapeutic potential in mitigating AKI. In this review, we discuss renal AR biology and potential clinical therapies for AKI. Recent Findings The 4 AR subtypes (A1AR, A2AAR, A2BAR and A3AR) have diverse effects on the kidney. The pathophysiology of AKI may dictate the specific AR subtype activation needed to produce renal protection. The A1AR activation in renal tubules and endothelial cells produces beneficial effects against ischemia and reperfusion (IR) injury by modulating metabolic demand, decreasing necrosis, apoptosis and inflammation. The A2AAR protects against AKI by modulating leukocyte-mediated renal and systemic inflammation whereas the A2BAR activation protects by direct activation of renal parenchymal ARs. In contrast, the A1AR antagonism may play a protective role in nephrotoxic AKI and radiocontrast induced nephropathy by reversing vascular constriction and inducing naturesis and diuresis. Furthermore, as the A3AR-activation exacerbates apoptosis and tissue damage due to renal IR, selective A3AR antagonism may hold promise to attenuate renal IR injury. Finally, renal A1AR activation also protects against renal endothelial dysfunction caused by hepatic IR injury. Summary Despite the current lack of therapies for the treatment and prevention of AKI, recent research suggests that modulation of renal ARs holds promise in treating AKI and extrarenal injury. PMID:22080856

  20. High salt diet exacerbates vascular contraction in the absence of adenosine A₂A receptor.

    PubMed

    Pradhan, Isha; Zeldin, Darryl C; Ledent, Catherine; Mustafa, Jamal S; Falck, John R; Nayeem, Mohammed A

    2014-05-01

    High salt (4% NaCl, HS) diet modulates adenosine-induced vascular response through adenosine A(2A) receptor (A(2A)AR). Evidence suggests that A(2A)AR stimulates cyp450-epoxygenases, leading to epoxyeicosatrienoic acids (EETs) generation. The aim of this study was to understand the vascular reactivity to HS and underlying signaling mechanism in the presence or absence of A(2A)AR. Therefore, we hypothesized that HS enhances adenosine-induced relaxation through EETs in A(2A)AR⁺/⁺, but exaggerates contraction in A(2A)AR⁻/⁻. Organ bath and Western blot experiments were conducted in HS and normal salt (NS, 0.18% NaCl)-fed A(2A)AR⁺/⁺ and A(2A)AR⁻/⁻ mice aorta. HS produced concentration-dependent relaxation to non-selective adenosine analog, NECA in A(2A)AR⁺/⁺, whereas contraction was observed in A(2A)AR⁻/⁻ mice and this was attenuated by A₁AR antagonist (DPCPX). CGS 21680 (selective A(2A)AR agonist) enhanced relaxation in HS-A(2A)AR⁺/⁺ versus NS-A(2A)AR⁺/⁺, which was blocked by EETs antagonist (14,15-EEZE). Compared with NS, HS significantly upregulated the expression of vasodilators A(2A)AR and cyp2c29, whereas vasoconstrictors A₁AR and cyp4a in A(2A)AR⁺/⁺ were downregulated. In A(2A)AR⁻/⁻ mice, however, HS significantly downregulated the expression of cyp2c29, whereas A₁AR and cyp4a were upregulated compared with A(2A)AR⁺/⁺ mice. Hence, our data suggest that in A(2A)AR⁺/⁺, HS enhances A(2A)AR-induced relaxation through increased cyp-expoxygenases-derived EETs and decreased A₁AR levels, whereas in A(2A)AR⁻/⁻, HS exaggerates contraction through decreased cyp-epoxygenases and increased A₁AR levels.

  1. No effect of nutritional adenosine receptor antagonists on exercise performance in the heat.

    PubMed

    Cheuvront, Samuel N; Ely, Brett R; Kenefick, Robert W; Michniak-Kohn, Bozena B; Rood, Jennifer C; Sawka, Michael N

    2009-02-01

    Nutritional adenosine receptor antagonists can enhance endurance exercise performance in temperate environments, but their efficacy during heat stress is not well understood. This double-blinded, placebo-controlled study compared the effects of an acute dose of caffeine or quercetin on endurance exercise performance during compensable heat stress (40 degrees C, 20-30% rh). On each of three occasions, 10 healthy men each performed 30-min of cycle ergometry at 50% Vo2peak followed by a 15-min performance time trial after receiving either placebo (Group P), caffeine (Group C; 9 mg/kg), or quercetin (Group Q; 2,000 mg). Serial blood samples, physiological (heart rate, rectal, and mean skin body temperatures), perceptual (ratings of perceived exertion, pain, thermal comfort, motivation), and exercise performance measures (total work and pacing strategy) were made. Supplementation with caffeine and quercetin increased preexercise blood concentrations of caffeine (55.62 +/- 4.77 microM) and quercetin (4.76 +/- 2.56 microM) above their in vitro inhibition constants for adenosine receptors. No treatment effects were observed for any physiological or perceptual measures, with the exception of elevated rectal body temperatures (0.20-0.30 degrees C; P < 0.05) for Group C vs. Groups Q and P. Supplementation did not affect total work performed (Groups P: 153.5 +/- 28.3, C: 157.3 +/- 28.9, and Q: 151.1 +/- 31.6 kJ; P > 0.05) or the self-selected pacing strategy employed. These findings indicate that the nutritional adenosine receptor antagonists caffeine and quercetin do not enhance endurance exercise performance during compensable heat stress.

  2. Metformin augments doxorubicin cytotoxicity in mammary carcinoma through activation of adenosine monophosphate protein kinase pathway.

    PubMed

    El-Ashmawy, Nahla E; Khedr, Naglaa F; El-Bahrawy, Hoda A; Abo Mansour, Hend E

    2017-05-01

    Since the incidence of breast cancer increases dramatically all over the world, the search for effective treatment is an urgent need. Metformin has demonstrated anti-tumorigenic effect both in vivo and in vitro in different cancer types. This work was designed to examine on molecular level the mode of action of metformin in mice bearing solid Ehrlich carcinoma and to evaluate the use of metformin in conjunction with doxorubicin as a combined therapy against solid Ehrlich carcinoma. Ehrlich ascites carcinoma cells were inoculated in 60 female mice as a model of breast cancer. The mice were divided into four equal groups: Control tumor, metformin, doxorubicin, and co-treatment. Metformin (15 mg/kg) and doxorubicin (4 mg/kg) were given intraperitoneally (i.p.) for four cycles every 5 days starting on day 12 of inoculation. The anti-tumorigenic effect of metformin was mediated by enhancement of adenosine monophosphate protein kinase activity and elevation of P53 protein as well as the suppression of nuclear factor-kappa B, DNA contents, and cyclin D1 gene expression. Metformin and doxorubicin mono-treatments exhibited opposing action regarding cyclin D1 gene expression, phosphorylated adenosine monophosphate protein kinase, and nuclear factor-kappa B levels. Co-treatment markedly decreased tumor volume, increased survival rate, and improved other parameters compared to doxorubicin group. In parallel, the histopathological findings demonstrated enhanced apoptosis and absence of necrosis in tumor tissue of co-treatment group. Metformin proved chemotherapeutic effect which could be mediated by the activation of adenosine monophosphate protein kinase and related pathways. Combining metformin and doxorubicin, which exhibited different mechanisms of action, produced greater efficacy as anticancer therapeutic regimen.

  3. Amplified fluorescence detection of adenosine via catalyzed hairpin assembly and host-guest interactions between β-cyclodextrin polymer and pyrene.

    PubMed

    Huang, Haihua; Yang, Xiaohai; Wang, Kemin; Wang, Qing; Guo, Qiuping; Huang, Jin; Liu, Jianbo; Song, Chunxia

    2016-04-21

    Nowadays, enzyme-free nucleic acid-based signal amplification strategies are frequently utilized in the design of biosensors due to their excellent sensitivity. Developing more extended analytical methods is fundamental for basic studies in the biological and biomedical fields. Herein, we introduce an enzyme-free amplified detection strategy for the small molecule adenosine. The approach is based on adenosine-aptamer binding triggered catalyzed hairpin assembly and host-guest interactions between β-cyclodextrin polymer (β-CDP) and pyrene. Two hairpin probes (probe H1 and probe H2) and an aptamer-trigger/inhibitor duplex probe were employed in the system and the pyrene-labeled probe H1 was chosen as the signal unit. In the absence of adenosine, the aptamer-trigger was inhibited by the inhibitor strand. The hairpin probes were in the closed hairpin formation without activation of the trigger strand. Pyrene labeled at the 5'-termini of the single-stranded stem of probe H1 could be easily trapped in the hydrophobic cavity of β-CDP because of weak steric hindrance, leading to a significant fluorescence enhancement. Once the hairpin assembly was catalyzed by the adenosine-aptamer binding event, a hybridized DNA duplex H1/H2 was created continuously. Pyrene labeled at the 5'-termini of the DNA duplex H1/H2 finds it difficult to enter the cavity of β-CDP due to steric hindrance, leading to a weaker fluorescence signal. Thus, the target could be detected by this simple mix-and-detect amplification method without a need for expensive and perishable protein enzymes. As low as 42 nM of adenosine was detected by this assay, which is comparable to that of some reported colorimetric methods. Meanwhile, the proposed method was further successfully applied to detect adenosine in human serum samples, showing great potential for adenosine detection from complex fluids.

  4. Effect of adenosine on the growth of human T-lymphocyte leukemia cell line MOLT-4.

    PubMed

    Streitová, Denisa; Weiterová, Lenka; Hofer, Michal; Holá, Jirina; Horváth, Viktor; Kozubík, Alois; Znojil, Vladimír

    2007-09-01

    Adenosine has been observed to suppress the growth of MOLT-4 human leukemia cells in vitro. Changes in the cell cycle, especially increased percentage of cells in S phase, prolonged generation time, and induction of apoptosis at higher adenosine concentrations have been found to be responsible for the growth suppression. Dipyridamole, a drug inhibiting the cellular uptake of adenosine, reversed partially but significantly the adenosine-induced growth suppression. It follows from these results that the action of adenosine on the MOLT-4 cells comprises its cellular uptake and intracellular operation. These findings present new data on anticancer efficacy of adenosine.

  5. High plasma adenosine levels in overweight/obese pregnant women.

    PubMed

    Badillo, Priscila; Salgado, Paola; Bravo, Patricia; Guevara, Katherine; Acurio, Jesenia; Gonzalez, Maria Angelica; Oyarzun, Carlos; San Martin, Rody; Escudero, Carlos

    2017-07-18

    We aim to investigate whether overweight/obese pregnant women have elevated plasma levels of adenosine associated with increased consumption of high-calorie food. Sixty women were included. They were divided into lean (n = 23 and n = 12) or overweight/obese (n = 7 and n = 18) non-pregnant and pregnant women, respectively. Clinical records and maternal blood samples were collected after informed consent. A self-reported dietary questionnaire was also completed. Plasma adenosine levels were determined with high-performance liquid chromatography. Biochemical parameters, including glucose, total protein, and lipid profile, were determined using standard colorimetric assays. Adenosine levels were higher in pregnant women than in non-pregnant women (18.7 ± 1.6 vs 10.8 ± 1.3 nM/μg protein, respectively, p < 0.0001). Overweight/obese pregnant women (21.9 ± 2.5 nM/μg protein) exhibited higher adenosine levels than lean pregnant (14.5 ± 1.0 nM/μg protein, p = 0.04) or non-pregnant women (11.7 ± 1.5 nM/μg protein, p = 0.0005). Also, pregnant women with elevated weight gain exhibited higher (26.2 ± 3.7 nM/μg protein) adenosine levels than those with adequate weight gain (14.9 ± 1.4 nM/μg protein, p = 0.03). These differences were not statistically significant compared with those of pregnant women with reduced weight gain (17.4 ± 2.1 nM/μg protein, p = 0.053). Body mass index and adenosine only in pregnant women were positively correlated (r = 0.39, p = 0.02). While, polyunsaturated fatty acid (PUFA) consumption was negatively correlated with plasma adenosine levels only in non-pregnant women (r = -0.33, p = 0.03). Pregnancy is associated with high plasma adenosine levels, which are further elevated in pregnant women who are overweight/obese. High PUFA intake might reduce plasma adenosine levels in non-pregnant women.

  6. Attenuation of exercise vasodilatation by adenosine deaminase in anaesthetized dogs.

    PubMed Central

    Goonewardene, I P; Karim, F

    1991-01-01

    1. In dogs anaesthetized with sodium pentobarbitone and artificially ventilated, the gracilis muscles were vascularly isolated and perfused at a constant flow of 28.4 +/- 4.6 ml min-1 (100 g muscle tissue)-1 (99.8 +/- 4.5% of maximum free flow, means +/- standard error of the mean (S.E.M.), n = 9). 2. Three to five minutes of electrical stimulation of the cut peripheral end of the obturator nerve (4 Hz, 6 V, 0.2 ms) resulted in muscle contraction (0.61 +/- 0.14 kg (100 g)-1 during solvent infusion and 0.56 +/- 0.10 kg (100 g)-1 during intra-arterial adenosine deaminase infusion (50 U min-1) and an immediate decrease in arterial perfusion pressure from 184.5 +/- 8.1 mmHg to 148.2 +/- 5.7 mmHg (18.7 +/- 3.4% decrease) during solvent infusion, and from 193.5 +/- 7.16 to 142.0 +/- 10.2 mmHg (25.4 +/- 6.1% decrease) during adenosine deaminase infusion 10 s after the commencement of muscle stimulation. After about 5 min of muscle contractions, the arterial perfusion pressure decreased to 120.8 +/- 7.8 mmHg (32.9 +/- 5.8% decrease) during solvent infusion, and to 152.8 +/- 11.2 mmHg (20.9 +/- 5.3% decrease) during adenosine deaminase infusion (i.e. 37.9 +/- 6.2% attenuation of the fall in arterial perfusion pressure). The time taken for 90% recovery of the arterial perfusion pressure was 72.1 +/- 10.9 s during solvent infusion, and 51.5 +/- 9.3 s during adenosine deaminase infusion (P less than 0.05). 3. Adenosine (2 x 10(-3) mol l-1) infusion in the resting muscle during solvent infusion (final concentration in arterial blood 1.3 x 10(-4) +/- 6.0 x 10(-5) mol l-1) resulted in a 34.8 +/- 7.2% fall in arterial perfusion pressure but a fall of only 7.2 +/- 1.8% during adenosine deaminase infusion (50 U min-1; P less than 0.05; n = 5) indicating that adenosine deaminase infused at 50 U min-1 was more than adequate to metabolize endogenous adenosine produced during muscle contractions. 4. These data suggest that adenosine contributes about 40% to the sustained

  7. Adenosine signaling and the regulation of chronic lung disease

    PubMed Central

    Zhou, Yang; Schneider, Daniel J.; Blackburn, Michael R.

    2009-01-01

    Chronic lung diseases such as asthma, chronic obstructive pulmonary disease and interstitial lung disease are characterized by inflammation and tissue remodeling processes that compromise pulmonary function. Adenosine is produced in the inflamed and damaged lung where it plays numerous roles in the regulation of inflammation and tissue remodeling. Extracellular adenosine serves as an autocrine and paracrine signaling molecule by engaging cell surface adenosine receptors. Preclinical and cellular studies suggest that adenosine plays an anti-inflammatory role in processes associated with acute lung disease, where activation of the A2AR and A2BR have promising implications for the treatment of these disorders. In contrast, there is growing evidence that adenosine signaling through the A1R, A2BR and A3R may serve pro-inflammatory and tissue remodeling functions in chronic lung diseases. This review discusses the current progress of research efforts and clinical trials aimed at understanding the complexities of this signaling pathway as they pertain to the development of treatment strategies for chronic lung diseases. PMID:19426761

  8. Intrathecal adenosine administration in abdominal hysterectomy lacks analgesic effect.

    PubMed

    Rane, K; Sollevi, A; Segerdahl, M

    2000-08-01

    Adenosine (Ado) is known, from studies in both animals and humans, to produce antinociception when administered systemically or intrathecally (IT). The current aim was to evaluate, in a placebo-controlled, randomised, double-blind study, whether IT adenosine given before surgery could reduce anaesthetic requirement and the need of opioids during 48 h after visceral surgery. Forty women (37-66 years, ASA I and II) scheduled for elective hysterectomy were included. Before inducing the standardised O2/N2O/isoflurane/fentanyl anaesthesia, the patients received an IT injection of either adenosine (500 microg in 1 ml volume) or placebo 1 ml (saline). Intraoperative anaesthetic drug doses and haemodynamics were recorded. Postoperative pain was assessed by visual analogue scale. For postoperative analgesia, cetobemidone was provided via intravenous patient-controlled analgesia (PCA). During surgery, there were no differences between groups in anaesthetic requirement or haemodynamic parameters. Postoperative cetobemidone requirements were similar in both groups (median 48 mg for adenosine/50 mg for saline) during the first 48 postoperative hours. IT adenosine did not influence the requirement of anaesthetic drug or postoperative analgesics after hysterectomy.

  9. Dicinnamoylquinides in roasted coffee inhibit the human adenosine transporter.

    PubMed

    de Paulis, Tomas; Schmidt, Dennis E; Bruchey, Aleksandra K; Kirby, Michael T; McDonald, Michael P; Commers, Patricia; Lovinger, David M; Martin, Peter R

    2002-05-10

    Preliminary screening of a minor, non-xanthine constituent of roasted coffee, 3,4-diferuloyl-1,5-quinolactone (DIFEQ), showed inhibition of the adenosine transporter at low micromolar concentration. DIFEQ is a neutral derivative of the chlorogenic acids, i.e. isomeric mono- and di-substituted coumaroyl-, caffeoyl-, and feruloyl-esters of quinic acid, formed in the roasting process of coffee. Displacement of the adenosine transporter antagonist [(3)H](S)-(nitrobenzyl)-6-thioinosine binding by DIFEQ in cultured U-937 cell preparations, expressing the human adenosine transporter protein (hENT1), showed a K(i) of 0.96+/-0.13 microM. Extracts of regular and decaffeinated coffee showed binding activities equivalent to 30-40 mg DIFEQ per three cups of coffee. Acute administration of a high dose of DIFEQ (100 mg/kg i.p.) reduced open field locomotion in mice for 20 min in correlation with brain levels of DIFEQ. Both 3,4-dicaffeoyl-1,5-quinide and 3,4-dicoumaroyl-1,5-quinide, two close structural analogs of DIFEQ also present in roasted coffee, showed similar affinities for the adenosine transporter, while the corresponding 3- and 4-mono caffeoyl- and feruloyl-quinides were one to two orders of magnitudes less active. This suggests that 3,4-dicinnamoyl-1,5-quinides in coffee could have the potential to raise extra-cellular adenosine levels, thereby counteracting the stimulant effect of caffeine.

  10. Prostaglandins and adenosine in the regulation of sleep and wakefulness.

    PubMed

    Huang, Zhi-Li; Urade, Yoshihiro; Hayaishi, Osamu

    2007-02-01

    Prostaglandin (PG) D2 and adenosine are potent humoral sleep-inducing factors that accumulate in the brain during prolonged wakefulness. PGD2 is produced in the brain by lipocalin-type PGD synthase, which is localized mainly in the leptomeninges, choroid plexus and oligodendrocytes, and circulates in the cerebrospinal fluid as a sleep hormone. It stimulates DP1 receptors on leptomeningeal cells of the basal forebrain to release adenosine as a paracrine signaling molecule to promote sleep. Adenosine activates adenosine A2A receptor-expressing sleep-active neurons in the basal forebrain and the ventrolateral preoptic area. Sleep-promoting neurons in the ventrolateral preoptic area send inhibitory signals to suppress the histaminergic neurons in the tuberomammillary nucleus, which contribute to arousal through histamine H1 receptors. Increased knowledge of the molecular mechanisms by which PGD2 induces sleep through activation of adenosine A2A receptors and inhibition of the histaminergic arousal system will be useful both for a better understanding of sleep/wake regulation and for the development of novel types of sleeping pills or anti-doze drugs.

  11. Regioselective 1-N-Alkylation and Rearrangement of Adenosine Derivatives.

    PubMed

    Oslovsky, Vladimir E; Drenichev, Mikhail S; Mikhailov, Sergey N

    2015-01-01

    Several methods for the preparation of some N(6)-substituted adenosines based on selective 1-N-alkylation with subsequent Dimroth rearrangement were developed. The proposed methods seem to be effective for the preparation of natural N(6)-isopentenyl- and N(6)-benzyladenosines, which are known to possess pronounced biological activities. Direct 1-N-alkylation of 2',3',5'-tri-O-acetyladenosine and 3',5'-di-O-acetyl-2'-deoxyadenosine with alkyl halides in N,N-dimethylformamide (DMF) in the presence of BaCO3 and KI gave 1-N-substituted derivatives with quantitative yields, whereas 1-N-alkylation of adenosine was accompanied by significant O-alkylation. Moreover, the reaction of trimethylsilyl derivatives of N(6)-acetyl-2',3',5'-tri-O-acetyladenosine and N(6)-acetyl-3',5'-di-O-acetyl-2'-deoxyadenosine with alkyl halides leads to the formation of the stable 1-N-substituted adenosines. Dimroth rearrangement of 1-N-substituted adenosines in aqueous ammonia yields pure N(6)-substituted adenosines.

  12. Unpredictable Chronic Stress Alters Adenosine Metabolism in Zebrafish Brain.

    PubMed

    Zimmermann, F F; Altenhofen, S; Kist, L W; Leite, C E; Bogo, M R; Cognato, G P; Bonan, C D

    2016-05-01

    Stress is considered a risk factor for several human disorders. Despite the broad knowledge of stress responses in mammals, data on the relationship between unpredictable chronic stress (UCS) and its effects on purinergic signaling are limited. ATP hydrolysis by ectonucleotidases is an important source of adenosine, and adenosine deaminase (ADA) contributes to the control of the nucleoside concentrations. Considering that some stress models could affect signaling systems, the objective of this study was to investigate whether UCS alters ectonucleotidase and ADA pathway in zebrafish brain. Additionally, we analyzed ATP metabolism as well as ada1, ada2.1, ada2.2, adaL, and adaasi gene expression in zebrafish brain. Our results have demonstrated that UCS did not alter ectonucleotidase and soluble ADA activities. However, ecto-ADA activity was significantly decreased (26.8%) in brain membranes of animals exposed to UCS when compared to the control group. Quantitative reverse transcription PCR (RT-PCR) analysis did not show significant changes on ADA gene expression after the UCS exposure. The brain ATP metabolism showed a marked increase in adenosine levels (ADO) in animals exposed to UCS. These data suggest an increase on extracellular adenosine levels in zebrafish brain. Since this nucleoside has neuromodulatory and anxiolytic effects, changes in adenosine levels could play a role in counteracting the stress, which could be related to a compensatory mechanism in order to restore the homeostasis.

  13. Differential adenosine sensitivity of diaphragm and skeletal muscle arterioles.

    PubMed

    Aaker, Aaron; Laughlin, M H

    2002-09-01

    The hyperemic response in exercising skeletal muscle is dependent on muscle fiber-type composition and fiber recruitment patterns, but the vascular control mechanisms producing exercise hyperemia in skeletal muscle remain poorly understood. The purpose of this study was to test the hypothesis that arterioles from white, low-oxidative skeletal muscle are less responsive to adenosine-induced dilation than are arterioles from diaphragm (Dia) and red, high-oxidative skeletal muscle. Second-order arterioles (2As) were isolated from the white portion of gastrocnemius muscle (WG; low-oxidative, fast-twitch muscle tissue) and two types of high-oxidative skeletal muscle [Dia and red portion of gastrocnemius muscle (RG)] of rats. Results reveal that 2As from all three types of muscle dilated in response to the endothelium-dependent dilator acetylcholine (WG: 48 +/- 3%, Dia: 51 +/- 3%, RG: 74 +/- 3%). In contrast, adenosine dilated only 2As from WG (48 +/- 4%) and Dia (46 +/- 5%) but not those from RG (5 +/- 5%). Thus adenosine-induced dilator responses differed among 2As of these different types of muscle tissue. However, the results do not support our hypothesis because 2As from Dia and WG dilated in response to adenosine, whereas 2As from RG did not. We conclude that the adenosine responsiveness of 2As from rat skeletal muscle cannot be predicted only by the fiber-type composition or oxidative capacity of the skeletal muscle tissue wherein the arteriole lies.

  14. Glutamate-induced depression of EPSP-spike coupling in rat hippocampal CA1 neurons and modulation by adenosine receptors.

    PubMed

    Ferguson, Alexandra L; Stone, Trevor W

    2010-04-01

    The presence of high concentrations of glutamate in the extracellular fluid following brain trauma or ischaemia may contribute substantially to subsequent impairments of neuronal function. In this study, glutamate was applied to hippocampal slices for several minutes, producing over-depolarization, which was reflected in an initial loss of evoked population potential size in the CA1 region. Orthodromic population spikes recovered only partially over the following 60 min, whereas antidromic spikes and excitatory postsynaptic potentials (EPSPs) showed greater recovery, implying a change in EPSP-spike coupling (E-S coupling), which was confirmed by intracellular recording from CA1 pyramidal cells. The recovery of EPSPs was enhanced further by dizocilpine, suggesting that the long-lasting glutamate-induced change in E-S coupling involves NMDA receptors. This was supported by experiments showing that when isolated NMDA-receptor-mediated EPSPs were studied in isolation, there was only partial recovery following glutamate, unlike the composite EPSPs. The recovery of orthodromic population spikes and NMDA-receptor-mediated EPSPs following glutamate was enhanced by the adenosine A1 receptor blocker DPCPX, the A2A receptor antagonist SCH58261 or adenosine deaminase, associated with a loss of restoration to normal of the glutamate-induced E-S depression. The results indicate that the long-lasting depression of neuronal excitability following recovery from glutamate is associated with a depression of E-S coupling. This effect is partly dependent on activation of NMDA receptors, which modify adenosine release or the sensitivity of adenosine receptors. The results may have implications for the use of A1 and A2A receptor ligands as cognitive enhancers or neuroprotectants.

  15. The interaction of gabapentin and N6-(2-phenylisopropyl)-adenosine R-(-)isomer (R-PIA) on mechanical allodynia in rats with a spinal nerve ligation.

    PubMed

    Park, Jong Yeon; Jun, In Gu

    2008-08-01

    We examined the antiallodynic interaction between gabapentin and adenosine A1 receptor agonist, N(6)-(2-phenylisopropyl)-adenosine R-(-)isomer (R-PIA), in a rat model of nerve ligation injury. Rats were prepared with ligation of left L5-6 spinal nerves and intrathecal catheter implantation for drug administration. Mechanical allodynia was measured by applying von Frey filaments. Gabapentin and R-PIA were administered to obtain the dose-response curve and the 50% effective dose (ED(50)). Fractions of ED(50)s were administered concurrently to establish the ED(50) of the drug combination. The drug interaction between gabapentin and R-PIA was analyzed using the isobolographic method. Adenosine A1 receptor antagonist was administered intrathecally to examine the reversal of the antiallodynic effect. Locomotor function changes were evaluated by rotarod testing. Intrathecal gabapentin and R-PIA and their combination produced a dose-dependent antagonism against mechanical allodynia without severe side effects. Intrathecal gabapentin synergistically enhanced the antiallodynic effect of R-PIA when coadministered. There were no significant changes in rotarod performance time, except gabapentin 300 microg. In the combination group, the maximal antiallodynic effect was reversed by A1 adenosine receptor antagonist. These results suggest that activation of adenosine A1 receptors at the spinal level is required for the synergistic interaction on the mechanical allodynia.

  16. Abasic Site-Containing DNAzyme and Aptamer for Label-free Fluorescent Detection of Pb2+ and Adenosine with High Sensitivity, Selectivity and Tunable Dynamic Range

    PubMed Central

    Xiang, Yu; Tong, Aijun; Lu, Yi

    2009-01-01

    An abasic site called dSpacer has been introduced into duplex regions of the 8–17 DNAzyme and adenosine aptamer for label-free fluorescent detection of Pb2+ and adenosine, respectively. The dSpacer can bind an extrinsic fluorescent compound, 2-amino-5,6,7-trimethyl-1,8-naphthyridine (ATMND), and quench its fluorescence. Addition of Pb2+ enables the DNAzyme to cleave its substrate and release ATMND from DNA duplex, recovering the fluorescence of ATMND. Similarly, the presence of adenosine induces structural switching of the aptamer, resulting in the release of ATMND from the DNA duplex and a subsequent fluorescence enhancement. Under optimized conditions, this label-free method exhibits detection limits of 4 nM for Pb2+ and 3.4 µM for adenosine, which are even lower than those of the corresponding labeled-DNAzyme and aptamer sensors. These low detection limits have been obtained without compromising any of the selectivity of the sensors. Finally, the dynamic range of the adenosine sensor has been tuned by varying the number of hybridized base-pairs in the aptamer duplex. The method demonstrated here can be applied for label-free detection and quantification of a broad range of analytes using other DNAzymes and aptamers. PMID:19807110

  17. Poly(glycidyl methacrylate-co-N-methylolacrylamide-co-ethylene dimethacrylate) monolith coupled to high-performance liquid chromatography for the determination of adenosine phosphates in royal jelly.

    PubMed

    Liu, Dan; Zhang, Tianbin; Cheng, Yechun; Jia, Qiong

    2014-07-01

    A polymer monolith microextraction method coupled with high-performance liquid chromatography was developed for the determination of adenosine triphosphate, adenosine diphosphate, and adenosine monophosphate. The monolithic column was synthesized inside fused-silica capillaries using thermal initiation free-radical polymerization with glycidyl methacrylate as the monomer, ethylene dimethacrylate as the cross-linker, cyclohexanol, and 1-dodecanol as the porogen. N-Methylolacrylamide, an important hydrophilic monomer, was incorporated into the polymerization mixture to enhance the hydrophilicity of the poly(glycidyl methacrylate-co-ethylene dimethacrylate) column. The obtained poly(glycidyl methacrylate-co-N-methylolacrylamide-co-ethylene dimethacrylate) monolith was characterized by scanning electron microscopy, Fourier-transform infrared spectra, and X-ray photoelectron spectroscopy. Optimum conditions for the preconcentration and separation of the target adenosines were also investigated. Under the optimum conditions, we obtained acceptable linearities, low limits of detection, and good relative standard deviations. The developed polymer monolith microextraction with high-performance liquid chromatography method exhibited a good performance with recovery values in the range of 76.9-104.7% when applied to the determination of the adenosines in five royal jelly samples.

  18. Identification and partial characterization of an adenosine(5')tetraphospho(5')adenosine hydrolase on intact bovine aortic endothelial cells.

    PubMed Central

    Ogilvie, A; Lüthje, J; Pohl, U; Busse, R

    1989-01-01

    The biologically active dinucleotides adenosine(5')tetraphospho(5')adenosine (Ap4A) and adenosine(5')-triphospho(5')adenosine (Ap3A), which are both releasable into the circulation from storage pools in thrombocytes, are catabolized by intact bovine aortic endothelial cells. 1. Compared with extracellular ATP and ADP, which are very rapidly hydrolysed, the degradation of Ap4A and Ap3A by endothelial ectohydrolases is relatively slow, resulting in a much longer half-life on the endothelial surface of the blood vessel. The products of hydrolysis are further degraded and finally taken up as adenosine. 2. Ap4A hydrolase has high affinity for its substrate (Km 10 microM). 3. ATP as well as AMP transiently accumulates in the extracellular fluid, suggesting an asymmetric split of Ap4A by the ectoenzyme. 4. Mg2+ or Mn2+ at millimolar concentration are needed for maximal activity; Zn2+ and Ca2+ are inhibitory. 5. The hydrolysis of Ap4A is retarded by other nucleotides, such as ATP and Ap3A, which are released from platelets simultaneously with Ap4A. PMID:2541689

  19. Effect of adenosine and inosine on ureagenesis in hepatocytes.

    PubMed Central

    Guinzberg, R; Laguna, I; Zentella, A; Guzman, R; Piña, E

    1987-01-01

    Adenosine and inosine produced a dose-dependent stimulation of ureagenesis in isolated rat hepatocytes. Hypoxanthine, xanthine and uric acid were without effect. Half-maximally effective concentrations were 0.08 microM for adenosine and 5 microM for inosine. Activation of ureagenesis by both nucleosides had the following characteristics: (a) it was observed with either glutamine or (NH4)2CO3, provided that glucose was present; (b) it was not detected when glucose was replaced by lactate plus oleate; (c) it was mutually antagonized by glucagon, but not by adrenaline; and (d) it was dependent on Ca2+. We suggest that the action of adenosine and inosine on ureagenesis might be of physiological significance. PMID:3663162

  20. Release of Adenosine and ATP During Ischemia and Epilepsy

    PubMed Central

    Dale, Nicholas; Frenguelli, Bruno G

    2009-01-01

    Eighty years ago Drury & Szent-Györgyi described the actions of adenosine, AMP (adenylic acid) and ATP (pyrophosphoric or diphosphoric ester of adenylic acid) on the mammalian cardiovascular system, skeletal muscle, intestinal and urinary systems. Since then considerable insight has been gleaned on the means by which these compounds act, not least of which in the distinction between the two broad classes of their respective receptors, with their many subtypes, and the ensuing diversity in cellular consequences their activation invokes. These myriad actions are of course predicated on the release of the purines into the extracellular milieu, but, surprisingly, there is still considerable ambiguity as to how this occurs in various physiological and pathophysiological conditions. In this review we summarise the release of ATP and adenosine during seizures and cerebral ischemia and discuss mechanisms by which the purines adenosine and ATP may be released from cells in the CNS under these conditions. PMID:20190959

  1. Correlation between blood adenosine metabolism and sleep in humans.

    PubMed

    Díaz-Muñoz, M; Hernández-Muñoz, R; Suárez, J; Vidrio, S; Yááñez, L; Aguilar-Roblero, R; Rosenthal, L; Villalobos, L; Fernández-Cancino, F; Drucker-Colín, R; Chagoya De Sanchez, V

    1999-01-01

    Blood adenosine metabolism, including metabolites and metabolizing enzymes, was studied during the sleep period in human volunteers. Searching for significant correlations among biochemical parameters found: adenosine with state 1 of slow-wave sleep (SWS); activity of 5'-nucleotidase with state 2 of SWS; inosine and AMP with state 3-4 of SWS; and activity of 5'-nucleotidase and lactate with REM sleep. The correlations were detected in all of the subjects that presented normal hypnograms, but not in those who had fragmented sleep the night of the experiment. The data demonstrate that it is possible to obtain information of complex brain operations such as sleep by measuring biochemical parameters in blood. The results strengthen the notion of a role played by adenosine, its metabolites and metabolizing enzymes, during each of the stages that constitute the sleep process in humans.

  2. Targeting adenosine receptors to prevent inflammatory skin diseases.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Borea, Pier Andrea

    2014-08-01

    Adenosine mediates its effects through activation of a family of four G-protein-coupled receptors, named A1 , A2A , A2B and A3 . This nucleoside plays an important role in immunity and inflammation, and the A2A adenosine receptor subtype has a key role in the inhibition of inflammatory processes besides promoting wound healing. In this issue of Experimental Dermatology, Arasa et al. show that the topical application of a selective A2A agonist, CGS 21680, to mouse skin reduced epidermal hyperplasia as well as skin inflammation, similarly to topical corticoids, without side effects like skin atrophy. Rigorously following up this work is important for the development of novel treatment strategies for chronic hyperproliferative inflammatory dermatoses, such as targeting the A2A adenosine receptor family.

  3. Demonstration of adenosine deaminase activity in human fibroblast lysosomes.

    PubMed Central

    Lindley, E R; Pisoni, R L

    1993-01-01

    Human fibroblast lysosomes, purified on Percoll density gradients, contain an adenosine deaminase (ADA) activity that accounts for approximately 10% of the total ADA activity in GM0010A human fibroblasts. In assays of lysosomal ADA, the conversion of [3H]adenosine into [3H]inosine was proportional to incubation time and the amount of lysosomal material added to reaction mixtures. Maximal activity was observed between pH 7 and 8, and lysosomal ADA displayed a Km of 37 microM for adenosine at 25 degrees C and pH 5.5. Lysosomal ADA was completely inhibited by 2.5 mM Cu2+ or Hg2+ salts, but not by other bivalent cations (Ba2+, Cd2+, Ca2+, Fe2+, Mg2+, Mn2+ and Zn2+). Coformycin (2.5 mM), deoxycoformycin (0.02 mM), 2'-deoxyadenosine (2.5 mM), 6-methylaminopurine riboside (2.5 mM), 2'-3'-isopropylidene-adenosine (2.5 mM) and erythro-9-(2-hydroxy-3-nonyl)adenine (0.2 mM) inhibited lysosomal ADA by > 97%. In contrast, 2.5 mM S-adenosyl-L-homocysteine and cytosine were poor inhibitors. Nearly all lysosomal ADA activity is eluted as a high-molecular-mass protein (> 200 kDa) just after the void volume on a Sephacryl S-200 column, and is very heat-stable, retaining 70% of its activity after incubation at 65 degrees C for 80 min. We speculate that compartmentalization of ADA within lysosomes would allow deamination of adenosine to occur without competition by adenosine kinase, which could assist in maintaining cellular energy requirements under conditions of nutritional deprivation. PMID:8452534

  4. 8-Amino-adenosine Inhibits Multiple Mechanisms of Transcription

    PubMed Central

    Frey, Jennifer Ann; Gandhi, Varsha

    2009-01-01

    Roscovitine and flavopiridol suppress CDK7 and CDK9 activity resulting in transcription inhibition, thus providing an alternative mechanism to traditional genotoxic chemotherapy. These agents have been effective in slow or non-replicative cell types. 8-Amino-adenosine is a transcription inhibitor which has proven very effective in multiple myeloma cell lines and primary indolent leukemia cells. The objective of the current work was to define mechanisms of action that lead to transcription inhibition by 8-amino-adenosine. 8-Amino-adenosine is metabolized into the active triphosphate (8-amino-ATP) in cells. This accumulation resulted in a simultaneous decrease of intracellular ATP and RNA synthesis. When the effects of established ATP synthesis inhibitors and transcription inhibitors on intracellular ATP concentrations and RNA synthesis were studied, there was a strong correlation between ATP decline and RNA synthesis. This correlation substantiated the hypothesis that the loss of ATP in 8-amino-adenosine treated cells contributes to the decrease in transcription due to the lack of substrate needed for mRNA body and poly(A) tail synthesis. RNA polymerase II C-terminal domain phosphorylation declined sharply in 8-amino-adenosine treated cells which may have been due to the lack of an ATP phosphate donor or competitive inhibition with 8-amino-ATP at CDK7 and CDK9. Furthermore, 8-amino-ATP was incorporated into nascent RNA in a dose dependent manner at the 3′-end resulting in transcription termination. Finally, in vitro transcription assays demonstrated that 8-amino-ATP competes with ATP for incorporation into mRNA. Collectively, we have concluded that 8-amino-adenosine elicits effects on multiple mechanisms of transcription, providing a new class of transcription inhibitors. PMID:20053765

  5. Stability of Diluted Adenosine Solutions in Polyolefin Infusion Bags

    PubMed Central

    Almagambetova, Elise; Hutchinson, David; Blais, Danielle M.; Zhao, Fang

    2013-01-01

    Background: Intravenous or intracoronary adenosine is used in the cardiac catherization lab to achieve maximal coronary blood flow and determine fractional flow reserve. Objective: To determine the stability of adenosine 10 and 50 µg/mL in either 0.9% sodium chloride injection or 5% dextrose injection in polyolefin infusion bags stored at 2 temperatures, refrigeration (2°C-8°C) or controlled room temperature (20°C-25°C). Methods: Adenosine 10 µg/mL and 50 µg/mL solutions were prepared in 50 mL polyolefin infusion bags containing 0.9% sodium chloride injection or 5% dextrose injection and stored at controlled room temperature or under refrigeration. Each combination of concentration, diluent, and storage was prepared in triplicate. Samples were assayed using stability-indicating, reversed-phase high-performance liquid chromatography immediately at time 0 and at 24 hours, 48 hours, 7 days, and 14 days. Stability was defined as retaining 90% to 110% of the initial adenosine concentration. The samples were also visually inspected against a light background for clarity, color, and the presence of particulate matter. Results: After 14 days, all samples retained 99% to 101% of the initial adenosine concentration. No considerable change in pH or visual appearance was noted. The stability data indicated no significant loss of drug due to chemical degradation or physical interactions during storage. Conclusion: Adenosine solutions of 10 and 50 µg/mL were stable for at least 14 days in 50 mL polyolefin infusion bags of 0.9% sodium chloride injection or 5% dextrose injection stored at controlled room temperature and refrigerated conditions. PMID:24421510

  6. Role of adenosine as adjunctive therapy in acute myocardial infarction.

    PubMed

    Forman, Mervyn B; Stone, Gregg W; Jackson, Edwin K

    2006-01-01

    Although early reperfusion and maintained patency is the mainstay therapy for ST elevation myocardial infarction, experimental studies demonstrate that reperfusion per se induces deleterious effects on viable ischemic cells. Thus "myocardial reperfusion injury" may compromise the full potential of reperfusion therapy and may account for unfavorable outcomes in high-risk patients. Although the mechanisms of reperfusion injury are complex and multifactorial, neutrophil-mediated microvascular injury resulting in a progressive decrease in blood flow ("no-reflow" phenomenon) likely plays an important role. Adenosine is an endogenous nucleoside found in large quantities in myocardial and endothelial cells. It activates four well-characterized receptors producing various physiological effects that attenuate many of the proposed mechanisms of reperfusion injury. The cardio-protective effects of adenosine are supported by its role as a mediator of pre- and post-conditioning. In experimental models, administration of adenosine in the peri-reperfusion period results in a marked reduction in infarct size and improvement in ventricular function. The cardioprotective effects in the canine model have a narrow time window with the drug losing its effect following three hours of ischemia. Several small clinical studies have demonstrated that administration of adenosine with reperfusion therapy reduces infarct size and improves ventricular function. In the larger AMISTAD and AMISTAD II trials a 3-h infusion of adenosine as an adjunct to reperfusion resulted in a striking reduction in infarct size (55-65%). Post hoc analysis of AMISTAD II showed that this was associated with significantly improved early and late mortality in patients treated within 3.17 h of symptoms. An intravenous infusion of adenosine for 3 h should be considered as adjunctive therapy in high risk-patients undergoing reperfusion therapy.

  7. Investigating real-time activation of adenosine receptors by bioluminescence resonance energy transfer technique

    NASA Astrophysics Data System (ADS)

    Huang, Yimei; Yang, Hongqin; Zheng, Liqin; Chen, Jiangxu; Wang, Yuhua; Li, Hui; Xie, Shusen

    2013-02-01

    Adenosine receptors play important roles in many physiological and pathological processes, for example regulating myocardial oxygen consumption and the release of neurotransmitters. The activations of adenosine receptors have been studied by some kinds of techniques, such as western blot, immunohistochemistry, etc. However, these techniques cannot reveal the dynamical response of adenosine receptors under stimulation. In this paper, bioluminescence resonance energy transfer technique was introduced to study the real-time activation of adenosine receptors by monitoring the dynamics of cyclic adenosine monophosphate (cAMP) level. The results showed that there were significant differences between adenosine receptors on real-time responses under stimulation. Moreover, the dynamics of cAMP level demonstrated that competition between adenosine receptors existed. Taken together, our study indicates that monitoring the dynamics of cAMP level using bioluminescence resonance energy transfer technique could be one potential approach to investigate the mechanism of competitions between adenosine receptors.

  8. Inert Reassessment Document for Adenosine - CAS No. 58-61-7

    EPA Pesticide Factsheets

    Adenosine is classified as a 4B inert ingredient. Based on the reasonable certainty of no harm safety finding and the existing 40 CFR 180.920 use limiation, the List 4B classification for adenosine is affirmed.

  9. Biologic implications of extracellular adenosine in hepatic ischemia and reperfusion injury

    PubMed Central

    Zimmerman, Michael A.; Kam, Igal; Eltzschig, Holger; Grenz, Almut

    2013-01-01

    The purine nucleoside adenosine is clinically employed in the treatment of supraventricular tachycardia. In addition, it has direct coronary vasodilatory effects, and may influence platelet aggregation. Experimental observations mechanistically link extracellular adenosine to cellular adaptation to hypoxia. Adenosine generation has been implicated in several pathophysiologic processes including angiogenesis, tumor defenses, and neurodegeneration. In solid organ transplantation, prolonged tissue ischemia and subsequent reperfusion injury may lead to profound graft dysfunction. Importantly, conditions of limited oxygen availability are associated with increased production of extracellular adenosine and subsequent tissue protection. Within the rapidly expanding field of adenosine biology, several enzymatic steps in adenosine production have been characterized and multiple receptor subtypes have been identified. In this review, we briefly examine the biologic steps involved in adenosine generation, and chronicle the current state of adenosine signaling in hepatic ischemia and reperfusion injury. PMID:23924168

  10. Intravenous Adenosine for Surgical Management of Penetrating Heart Wounds

    PubMed Central

    Kokotsakis, John; Hountis, Panagiotis; Antonopoulos, Nikolaos; Skouteli, Elian; Athanasiou, Thanos; Lioulias, Achilleas

    2007-01-01

    Accurate suturing of penetrating cardiac injuries is difficult. Heart motion, ongoing blood loss, arrhythmias due to heart manipulation, and the near-death condition of the patient can all affect the outcome. Rapid intravenous injection of adenosine induces temporary asystole that enables placement of sutures in a motionless surgical field. Use of this technique improves surgical conditions, and it is faster than other methods. Herein, we describe our experience with the use of intravenous adenosine to successfully treat 3 patients who had penetrating heart wounds. PMID:17420798

  11. Why do premature newborn infants display elevated blood adenosine levels?

    PubMed

    Panfoli, Isabella; Cassanello, Michela; Bruschettini, Matteo; Colella, Marina; Cerone, Roberto; Ravera, Silvia; Calzia, Daniela; Candiano, Giovanni; Ramenghi, Luca

    2016-05-01

    Our preliminary data show high levels of adenosine in the blood of very low birth weight (VLBW) infants, positively correlating to their prematurity (i.e. body weight class). This prompted us to look for a mechanism promoting such impressive adenosine increase. We hypothesized a correlation with oxygen challenge. In fact, it is recognized that either oxygen lack or its excess contribute to the pathogenesis of the injuries of prematurity, such as retinopathy (ROP) and periventricular white matter lesions (PWMI). The optimal concentration of oxygen for resuscitation of VLBW infants is currently under revision. We propose that the elevated adenosine blood concentrations of VLBW infants recognizes two sources. The first could be its activity-dependent release from unmyelinated brain axons. Adenosine in this respect would be an end-product of the hypometabolic VLBW newborn unmyelinated axon intensely firing in response to the environmental stimuli consequent to premature birth. Adenosine would be eventually found in the blood due to blood-brain barrier immaturity. In fact, adenosine is the primary activity-dependent signal promoting differentiation of premyelinating oligodendrocyte progenitor cells (OPC) into myelinating cells in the Central Nervous System, while inhibiting their proliferation and inhibiting synaptic function. The second, would be the ecto-cellular ATP synthesized by the endothelial cell plasmalemma exposed to ambient oxygen concentrations due to premature breathing, especially in lung. ATP would be rapidly transformed into adenosine by the ectonucleotidase activities such as NTPDase I (CD39), and NT5E (CD73). An ectopic extra-mitochondrial aerobic ATP synthetic ability was reported in many cell plasma-membranes, among which endothelial cells. The potential implications of the cited hypotheses for the neonatology area would be great. The amount of oxygen administration for reviving of newborns would find a molecular basis for its assessment. VLBW

  12. Computer-assisted analysis of adenosine triphosphate data.

    PubMed

    Erkenbrecher, C W; Crabtree, S J; Stevenson, L H

    1976-09-01

    A computer program has been written to assist in the analysis of adenosine 5'-triphosphate data. The program is designed to calculate a dilution curve and to correct sample and adenosine 5'-triphosphate standard data for background and dilution effects. In addition, basic statistical parameters and estimates of biomass carbon are also calculated for each group of samples and printed in a convenient format. The versatility of the program to analyze data from both qauatic and terrestrial samples is noted as well as its potential use with various types of instrumentation and extraction techniques.

  13. Synthesis of 1,N6-etheno-2-aza-adenosine (2-aza-ε-adenosine): a new cytotoxic fluorescent nucleoside

    PubMed Central

    Tsou, K.C.; Yip, K.F.; Miller, E.E.; Lo, K.W.

    1974-01-01

    1,N6-Etheno-2-aza-adenosine was synthesized by treating 1,N6-etheno-adenosine with alkali, followed by nitrosation. The mechanism of formation of this novel nucleoside was elucidated using adenosine tritiated at C-8 and C-2, and was found to deformylate exclusively at C-2. This new 2-aza nucleoside fluoresces at 494 nm when excited at 358 nm. Toxicity study showed the compound is active in a rat mammary tumor tissue culture line, but inactive in HeLa and Glioma 26 tissue culture lines. It was also found to selectively inhibit the thymidine incorporation into DNA in a rat mammary tumor, but exhibits no ill effect on normal proliferative tissue. The reactive intermediate 3-β-D-ribofuranosyl-4-amino-5-(imidazol-2-yl) imidazole was identified and was found to be an active agent in tissue culture. PMID:10793738

  14. Phosphorylation of adenosine in renal brush-border membrane vesicles by an exchange reaction catalysed by adenosine kinase.

    PubMed Central

    Sayós, J; Solsona, C; Mallol, J; Lluis, C; Franco, R

    1994-01-01

    Uptake of [3H]adenosine in brush-border membrane (BBM) vesicles from either rat or pig kidney leads to an accumulation of intravesicular [3H]AMP. The lack of significant levels of ATP and the presence of AMP in BBM indicated that a phosphotransfer between [3H]adenosine and AMP occurs. The phosphotransfer activity is inhibited by iodotubercidin, which suggests that it is performed by adenosine kinase acting in an ATP-independent manner. The existence of a similar phosphotransferase activity was demonstrated in membrane-free extracts from pig kidney. From the compounds tested it was shown that a variety of mononucleotides could act as phosphate donors. The results suggest that phosphotransfer reactions may be physiologically relevant in kidney. PMID:8110185

  15. Development of Coronary Vasospasm during Adenosine-Stress Myocardial Perfusion CT Imaging.

    PubMed

    Nam, Jeong Gu; Choi, Seong Hoon; Kang, Byeong Seong; Bang, Min Seo; Kwon, Woon Jeong

    2015-01-01

    Adenosine is a short-acting coronary vasodilator, and it is widely used during pharmacological stress myocardial perfusion imaging. It has a well-established safety profile, and most of its side effects are known to be mild and transient. Until now, coronary vasospasm has been rarely reported as a side effect of adenosine during or after adenosine stress test. This study reports a case of coronary vasospasm which was documented on stress myocardial perfusion CT imaging during adenosine stress test.

  16. A2B Adenosine Receptor Agonist Improves Erectile Function in Diabetic Rats.

    PubMed

    Wen, Jiaming; Wang, Bohan; Du, Chuanjun; Xu, Gang; Zhang, Zhewei; Li, Yi; Zhang, Nan

    2015-10-01

    Diabetes is an important risk factor for erectile dysfunction (ED). Recent studies have indicated that A2B adenosine receptor (ADORA2B) signaling is essential for penile erection. Thus, we hypothesize that diabetic ED may be attributed to impaired A2B adenosine signaling. To test this hypothesis, we generated diabetic rats by injecting streptozocin as animal model. After 12 weeks, immunohistochemistry staining was used to localize the expression of ADORA2B. Western Blot and quantitative PCR were employed to determine ADORA2B expression level. Intracavernosal pressure (ICP) measurement was used to evaluate erectile function. Diabetic rats received a single intravenous injection of BAY 60-6583, an ADORA2B agonist, or vehicle solution, at 60 min before the ICP measurement. The results showed that ADORA2B expressed in the nerve bundle, smooth muscle, and endothelium in penile tissue of control mice. Western Blot and quantitative PCR results indicated that the expression levels of ADORA2B protein and mRNA were significantly reduced in penile tissues of diabetic rats. Functional studies showed that the erectile response induced by electrical stimulation was remarkably decreased in diabetic rats, compared with age-matched control rats. However, at 60 min after BAY 60-6583 treatment, the erectile function was improved in diabetic rats, suggesting that enhancement of ADORA2B signaling may improve erectile function in diabetic ED. This preclinical study has revealed a previously unrecognized therapeutic possibility of BAY 60-6583 as an effective and mechanism-based drug to treat diabetic ED. In conclusion, we propose that impaired A2B adenosine signaling is one of the pathological mechanisms of diabetic ED.

  17. Adenosine-dependent activation of tyrosine hydroxylase is defective in adenosine kinase-deficient PC12 cells.

    PubMed Central

    Erny, R; Wagner, J A

    1984-01-01

    (R)-N6-Phenylisopropyladenosine (PIA) stimulates dopa production 3- to 5-fold in PC12 cells, with a half-maximal effective concentration (EC50) of 50 nM. This increase can be explained by a stable activation of tyrosine hydroxylase [TyrOHase; L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] when it is phosphorylated by a cAMP-dependent protein kinase. The activation of TyrOHase is mediated by the adenosine-dependent activation of adenylate cyclase (EC50 = 600 nM). PIA (10 microM) is as effective as cholera toxin or dibutyryl cAMP in activating TyrOHase in wild-type cells. Adenosine kinase-deficient mutants of PC12 were found to be resistant to PIA-dependent activation of TyrOHase (EC50 = 100-1000 nM). This phenomenon was explored in detail in one adenosine kinase-deficient mutant and was shown to occur because the mutant was resistant to the adenosine-dependent activation of adenylate cyclase. In this mutant, TyrOHase was activated 14-fold by cholera toxin, suggesting that activated TyrOHase is about 14 times as active as unactivated TyrOHase. These studies with kinase-deficient PC12 cells provide genetic evidence that adenosine-dependent activation of TyrOHase is mediated by acute increases in cAMP. When the adenosine receptor found on PC12 cells is expressed in vivo, it might function as either a presynaptic (i.e., localized on the nerve terminal) or a postsynaptic (i.e., localized on the cell body or dendrite) receptor that regulates rates of transmitter synthesis in response to cell activity. PMID:6146982

  18. In vivo assessment of coronary flow and cardiac function after bolus adenosine injection in adenosine receptor knockout mice.

    PubMed

    Teng, Bunyen; Tilley, Stephen L; Ledent, Catherine; Mustafa, S Jamal

    2016-06-01

    Bolus injections of adenosine and the A2A adenosine receptor (AR) selective agonist (regadenoson) are used clinically as a substitute for a stress test in people who cannot exercise. Using isolated tissue preparations, our lab has shown that coronary flow and cardiac effects of adenosine are mostly regulated by the AR subtypes A1, A2A, and A2B In this study, we used ultrasound imaging to measure the in vivo effects of adenosine on coronary blood flow (left coronary artery) and cardiac function in anesthetized wild-type, A1 knockout (KO), A2AKO, A2BKO, A3KO, A1, and A3 double KO (A1/3 DKO) and A2A and A2B double KO (A2A/2B DKO) mice in real time. Echocardiographic and Doppler studies were performed using a Visualsonic Vevo 2100 ultrasound system. Coronary blood flow (CBF) baseline data were obtained when animals were anesthetized with 1% isoflourane. Diameter (D) and velocity time integral (VTI) were measured on the left coronary arteries (CBF = ((π/4) × D(2) × VTI × HR)/1000). CBF changes were the highest within 2 min of injection (about 10 mg/kg). Heart rate, cardiac output, and stroke volume were measured by tracing the left ventricle long axis. Our data support a role for the A2 AR in CBF and further support our conclusions of previous studies from isolated tissues. Adenosine-mediated decreases in cardiac output and stroke volume may be A2B and/or A3 AR-mediated; however, the A1 and A2 ARs also play roles in overall cardiac function. These data further provide a powerful translational tool in studying the cardiovascular effects of adenosine in disease states.

  19. Elevated Adenosine Induces Placental DNA Hypomethylation Independent of A2B Receptor Signaling in Preeclampsia.

    PubMed

    Huang, Aji; Wu, Hongyu; Iriyama, Takayuki; Zhang, Yujin; Sun, Kaiqi; Song, Anren; Liu, Hong; Peng, Zhangzhe; Tang, Lili; Lee, Minjung; Huang, Yun; Ni, Xin; Kellems, Rodney E; Xia, Yang

    2017-07-01

    Preeclampsia is a prevalent pregnancy hypertensive disease with both maternal and fetal morbidity and mortality. Emerging evidence indicates that global placental DNA hypomethylation is observed in patients with preeclampsia and is linked to altered gene expression and disease development. However, the molecular basis underlying placental epigenetic changes in preeclampsia remains unclear. Using 2 independent experimental models of preeclampsia, adenosine deaminase-deficient mice and a pathogenic autoantibody-induced mouse model of preeclampsia, we demonstrate that elevated placental adenosine not only induces hallmark features of preeclampsia but also causes placental DNA hypomethylation. The use of genetic approaches to express an adenosine deaminase minigene specifically in placentas, or adenosine deaminase enzyme replacement therapy, restored placental adenosine to normal levels, attenuated preeclampsia features, and abolished placental DNA hypomethylation in adenosine deaminase-deficient mice. Genetic deletion of CD73 (an ectonucleotidase that converts AMP to adenosine) prevented the elevation of placental adenosine in the autoantibody-induced preeclampsia mouse model and ameliorated preeclampsia features and placental DNA hypomethylation. Immunohistochemical studies revealed that elevated placental adenosine-mediated DNA hypomethylation predominantly occurs in spongiotrophoblasts and labyrinthine trophoblasts and that this effect is independent of A2B adenosine receptor activation in both preeclampsia models. Extending our mouse findings to humans, we used cultured human trophoblasts to demonstrate that adenosine functions intracellularly and induces DNA hypomethylation without A2B adenosine receptor activation. Altogether, both mouse and human studies reveal novel mechanisms underlying placental DNA hypomethylation and potential therapeutic approaches for preeclampsia. © 2017 American Heart Association, Inc.

  20. Neuronal transporter and astrocytic ATP exocytosis underlie activity-dependent adenosine release in the hippocampus

    PubMed Central

    Wall, Mark J; Dale, Nicholas

    2013-01-01

    The neuromodulator adenosine plays an important role in many physiological and pathological processes within the mammalian CNS. However, the precise mechanisms of how the concentration of extracellular adenosine increases following neural activity remain contentious. Here we have used microelectrode biosensors to directly measure adenosine release induced by focal stimulation in stratum radiatum of area CA1 in mouse hippocampal slices. Adenosine release was both action potential and Ca2+ dependent and could be evoked with low stimulation frequencies and small numbers of stimuli. Adenosine release required the activation of ionotropic glutamate receptors and could be evoked by local application of glutamate receptor agonists. Approximately 40% of stimulated-adenosine release occurred by translocation of adenosine via equilibrative nucleoside transporters (ENTs). This component of release persisted in the presence of the gliotoxin fluoroacetate and thus results from the direct release of adenosine from neurons. A reduction of adenosine release in the presence of NTPDase blockers, in slices from CD73−/− and dn-SNARE mice, provides evidence that a component of adenosine release arises from the extracellular metabolism of ATP released from astrocytes. This component of release appeared to have slower kinetics than the direct ENT-mediated release of adenosine. These data suggest that activity-dependent adenosine release is surprisingly complex and, in the hippocampus, arises from at least two distinct mechanisms with different cellular sources. PMID:23713028

  1. Anticancer effect of adenosine on gastric cancer via diverse signaling pathways.

    PubMed

    Tsuchiya, Ayako; Nishizaki, Tomoyuki

    2015-10-21

    Extracellular adenosine induces apoptosis in a variety of cancer cells via intrinsic and extrinsic pathways. In the former pathway, adenosine uptake into cells triggers apoptosis, and in the latter pathway, adenosine receptors mediate apoptosis. Extracellular adenosine also induces apoptosis of gastric cancer cells. Extracellular adenosine is transported into cells through an adenosine transporter and converted to AMP by adenosine kinase. In turn, AMP activates AMP-activated protein kinase (AMPK). AMPK is the factor responsible for caspase-independent apoptosis of GT3-TKB gastric cancer cells. Extracellular adenosine, on the other hand, induces caspase-dependent apoptosis of MKN28 and MKN45 gastric cancer cells by two mechanisms. Firstly, AMP, converted from intracellularly transported adenosine, initiates apoptosis, regardless of AMPK. Secondly, the A3 adenosine receptor, linked to Gi/Gq proteins, mediates apoptosis by activating the Gq protein effector, phospholipase Cγ, to produce inositol 1,4,5-trisphosphate and diacylglycerol, which activate protein kinase C. Consequently, the mechanisms underlying adenosine-induced apoptosis vary, depending upon gastric cancer cell types. Understand the contribution of each downstream target molecule of adenosine to apoptosis induction may aid the establishment of tailor-made chemotherapy for gastric cancer.

  2. [The mechanisms of depression by benzodiazepines, barbiturates and propofol of excitatory synaptic transmissions mediated by adenosine neuromodulation].

    PubMed

    Narimatsu, Eichi; Niiya, Tomohisa; Kawamata, Mikito; Namiki, Akiyoshi

    2006-06-01

    Adenosine is known to modulate synaptic functions through adenosine receptors and the related adenosine neuromodulatory system. Benzodiazepines, barbiturates and propofol, the main actins of which are facilitation of GABAA receptor functions, also facilitate accumulation of endogenous adenosine in the extracellular space by inhibiting adenosine uptake via adenosine transporters. The accumulated adenosine depresses excitatory synaptic transmission by decreasing transmitter release, depressing postsynaptic sensitivity and inhibiting neuronal excitability through adenosine A1 receptors and the related adenosine neuromodulatory system. The adenosine-induced depressions of excitatory synaptic transmissions probably contribute to the mechanisms of the anesthetic, sedative, anticonvulsant and neuroprotective effects of these anesthetics and sedatives. An understanding of the synaptic mechanisms of these anesthetics and sedatives through the adenosine neuromodulatory system is needed for rational clinical use of the drugs.

  3. Purification and properties of adenylyl sulphate:ammonia adenylyltransferase from Chlorella catalysing the formation of adenosine 5' -phosphoramidate from adenosine 5' -phosphosulphate and ammonia.

    PubMed

    Fankhauser, H; Schiff, J A; Garber, L J

    1981-06-01

    Extracts of Chlorella pyrenoidosa, Euglena gracilis var. bacillaris, spinach, barley, Dictyostelium discoideum and Escherichia coli form an unknown compound enzymically from adenosine 5'-phosphosulphate in the presence of ammonia. This unknown compound shares the following properties with adenosine 5'-phosphoramidate: molar proportions of constituent parts (1 adenine:1 ribose:1 phosphate:1 ammonia released at low pH), co-electrophoresis in all buffers tested including borate, formation of AMP at low pH through release of ammonia, mass and i.r. spectra and conversion into 5'-AMP by phosphodiesterase. This unknown compound therefore appears to be identical with adenosine 5'-phosphoramidate. The enzyme that catalyses the formation of adenosine 5'-phosphoramidate from ammonia and adenosine 5'-phosphosulphate was purified 1800-fold (to homogeneity) from Chlorella by using (NH(4))(2)SO(4) precipitation and DEAE-cellulose, Sephadex and Reactive Blue 2-agarose chromatography. The purified enzyme shows one band of protein, coincident with activity, at a position corresponding to 60000-65000 molecular weight, on polyacrylamide-gel electrophoresis, and yields three subunits on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of 26000, 21000 and 17000 molecular weight, consistent with a molecular weight of 64000 for the native enzyme. Isoelectrofocusing yields one band of pI4.2. The pH optimum of the enzyme-catalysed reaction is 8.8. ATP, ADP or adenosine 3'-phosphate 5'-phosphosulphate will not replace adenosine 5'-phosphosulphate, and the apparent K(m) for the last-mentioned compound is 0.82mm. The apparent K(m) for ammonia (assuming NH(3) to be the active species) is about 10mm. A large variety of primary, secondary and tertiary amines or amides will not replace ammonia. One mol.prop. of adenosine 5'-phosphosulphate reacts with 1 mol.prop. of ammonia to yield 1 mol.prop. each of adenosine 5'-phosphoramidate and sulphate; no AMP is found. The highly purified enzyme

  4. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages §...

  5. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7040...

  6. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7040...

  7. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7040...

  8. The Role of Adenosine in Alzheimer’s Disease

    PubMed Central

    Rahman, Anisur

    2009-01-01

    Alzheimer’s disease (AD) is a neurodegenerative disorder of the central nervous system manifested by cognitive and memory deterioration, a variety of neuropsychiatric symptoms, behavioral disturbances, and progressive impairment of daily life activities. Current pharmacotherapies are restricted to symptomatic interventions but do not prevent progressive neuronal degeneration. Therefore, new therapeutic strategies are needed to intervene with these progressive pathological processes. In the past several years adenosine, a ubiquitously released purine ribonucleoside, has become important for its neuromodulating capability and its emerging positive experimental effects in neurodegenerative diseases. Recent research suggests that adenosine receptors play important roles in the modulation of cognitive function. The present paper attempts to review published reports and data from different studies showing the evidence of a relationship between adenosinergic function and AD-related cognitive deficits. Epidemiological studies have found an association between coffee (a nonselective adenosine receptor antagonist) consumption and improved cognitive function in AD patients and in the elderly. Long-term administration of caffeine in transgenic animal models showed a reduced amyloid burden in brain with better cognitive performance. Antagonists of adenosine A2A receptors mimic these beneficial effects of caffeine on cognitive function. Neuronal cell cultures with amyloid beta in the presence of an A2A receptor antagonist completely prevented amyloid beta-induced neurotoxicity. These findings suggest that the adenosinergic system constitutes a new therapeutic target for AD, and caffeine and A2A receptor antagonists may have promise to manage cognitive dysfunction in AD. PMID:20190962

  9. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... device that measures the release of adenosine triphosphate (ATP) from platelets following aggregation. This measurement is made on platelet-rich plasma using a photometer and a luminescent firefly extract. Simultaneous measurements of platelet aggregation and ATP release are used to evaluate platelet...

  10. Laser photobleaching leads to a fluorescence grade adenosine deaminase.

    PubMed

    Parola, A H; Caiolfa, V R; Bar, I; Rosenwaks, S

    1989-09-01

    The enzyme adenosine deaminase (adenosine aminohydrolase EC 3.5.4.4) from calf intestinal mucosa is commercially available at high purity grade yet, at the sensitivity at which fluorescence studies may be undertaken, a nonpeptidic fluorescence is detectable at lambda exmax = 350 nm and lambda emmax = 420 nm. A sevenfold decrease of this nonpeptidic fluorescence was obtained upon irradiation by the third harmonic (355 nm) of a Nd:YAG laser for 16 min, at 5 mJ/pulse, with a pulse width of 6 ns at a repetition rate of 10 Hz. The decline of fluorescence was accompanied by a negligible loss of enzymatic activity. Moreover, the integrity of the protein was ascertained by (i) its fluorescence (lambda exmax = 305 nm, lambda emmax = 335 nm) and lifetime distribution and (ii) its kinetics in the presence of the substrate adenosine and two inhibitors, all of which remained essentially unaltered. Laser photobleaching is a simple way to achieve a fluorescence grade adenosine deaminase.

  11. Adenosine receptor blockade reduces splanchnic hyperemia in cirrhotic rats.

    PubMed

    Lee, S S; Chilton, E L; Pak, J M

    1992-06-01

    To explore a possible role for adenosine in the pathogenesis of the splanchnic hyperemia of cirrhosis, we administered 8-phenyltheophylline, a specific adenosine receptor antagonist, to rats with biliary cirrhosis caused by bile duct ligation and to control sham-operated rats. Micro-Doppler flow studies showed that a 10-mumol/kg dose of 8-phenyltheophylline completely abolished the superior mesenteric hyperemic response to infusions of exogenous adenosine in both cirrhotic and control rats. Analysis of regional blood flows by radioactive microspheres demonstrated that this dose of 8-phenyltheophylline in cirrhotic rats significantly increased portal tributary vascular resistance by 60% and decreased portal tributary blood flow by 26%. This decrease was entirely the result of a 42% reduction in the intestinal blood flow. 8-phenyltheophylline did not affect cardiac output, arterial pressure or any other extrasplanchnic hemodynamic variables in cirrhotic rats. No detectable effect of 8-phenyltheophylline was seen in sham-operated rats. These results suggest that adenosine may be involved in the genesis of splanchnic hyperemia in cirrhotic rats.

  12. Adenosine receptor modulation of seizure susceptibility in rats

    SciTech Connect

    Szot, P.

    1987-01-01

    Adenosine is considered to be a neuromodulator or cotransmitter in the periphery and CNS. This neuromodulatory action of adenosine may be observed as an anticonvulsant effect. Dose-response curves for R-phenylisopropyladenosine (PIA), cycohexyladenosine (CHA), 2-chloroadenosine (2-ClAdo), N-ethylcarboxamidoadenosine (NECA) and S-PIA were generated against PTZ seizure thresholds in the rat. The rank order of potency for adenosine agonists to elevate PTZ seizure threshold was R-PIA > 2-ClAdo > NECA > CHA > S-PIA. R-PIA was approximately 80-fold more potent than S-PIA. This 80-fold difference in potency between the diasteriomers of PIA was consistent with an A{sub 1} adenoise receptor-mediated response. The anticonvulsant action of 2-ClAdo was reversed by pretreatment with theoplylline. Chronic administration of theophylline significantly increased the specific binding of {sup 3}H-cyclohexyladenosine in membranes of the cerebral cortex and cerebellum of the rat. Chronic exposure to theophylline produced a significant increase in the densities of both the high- and low-affinity forms of A{sub 1} adenosine receptors in the cerebral cortex.

  13. Quantitative changes in adenosine deaminase isoenzymes in human colorectal adenocarcinomas.

    PubMed

    ten Kate, J; Wijnen, J T; van der Goes, R G; Quadt, R; Griffioen, G; Bosman, F T; Khan, P M

    1984-10-01

    Several reports have suggested that a decrease or absence of adenosine deaminase complexing protein (ADCP) is consistently associated with cancer. However, in other studies, decreased as well as increased ADCP levels were found. In the present study, we investigated ADCP levels in 37 colorectal adenocarcinomas and correlated the results with clinicopathological characteristics in individual carcinomas. The levels of adenosine deaminase (EC 3.5.4.4) and soluble ADCP were determined in tissue samples by, respectively, a spectrophotometric assay and an ADCP specific radioimmunoassay. The values in the individual tumors were compared with their histological characteristics, such as degree of differentiation, nuclear grading, and the preoperative plasma carcinoembryonic antigen levels in the patients. It was found that ADCP was decreased in about a third of the tumors but unaltered or even increased in others. However, there was an overall 40% increase of the adenosine deaminase activity in the tumors compared to normal tissue. There seems to be no simple correlation between any of the clinicopathological parameters and the ADCP or adenosine deaminase levels. Methods detecting ADCP at single cell level might be helpful in exploring its potential use as a cancer-associated marker.

  14. Striatal adenosine-cannabinoid receptor interactions in rats over-expressing adenosine A2A receptors.

    PubMed

    Chiodi, Valentina; Ferrante, Antonella; Ferraro, Luca; Potenza, Rosa Luisa; Armida, Monica; Beggiato, Sarah; Pèzzola, Antonella; Bader, Michael; Fuxe, Kjell; Popoli, Patrizia; Domenici, Maria Rosaria

    2016-03-01

    Adenosine A2A receptors (A2 A Rs) and cannabinoid CB1 receptors (CB1 Rs) are highly expressed in the striatum, where they functionally interact and form A2A /CB1 heteroreceptor complexes. We investigated the effects of CB1 R stimulation in a transgenic rat strain over-expressing A2 A Rs under the control of the neural-specific enolase promoter (NSEA2A rats) and in age-matched wild-type (WT) animals. The effects of the CB1 R agonist WIN 55,212-2 (WIN) were significantly lower in NSEA2A rats than in WT animals, as demonstrated by i) electrophysiological recordings of synaptic transmission in corticostriatal slices; ii) the measurement of glutamate outflow from striatal synaptosomes and iii) in vivo experiments on locomotor activity. Moreover, while the effects of WIN were modulated by both A2 A R agonist (CGS 21680) and antagonists (ZM 241385, KW-6002 and SCH-442416) in WT animals, the A2 A R antagonists failed to influence WIN-mediated effects in NSEA2A rats. The present results demonstrate that in rats with genetic neuronal over-expression of A2 A Rs, the effects mediated by CB1 R activation in the striatum are significantly reduced, suggesting a change in the stoichiometry of A2A and CB1 receptors and providing a strategy to dissect the involvement of A2 A R forming or not forming heteromers in the modulation of striatal functions. These findings add additional evidence for the existence of an interaction between striatal A2 A Rs and CB1 Rs, playing a fundamental role in the regulation of striatal functions. We studied A2A -CB1 receptor interaction in transgenic rats over-expressing adenosine A2A receptors under the control of the neuron-specific enolase promoter (NSEA2A ). In these rats, we demonstrated a reduced effect of the CB1 receptor agonist WIN 55,212-2 in the modulation of corticostriatal synaptic transmission and locomotor activity, while CB1 receptor expression level did not change with respect to WT rats. A reduction in the expression of A2A -CB1

  15. Correlation of transient adenosine release and oxygen changes in the caudate-putamen.

    PubMed

    Wang, Ying; Venton, B Jill

    2017-01-01

    Adenosine is an endogenous nucleoside that modulates important physiological processes, such as vasodilation, in the central nervous system. A rapid, 2-4 s, mode of adenosine signaling has been recently discovered, but the relationship between this type of adenosine and blood flow change has not been characterized. In this study, adenosine and oxygen changes were simultaneously measured using fast-scan cyclic voltammetry. Oxygen changes occur when there is an increase in local cerebral blood flow and thus are a measure of vasodilation. About 34% of adenosine transients in the rat caudate-putamen are correlated with a subsequent transient change in oxygen. The amount of oxygen was correlated with the concentration of adenosine release and larger adenosine transients (over 0.4 μM) always had subsequent oxygen changes. The average duration of adenosine and oxygen transients was 3.2 and 3.5 s, respectively. On average, the adenosine release starts and peaks 0.2 s prior to the oxygen. The A2a antagonist, SCH442416, decreased the number of both adenosine and oxygen transient events by about 32%. However, the A1 antagonist, DPCPX, did not significantly affect simultaneous adenosine and oxygen release. The nitric oxide (NO) synthase inhibitor l-NAME also did not affect the concentration or number of adenosine and oxygen release events. These results demonstrate that both adenosine and oxygen release are modulated via A2a receptors. The correlation of transient concentrations, time delay between adenosine and oxygen peaks, and effect of A2a receptors suggests that adenosine modulates blood flow on a rapid, sub-second time scale. Read the Editorial Highlight for this article on page 10. © 2016 International Society for Neurochemistry.

  16. Prognostic value of normal adenosine-stress cardiac magnetic resonance imaging.

    PubMed

    Pilz, Guenter; Jeske, Andrea; Klos, Markus; Ali, Eman; Hoefling, Berthold; Scheck, Roland; Bernhardt, Peter

    2008-05-15

    We investigated the prognostic value of normal adenosine stress cardiac magnetic resonance (CMR) in suspected coronary artery disease (CAD). Prospectively enrolled in the study were 218 patients with suspected CAD, no stress hypoperfusion, and no delayed enhancement in CMR, and consecutively deferred coronary angiography. The primary end point was a 12-month rate of major adverse cardiac events (MACE; cardiovascular mortality, myocardial infarction, revascularization, hospitalization due to cardiovascular event). CMR indication was symptomatic angina (Canadian Cardiovascular Society II in 42% and III in 7%) or evaluation of myocardial ischemia in patients with arrhythmia, syncope, and/or equivocal stress tests and cardiovascular risk factors (51%). As the main result, the 12-month MACE rate was 2/218 (1 stent implantation, 1 bypass surgery) and CMR negative predictive value 99.1%. There was no cardiac death or myocardial infarction. In conclusion, normal adenosine stress CMR predicts a very low MACE rate and an excellent 1-year prognosis in patients with suspected CAD. Our results provide clinical reassurance that patients at risk for CAD-associated MACE were not missed by CMR. Hence, CMR may serve as a reliable noninvasive gatekeeper to reduce the number of redundant coronary angiographies.

  17. Nucleotide Binding Site Communication in Arabidopsis thaliana Adenosine 5′-Phosphosulfate Kinase*

    PubMed Central

    Ravilious, Geoffrey E.; Jez, Joseph M.

    2012-01-01

    Adenosine 5′-phosphosulfate kinase (APSK) catalyzes the ATP-dependent synthesis of adenosine 3′-phosphate 5′-phosphosulfate (PAPS), which is an essential metabolite for sulfur assimilation in prokaryotes and eukaryotes. Using APSK from Arabidopsis thaliana, we examine the energetics of nucleotide binary and ternary complex formation and probe active site features that coordinate the order of ligand addition. Calorimetric analysis shows that binding can occur first at either nucleotide site, but that initial interaction at the ATP/ADP site was favored and enhanced affinity for APS in the second site by 50-fold. The thermodynamics of the two possible binding models (i.e. ATP first versus APS first) differs and implies that active site structural changes guide the order of nucleotide addition. The ligand binding analysis also supports an earlier suggestion of intermolecular interactions in the dimeric APSK structure. Crystallographic, site-directed mutagenesis, and energetic analyses of oxyanion recognition by the P-loop in the ATP/ADP binding site and the role of Asp136, which bridges the ATP/ADP and APS/PAPS binding sites, suggest how the ordered nucleotide binding sequence and structural changes are dynamically coordinated for catalysis. PMID:22810229

  18. Nucleotide Binding Site Communication in Arabidopsis thaliana Adenosine 5;-Phosphosulfate Kinase

    SciTech Connect

    Ravilious, Geoffrey E.; Jez, Joseph M.

    2012-08-31

    Adenosine 5{prime}-phosphosulfate kinase (APSK) catalyzes the ATP-dependent synthesis of adenosine 3{prime}-phosphate 5{prime}-phosphosulfate (PAPS), which is an essential metabolite for sulfur assimilation in prokaryotes and eukaryotes. Using APSK from Arabidopsis thaliana, we examine the energetics of nucleotide binary and ternary complex formation and probe active site features that coordinate the order of ligand addition. Calorimetric analysis shows that binding can occur first at either nucleotide site, but that initial interaction at the ATP/ADP site was favored and enhanced affinity for APS in the second site by 50-fold. The thermodynamics of the two possible binding models (i.e. ATP first versus APS first) differs and implies that active site structural changes guide the order of nucleotide addition. The ligand binding analysis also supports an earlier suggestion of intermolecular interactions in the dimeric APSK structure. Crystallographic, site-directed mutagenesis, and energetic analyses of oxyanion recognition by the P-loop in the ATP/ADP binding site and the role of Asp136, which bridges the ATP/ADP and APS/PAPS binding sites, suggest how the ordered nucleotide binding sequence and structural changes are dynamically coordinated for catalysis.

  19. Adenosine Deaminases Acting on RNA (ADARs) are both Antiviral and Proviral Dependent upon the Virus

    PubMed Central

    Samuel, Charles E.

    2010-01-01

    A-to-I RNA editing, the deamination of adenosine (A) to inosine (I) that occurs in regions of RNA with double-stranded character, is catalyzed by a family of Adenosine Deaminases Acting on RNA (ADARs). In mammals there are three ADAR genes. Two encode proteins that possess demonstrated deaminase activity: ADAR1, which is interferon-inducible, and ADAR2 which is constitutively expressed. ADAR3, by contrast, has not yet been shown to bean active enzyme. The specificity of the ADAR1 and ADAR2 deaminases ranges from highly site-selective to non-selective, dependent on the duplex structure of the substrate RNA. A-to-I editing is a form of nucleotide substitution editing, because I is decoded as guanosine (G) instead of A by ribosomes during translation and by polymerases during RNA-dependent RNA replication. Additionally, A-to-I editing can alter RNA structure stability as I:U mismatches are less stable than A:U base pairs. Both viral and cellular RNAs are edited by ADARs. A-to-I editing is of broad physiologic significance. Among the outcomes of A-to-I editing are biochemical changes that affect how viruses interact with their hosts, changes that can lead to either enhanced or reduced virus growth and persistence dependent upon the specific virus. PMID:21211811

  20. Triggering neurotrophic factor actions through adenosine A2A receptor activation: implications for neuroprotection

    PubMed Central

    Sebastião, Ana M; Ribeiro, Joaquim A

    2009-01-01

    G protein coupled receptors and tropomyosin-related kinase (Trk) receptors have distinct structure and transducing mechanisms; therefore, cross-talk among them was unexpected. Evidence has, however, accumulated showing that tonic adenosine A2A receptor activity is a required step to allow synaptic actions of neurotrophic factors, namely upon synaptic transmission at both pre- and post-synaptic level as well as upon synaptic plasticity. An enhancement of A2A receptor tonus upon ageing may partially compensate the loss of TrkB receptors, rescuing to certain degree the facilitatory action of brain derived neurotrophic factor in aged animals, which might prove particularly relevant in the prevention of neurodegeneration upon ageing. A2A receptors also trigger synaptic actions of other neurotrophic factors, such as glial derived neurotrophic factor at dopaminergic striatal nerve endings. The growing evidence that tonic adenosine A2A receptor activity is a crucial step to allow actions of neurotrophic factors in neurones will be reviewed and discussed in the light of therapeutic strategies for neurodegenerative diseases. PMID:19508402

  1. Water-mediated recognition of t1-adenosine anchors Argonaute2 to microRNA targets

    PubMed Central

    Schirle, Nicole T; Sheu-Gruttadauria, Jessica; Chandradoss, Stanley D; Joo, Chirlmin; MacRae, Ian J

    2015-01-01

    MicroRNAs (miRNAs) direct post-transcriptional regulation of human genes by guiding Argonaute proteins to complementary sites in messenger RNAs (mRNAs) targeted for repression. An enigmatic feature of many conserved mammalian miRNA target sites is that an adenosine (A) nucleotide opposite miRNA nucleotide-1 confers enhanced target repression independently of base pairing potential to the miRNA. In this study, we show that human Argonaute2 (Ago2) possesses a solvated surface pocket that specifically binds adenine nucleobases in the 1 position (t1) of target RNAs. t1A nucleotides are recognized indirectly through a hydrogen-bonding network of water molecules that preferentially interacts with the N6 amine on adenine. t1A nucleotides are not utilized during the initial binding of Ago2 to its target, but instead function by increasing the dwell time on target RNA. We also show that N6 adenosine methylation blocks t1A recognition, revealing a possible mechanism for modulation of miRNA target site potency. DOI: http://dx.doi.org/10.7554/eLife.07646.001 PMID:26359634

  2. Platelet aggregation and serum adenosine deaminase (ADA) activity in pregnancy associated with diabetes, hypertension and HIV.

    PubMed

    Leal, Claudio A M; Leal, Daniela B R; Adefegha, Stephen A; Morsch, Vera M; da Silva, José E P; Rezer, João F P; Schrekker, Clarissa M L; Abdalla, Faida H; Schetinger, Maria R C

    2016-07-01

    Platelet aggregation and adenosine deaminase (ADA) activity were evaluated in pregnant women living with some disease conditions including hypertension, diabetes mellitus and human immunodeficiency virus infection. The subject population is consisted of 15 non-pregnant healthy women [control group (CG)], 15 women with normal pregnancy (NP), 7 women with hypertensive pregnancy (HP), 10 women with gestational diabetes mellitus (GDM) and 12 women with human immunodeficiency virus-infected pregnancy (HIP) groups. The aggregation of platelets was checked using an optical aggregometer, and serum ADA activity was determined using the colorimetric method. After the addition of 5 µM of agonist adenosine diphosphate, the percentage of platelet aggregation was significantly (p < 0·05) increased in NP, HP, GDM and HIP groups when compared with the CG, while the addition of 10 µM of the same agonist caused significant (p < 0·05) elevations in HP, GDM and HIP groups when compared with CG. Furthermore, ADA activity was significantly (p < 0·05) enhanced in NP, HP, GDM and HIP groups when compared with CG. In this study, the increased platelet aggregation and ADA activity in pregnancy and pregnancy-associated diseases suggest that platelet aggregation and ADA activity could serve as peripheral markers for the development of effective therapy in the maintenance of homeostasis and some inflammatory process in these pathophysiological conditions. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  3. Caffeine May Reduce Perceived Sweet Taste in Humans, Supporting Evidence That Adenosine Receptors Modulate Taste.

    PubMed

    Choo, Ezen; Picket, Benjamin; Dando, Robin

    2017-09-01

    Multiple recent reports have detailed the presence of adenosine receptors in sweet sensitive taste cells of mice. These receptors are activated by endogenous adenosine in the plasma to enhance sweet signals within the taste bud, before reporting to the primary afferent. As we commonly consume caffeine, a powerful antagonist for such receptors, in our daily lives, an intriguing question we sought to answer was whether the caffeine we habitually consume in coffee can inhibit the perception of sweet taste in humans. 107 panelists were randomly assigned to 2 groups, sampling decaffeinated coffee supplemented with either 200 mg of caffeine, about the level found in a strong cup of coffee, or an equally bitter concentration of quinine. Participants subsequently performed sensory testing, with the session repeated in the alternative condition in a second session on a separate day. Panelists rated both the sweetened coffee itself and subsequent sucrose solutions as less sweet in the caffeine condition, despite the treatment having no effect on bitter, sour, salty, or umami perception. Panelists were also unable to discern whether they had consumed the caffeinated or noncaffeinated coffee, with ratings of alertness increased equally, but no significant improvement in reaction times, highlighting coffee's powerful placebo effect. This work validates earlier observations in rodents in a human population. © 2017 Institute of Food Technologists®.

  4. Robust aptamer sol-gel solid phase microextraction of very polar adenosine from human plasma.

    PubMed

    Mu, Li; Hu, Xiangang; Wen, Jianping; Zhou, Qixing

    2013-03-01

    Conventional solid phase microextraction (SPME) has a limited capacity to extract very polar analytes, such as adenosine. To solve this problem, aptamer conjugating sol-gel methodology was coupled with an SPME fiber. According to the authors' knowledge, this is the first reported use of aptamer SPME. The fiber of aptamer sol-gel SPME with a mesoporous structure has high porosity, large surface area, and small water contact angle. Rather than employing direct entrapment, covalent immobilization was the dominant method of aptamer loading in sol-gel. Aptamer sol-gel fiber captured a specified analyte from among the analog molecules, thereby, exhibiting an excellent selective property. Compared with commercial SPME fibers, this aptamer fiber was suitable for extracting adenosine, presenting an extraction efficiency higher than 20-fold. The values of repeatability and reproducibility expressed by relative standard deviation were low (9.4%). Interestingly, the sol-gel network enhanced the resistance of aptamer SPME to both nuclease and nonspecific proteins. Furthermore, the aptamer sol-gel fiber was applied in human plasma with LOQ 1.5 μg/L, which is an acceptable level. This fiber also demonstrates durability and regeneration over 20-cycles without significant loss of efficiency. Given the various targets (from metal ions to biomacromolecules and cells) of aptamers, this methodology will extend the multi-domain applications of SPME.

  5. Adenosine induces G2/M cell-cycle arrest by inhibiting cell mitosis progression.

    PubMed

    Jia, Kun-Zhi; Tang, Bo; Yu, Lu; Cheng, Wei; Zhang, Rong; Zhang, Jian-Fa; Hua, Zi-Chun

    2009-12-16

    Cellular adenosine accumulates under stress conditions. Few papers on adenosine are concerned with its function in the cell cycle. The cell cycle is the essential mechanism by which all living things reproduce and the target machinery when cells encounter stresses, so it is necessary to examine the relationship between adenosine and the cell cycle. In the present study, adenosine was found to induce G-2/M cell-cycle arrest. Furthermore, adenosine was found to modulate the expression of some important proteins in the cell cycle, such as cyclin B and p21, and to inhibit the transition of metaphase to anaphase in mitosis.

  6. Label-free fluorescence dual-amplified detection of adenosine based on exonuclease III-assisted DNA cycling and hybridization chain reaction.

    PubMed

    Sun, Jiewei; Jiang, Wei; Zhu, Jing; Li, Wei; Wang, Lei

    2015-08-15

    In this work, we constructed a label-free and dual-amplified fluorescence aptasensor for sensitive analysis of adenosine based on exonuclease III (Exo III)-assisted DNA cycling and hybridization chain reaction (HCR). Firstly, we fabricated a trifunctional probe that consisting of the catalytic strand, the aptamer sequence and a streptavidin-magnetic nanobead (streptavidin-MNB). The streptavidin-MNB played a role of enrichment and separation to achieve a low background. The aptamer sequence was employed as a recognition element to bind the target adenosine, leading to the releasing of the catalytic stand. Then, the catalytic strand induced the Exo III-assisted DNA cycling reaction and produced a large amount of DNA fragments, which got a primary amplification. Subsequently, the DNA fragments acted as trigger strands to initiate HCR, forming nicked double helices with multiple G-quadruplex structures, which achieved a secondary amplification. Finally, the G-quadruplex structures bonded with the N-nethyl mesopor-phyrin IX (NMM) and yielded an enhanced fluorescence signal, realizing the label-free detection. In the proposed strategy, a small amount of adenosine can be converted to a large amount of DNA triggers, leading to a significant amplification for the target. This method exhibited a high sensitivity toward adenosine with a detection limit of 4.2×10(-7) mol L(-1), which was about 10 times lower than that of the reported label-free strategies. Moreover, this assay can significantly distinguish the content of adenosine in urine samples of cancer patients and normal human, indicating that our method will offer a new strategy for reliable quantification of adenosine in medical research and early clinical diagnosis. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Adenosine A3 receptor elicits chemoresistance mediated by multiple resistance-associated protein-1 in human glioblastoma stem-like cells.

    PubMed

    Torres, Angelo; Vargas, Yosselyn; Uribe, Daniel; Jaramillo, Catherine; Gleisner, Alejandra; Salazar-Onfray, Flavio; López, Mercedes N; Melo, Rómulo; Oyarzún, Carlos; San Martín, Rody; Quezada, Claudia

    2016-10-11

    MRP1 transporter correlates positively with glioma malignancy and the Multiple Drug Resistance (MDR) phenotype in Glioblastoma Multiforme (GBM). Evidence shows that the MRP1 transporter is controlled by the adenosine signalling axis. The aim of this study was to identify the role of adenosine on the MDR phenotype in Glioblastoma Stem-like Cells (GSCs), the cell population responsible for the tumorigenic and chemoresistance capabilities of this tumour. We found that GSCs have increased intrinsic capacity to generate extracellular adenosine, thus controlling MRP1 transporter expression and activity via activation of the adenosine A3 receptor (A3AR). We showed PI3K/Akt and MEK/ERK1/2 signaling pathways downstream A3AR to control MRP1 in GSCs. In vitro pharmacological blockade of A3AR had a chemosensitizing effect, enhancing the actions of antitumour drugs and decreasing cell viability and proliferation of GSCs. In addition, we produced an in vivo xenograft model by subcutaneous inoculation of human GSCs in NOD/SCID-IL2Rg null mice. Pharmacological blockade of A3AR generated a chemosensitizing effect, enhancing the effectiveness of the MRP1 transporter substrate, vincristine, reducing tumour size and the levels of CD44 and Nestin stem cell markers as well as the Ki-67 proliferation indicator. In conclusion, we demonstrated the chemosensitizing effect of A3AR blockade on GSCs.

  8. Adenosine A3 receptor elicits chemoresistance mediated by multiple resistance-associated protein-1 in human glioblastoma stem-like cells

    PubMed Central

    Torres, Angelo; Vargas, Yosselyn; Uribe, Daniel; Jaramillo, Catherine; Gleisner, Alejandra; Salazar-Onfray, Flavio; López, Mercedes N.; Melo, Rómulo; Oyarzún, Carlos; Martín, Rody San; Quezada, Claudia

    2016-01-01

    MRP1 transporter correlates positively with glioma malignancy and the Multiple Drug Resistance (MDR) phenotype in Glioblastoma Multiforme (GBM). Evidence shows that the MRP1 transporter is controlled by the adenosine signalling axis. The aim of this study was to identify the role of adenosine on the MDR phenotype in Glioblastoma Stem-like Cells (GSCs), the cell population responsible for the tumorigenic and chemoresistance capabilities of this tumour. We found that GSCs have increased intrinsic capacity to generate extracellular adenosine, thus controlling MRP1 transporter expression and activity via activation of the adenosine A3 receptor (A3AR). We showed PI3K/Akt and MEK/ERK1/2 signaling pathways downstream A3AR to control MRP1 in GSCs. In vitro pharmacological blockade of A3AR had a chemosensitizing effect, enhancing the actions of antitumour drugs and decreasing cell viability and proliferation of GSCs. In addition, we produced an in vivo xenograft model by subcutaneous inoculation of human GSCs in NOD/SCID-IL2Rg null mice. Pharmacological blockade of A3AR generated a chemosensitizing effect, enhancing the effectiveness of the MRP1 transporter substrate, vincristine, reducing tumour size and the levels of CD44 and Nestin stem cell markers as well as the Ki-67 proliferation indicator. In conclusion, we demonstrated the chemosensitizing effect of A3AR blockade on GSCs. PMID:27634913

  9. Mechanism of A2 adenosine receptor activation. I. Blockade of A2 adenosine receptors by photoaffinity labeling

    SciTech Connect

    Lohse, M.J.; Klotz, K.N.; Schwabe, U.

    1991-04-01

    It has previously been shown that covalent incorporation of the photoreactive adenosine derivative (R)-2-azido-N6-p-hydroxy-phenylisopropyladenosine ((R)-AHPIA) into the A1 adenosine receptor of intact fat cells leads to a persistent activation of this receptor, resulting in a reduction of cellular cAMP levels. In contrast, covalent incorporation of (R)-AHPIA into human platelet membranes, which contain only stimulatory A2 adenosine receptors, reduces adenylate cyclase stimulation via these receptors. This effect of (R)-AHPIA is specific for the A2 receptor and can be prevented by the adenosine receptor antagonist theophylline. Binding studies indicate that up to 90% of A2 receptors can be blocked by photoincorporation of (R)-AHPIA. However, the remaining 10-20% of A2 receptors are sufficient to mediate an adenylate cyclase stimulation of up to 50% of the control value. Similarly, the activation via these 10-20% of receptors occurs with a half-life that is only 2 times longer than that in control membranes. This indicates the presence of a receptor reserve, with respect to both the extent and the rate of adenylate cyclase stimulation. These observations require a modification of the models of receptor-adenylate cyclase coupling.

  10. Submaximal adenosine-induced coronary hyperaemia with 12 h caffeine abstinence: implications for clinical adenosine perfusion imaging tests.

    PubMed

    Carlsson, Marcus; Jögi, Jonas; Bloch, Karin M; Hedén, Bo; Ekelund, Ulf; Ståhlberg, Freddy; Arheden, Håkan

    2015-01-01

    Adenosine is widely used as a vasodilator agent in myocardial perfusion imaging. Caffeine inhibits the effect, but the time of caffeine abstinence needed is under discussion and varies from 12 to 24 h. Therefore, our aim was to examine whether the time of caffeine abstinence affects the hyperaemic response using quantification of coronary sinus flow (CS F) with cardiac magnetic resonance (CMR) during adenosine infusion. Healthy individuals (n = 16, eight females, age 41 ± 3 years) underwent two CMR examinations with 12 and 24 h of caffeine abstinence. CS F was quantified with phase-contrast velocity mapping (PC-)CMR during adenosine infusion (140 μg kg(-1)  min(-1) ) and rest and the CS F reserve between adenosine and rest was calculated. Myocardial perfusion (MP) was calculated as CS F × heart rate/left ventricular mass. Cardiac output (CO) was quantified using PC-CMR of the ascending aorta. The CS F reserve was lower after 12 h abstinence compared to 24 h (4·31 ± 0·57 versus 5·32 ± 0·76, P = 0·03). In six of 16 subjects (38%), CS F reserve was >30% higher with longer caffeine abstinence. MP during adenosine was lower after 12 h compared to 24 h caffeine abstinence (3·59 ± 0·37 versus 4·23 ± 0·28 ml min(-1) g(-1) ; P = 0·046). The increase in CO during adenosine between the two occasions did not differ (55 ± 7% and 55 ± 6%, P = 0·11). Interobserver variability for CS F/heartbeat was -0·05 ± 1·00 ml. Hyperaemia during adenosine is lower in some patients with 12 h of caffeine abstinence compared to 24 h. Longer caffeine abstinence, that is 24 h, is of value before pharmacological stress testing as the individual response is not known and the individual variation is large. © 2014 Scandinavian Society of Clinical Physiology and Nuclear Medicine. Published by John Wiley & Sons Ltd.

  11. Fast-scan Cyclic Voltammetry for the Characterization of Rapid Adenosine Release

    PubMed Central

    Nguyen, Michael D.; Venton, B. Jill

    2014-01-01

    Adenosine is a signaling molecule and downstream product of ATP that acts as a neuromodulator. Adenosine regulates physiological processes, such as neurotransmission and blood flow, on a time scale of minutes to hours. Recent developments in electrochemical techniques, including fast-scan cyclic voltammetry (FSCV), have allowed direct detection of adenosine with sub-second temporal resolution. FSCV studies have revealed a novel mode of rapid signaling that lasts only a few seconds. This rapid release of adenosine can be evoked by electrical or mechanical stimulations or it can be observed spontaneously without stimulation. Adenosine signaling on this time scale is activity dependent; however, the mode of release is not fully understood. Rapid adenosine release modulates oxygen levels and evoked dopamine release, indicating that adenosine may have a rapid modulatory role. In this review, we outline how FSCV can be used to detect adenosine release, compare FSCV with other techniques used to measure adenosine, and present an overview of adenosine signaling that has been characterized using FSCV. These studies point to a rapid mode of adenosine modulation, whose mechanism and function will continue to be characterized in the future. PMID:26900429

  12. Fast-scan Cyclic Voltammetry for the Characterization of Rapid Adenosine Release.

    PubMed

    Nguyen, Michael D; Venton, B Jill

    2015-01-01

    Adenosine is a signaling molecule and downstream product of ATP that acts as a neuromodulator. Adenosine regulates physiological processes, such as neurotransmission and blood flow, on a time scale of minutes to hours. Recent developments in electrochemical techniques, including fast-scan cyclic voltammetry (FSCV), have allowed direct detection of adenosine with sub-second temporal resolution. FSCV studies have revealed a novel mode of rapid signaling that lasts only a few seconds. This rapid release of adenosine can be evoked by electrical or mechanical stimulations or it can be observed spontaneously without stimulation. Adenosine signaling on this time scale is activity dependent; however, the mode of release is not fully understood. Rapid adenosine release modulates oxygen levels and evoked dopamine release, indicating that adenosine may have a rapid modulatory role. In this review, we outline how FSCV can be used to detect adenosine release, compare FSCV with other techniques used to measure adenosine, and present an overview of adenosine signaling that has been characterized using FSCV. These studies point to a rapid mode of adenosine modulation, whose mechanism and function will continue to be characterized in the future.

  13. Metabolic changes of cultured DRG neurons induced by adenosine using confocal microscopy imaging

    NASA Astrophysics Data System (ADS)

    Zheng, Liqin; Huang, Yimei; Chen, Jiangxu; Wang, Yuhua; Yang, Hongqin; Zhang, Yanding; Xie, Shusen

    2012-12-01

    Adenosine exerts multiple effects on pain transmission in the peripheral nervous system. This study was performed to use confocal microscopy to evaluate whether adenosine could affect dorsal root ganglia (DRG) neurons in vitro and test which adenosine receptor mediates the effect of adenosine on DRG neurons. After adding adenosine with different concentration, we compared the metabolic changes by the real time imaging of calcium and mitochondria membrane potential using confocal microscopy. The results showed that the effect of 500 μM adenosine on the metabolic changes of DRG neurons was more significant than others. Furthermore, four different adenosine receptor antagonists were used to study which receptor mediated the influences of adenosine on the cultured DRG neurons. All adenosine receptor antagonists especially A1 receptor antagonist (DPCPX) had effect on the Ca2+ and mitochondria membrane potential dynamics of DRG neurons. The above studies demonstrated that the effect of adenosine which may be involved in the signal transmission on the sensory neurons was dose-dependent, and all the four adenosine receptors especially the A1R may mediate the transmission.

  14. Adenosine transiently modulates stimulated dopamine release in the caudate putamen via A1 receptors

    PubMed Central

    Ross, Ashley E.; Venton, B. Jill

    2014-01-01

    Adenosine modulates dopamine in the brain via A1 and A2A receptors, but that modulation has only been characterized on a slow time scale. Recent studies have characterized a rapid signaling mode of adenosine that suggests a possible rapid modulatory role. Here, fast-scan cyclic voltammetry was used to characterize the extent to which transient adenosine changes modulate stimulated dopamine release (5 pulses at 60 Hz) in rat caudate putamen brain slices. Exogenous adenosine was applied and dopamine concentration monitored. Adenosine only modulated dopamine when it was applied 2 or 5 s before stimulation. Longer time intervals and bath application of 5 µM adenosine did not decrease dopamine release. Mechanical stimulation of endogenous adenosine 2s before dopamine stimulation also decreased stimulated dopamine release by 41 ± 7 %, similar to the 54 ± 6 % decrease in dopamine after exogenous adenosine application. Dopamine inhibition by transient adenosine was recovered within 10 minutes. The A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) blocked the dopamine modulation, whereas dopamine modulation was unaffected by the A2A receptor antagonist SCH 442416. Thus, transient adenosine changes can transiently modulate phasic dopamine release via A1 receptors. These data demonstrate that adenosine has a rapid, but transient, modulatory role in the brain. PMID:25219576

  15. Adenosine augmentation therapies (AATs) for epilepsy: prospect of cell and gene therapies

    PubMed Central

    Boison, Detlev

    2009-01-01

    Deficiencies in the brain’s own adenosine-based seizure control system contribute to seizure generation. Consequently, reconstitution of adenosinergic neuromodulation constitutes a rational approach for seizure control. This review will critically discuss focal adenosine augmentation strategies and their potential for antiepileptic and disease modifying therapy. Due to systemic side effects of adenosine focal adenosine augmentation – ideally targeted to an epileptic focus – becomes a therapeutic necessity. This has experimentally been achieved in kindled seizure models as well as in post status epilepticus models of spontaneous recurrent seizures using three different therapeutic strategies that will be discussed here: (i) Polymer-based brain implants that were loaded with adenosine; (ii) Brain implants comprised of cells engineered to release adenosine and embedded in a cell-encapsulation device; (iii) Direct transplantation of stem cells engineered to release adenosine. To meet the therapeutic goal of focal adenosine augmentation, genetic disruption of the adenosine metabolizing enzyme adenosine kinase (ADK) in rodent and human cells was used as a molecular strategy to induce adenosine release from cellular brain implants, which demonstrated antiepileptic and neuroprotective properties. New developments and therapeutic challenges in using AATs for epilepsy therapy will critically be evaluated. PMID:19428218

  16. Protective effect of adenosine against a calcium paradox in the isolated frog heart.

    PubMed

    Touraki, M; Lazou, A

    1992-01-01

    The effect of adenosine on the calcium paradox in the isolated frog heart was studied. Addition of adenosine during calcium depletion protected the frog heart against a calcium paradox. This protective effect was indicated by reduced protein and creatine kinase release, maintenance of electrical activity, and recovery of mechanical activity during reperfusion. Tissue calcium determination results showed that adenosine protected frog myocardial cells by reducing the massive calcium influx during reperfusion possibly through an action on calcium channels. Adenosine exerted its action in a dose-dependent manner; a concentration of 10 microM adenosine provided maximum protection of myocardial cells against the calcium paradox damage. Higher concentrations of adenosine produced side effects on both electrical and mechanical activity. These results are discussed in terms of the possible mechanism involved in the protective effect of adenosine.

  17. Adenosine Signaling Increases Proinflammatory and Profibrotic Mediators through Activation of a Functional Adenosine 2B Receptor in Renal Fibroblasts.

    PubMed

    Wilkinson, Patrick F; Farrell, Francis X; Morel, Diane; Law, William; Murphy, Suzanne

    2016-07-01

    Interstitial renal fibrosis is a major pathophysiological manifestation of patients diagnosed with Chronic Kidney Disease (CKD), Diabetic Nephropathy (DN) and other inflammatory diseases. Adenosine signaling is an innate autocrine and paracrine cellular signaling pathway involving several key mediators that are elevated in the blood and kidneys of patients with DN. In these studies, we hypothesized that extracellular adenosine signals through one or more functional adenosine GPCRs on renal fibroblasts which increases profibrotic and proinflammatory mediators by inducing an activated fibroblast phenotype. Utilizing the renal fibroblast cell line NRK-49F, the presence and relative abundance of adenosine receptors (AR) A1, A2A, A2B, and A3 were quantified by RT-PCR. Under normal homeostatic conditions, only AR1 and AR2B were detected. The functionality of each receptor was then assessed by receptor specific pharmacological agonism and antagonism and assessed for modulation of the GPCR associated secondary messenger molecule, cyclic adenosine monophosphate (cAMP). Agonism of the AR2B receptor resulted in increased intracellular cAMP while agonism of the AR1 receptor inhibited cAMP modulation. Upon direct agonism of the AR2B receptor, transcripts for profibrotic and inflammatory mediators including SMA-α, IL-6, TGF-β, CTGF, and fibronectin were elevated between 2-4 fold. These data indicate that renal fibroblasts express a functional AR1 receptor that inhibits cAMP upon stimulation, leading to a functional AR2B receptor that increases cAMP upon stimulation and also induces an activated fibroblast phenotype resulting in increased fibrotic and inflammatory mediators.

  18. Epithelial-specific A2B adenosine receptor signaling protects the colonic epithelial barrier during acute colitis

    PubMed Central

    Aherne, CM; Saeedi, B; Collins, CB; Masterson, JC; McNamee, EN; Perrenoud, L; Rapp, CR; Curtis, VF; Bayless, A; Fletcher, A; Glover, LE; Evans, CM; Jedlicka, P; Furuta, GT; de Zoeten, EF; Colgan, SP; Eltzschig, HK

    2015-01-01

    Central to inflammatory bowel disease (IBD) pathogenesis is loss of mucosal barrier function. Emerging evidence implicates extracellular adenosine signaling in attenuating mucosal inflammation. We hypothesized that adenosine-mediated protection from intestinal barrier dysfunction involves tissue-specific signaling through the A2B adenosine receptor (Adora2b) at the intestinal mucosal surface. To address this hypothesis, we combined pharmacologic studies and studies in mice with global or tissue-specific deletion of the Adora2b receptor. Adora2b−/− mice experienced a significantly heightened severity of colitis, associated with a more acute onset of disease and loss of intestinal epithelial barrier function. Comparison of mice with Adora2b deletion on vascular endothelial cells (Adora2bfl/flVeCadCre+) or intestinal epithelia (Adora2bfl/flVillinCre+) revealed a selective role for epithelial Adora2b signaling in attenuating colonic inflammation. In vitro studies with Adora2b knockdown in intestinal epithelial cultures or pharmacologic studies highlighted Adora2b-driven phosphorylation of vasodilator-stimulated phosphoprotein (VASP) as a specific barrier repair response. Similarly, in vivo studies in genetic mouse models or treatment studies with an Adora2b agonist (BAY 60-6583) recapitulate these findings. Taken together, our results suggest that intestinal epithelial Adora2b signaling provides protection during intestinal inflammation via enhancing mucosal barrier responses. PMID:25850656

  19. Adenosine A2B Receptor Deficiency Promotes Host Defenses against Gram-Negative Bacterial Pneumonia

    PubMed Central

    Barletta, Kathryn E.; Cagnina, R. Elaine; Burdick, Marie D.; Linden, Joel

    2012-01-01

    Rationale: Activation of the adenosine A2B receptor (A2BR) promotes antiinflammatory effects in diverse biological settings, but the role of this receptor in antimicrobial host defense in the lung has not been established. Gram-negative bacillary pneumonia is a common and serious illness associated with high morbidity and mortality, the treatment of which is complicated by increasing rates of antibiotic resistance. Objectives: To test the hypothesis that absence of adenosine A2B receptor signaling promotes host defense against bacterial pneumonia. Methods: We used a model of Klebsiella pneumoniae pneumonia in wild-type mice and mice with targeted deletion of the A2BR. Host responses were compared in vivo and leukocyte responses to the bacteria were examined in vitro. Measurements and Main Results: A2BR–/– mice demonstrated enhanced bacterial clearance from the lung and improved survival after infection with K. pneumoniae compared with wild-type controls, an effect that was mediated by bone marrow–derived cells. Leukocyte recruitment to the lungs and expression of inflammatory cytokines did not differ between A2BR–/– and wild-type mice, but A2BR–/– neutrophils exhibited sixfold greater bactericidal activity and enhanced production of neutrophil extracellular traps compared with wild-type neutrophils when incubated with K. pneumoniae. Consistent with this finding, bronchoalveolar lavage fluid from A2BR–/– mice with Klebsiella pneumonia contained more extracellular DNA compared with wild-type mice with pneumonia. Conclusions: These data suggest that the absence of A2BR signaling enhances antimicrobial activity in gram-negative bacterial pneumonia. PMID:22997203

  20. Increase in adenosine sensitivity in the nucleus accumbens following chronic morphine treatment.

    PubMed

    Brundege, James M; Williams, John T

    2002-03-01

    There is a growing body of evidence suggesting that the neuromodulator adenosine is involved in drug addiction and withdrawal and that adenosine signaling pathways may offer new targets for therapeutic treatments of addiction. Recent studies have suggested that chronic exposure to drugs of abuse may alter adenosine metabolism in the nucleus accumbens, a brain region critically involved in drug addiction and withdrawal. The present study examined the effects of chronic morphine treatment on the ability of adenosine to inhibit excitatory postsynaptic currents in nucleus accumbens medium spiny neurons. It was found that chronic morphine treatment via subcutaneous implantation of morphine pellets in rats for 1 wk did not alter the level of adenosine-mediated tonic inhibition of nucleus accumbens excitatory synapses. However, chronic morphine treatment did induce a leftward shift in the adenosine dose-response curve, indicating an increase in the sensitivity of synaptic currents to exogenously applied adenosine. This shift was not due to a change in adenosine receptors or their effectors, because chronic morphine treatment had no effect on the dose-response relationship of a nonmetabolized adenosine receptor agonist. When adenosine transport was blocked, the ability of chronic morphine to shift the adenosine dose-response curve was eliminated. These experiments suggest that the increase in the sensitivity of nucleus accumbens synapses to the inhibitory effects of adenosine may be due to a decrease in adenosine transport. The identification of these changes in the adenosine system after chronic drug exposure may help identify new therapeutic strategies aimed at easing withdrawal from opioids.

  1. Extracellular adenosine levels are associated with the progression and exacerbation of pulmonary fibrosis

    PubMed Central

    Luo, Fayong; Le, Ngoc-Bao; Mills, Tingting; Chen, Ning-Yuan; Karmouty-Quintana, Harry; Molina, Jose G.; Davies, Jonathan; Philip, Kemly; Volcik, Kelly A.; Liu, Hong; Xia, Yang; Eltzschig, Holger K.; Blackburn, Michael R.

    2015-01-01

    Idiopathic pulmonary fibrosis is a devastating lung disease with limited treatment options. The signaling molecule adenosine is produced in response to injury and serves a protective role in early stages of injury and is detrimental during chronic stages of disease such as seen in lung conditions such as pulmonary fibrosis. Understanding the association of extracellular adenosine levels and the progression of pulmonary fibrosis is critical for designing adenosine based approaches to treat pulmonary fibrosis. The goal of this study was to use various models of experimental lung fibrosis to understand when adenosine levels are elevated during pulmonary fibrosis and whether these elevations were associated with disease progression and severity. To accomplish this, extracellular adenosine levels, defined as adenosine levels found in bronchioalveolar lavage fluid, were determined in mouse models of resolvable and progressive pulmonary fibrosis. We found that relative bronchioalveolar lavage fluid adenosine levels are progressively elevated in association with pulmonary fibrosis and that adenosine levels diminish in association with the resolution of lung fibrosis. In addition, treatment of these models with dipyridamole, an inhibitor of nucleoside transporters that potentiates extracellular adenosine levels, demonstrated that the resolution of lung fibrosis is blocked by the failure of adenosine levels to subside. Furthermore, exacerbating adenosine levels led to worse fibrosis in a progressive fibrosis model. Increased adenosine levels were associated with elevation of IL-6 and IL-17, which are important inflammatory cytokines in pulmonary fibrosis. These results demonstrate that extracellular adenosine levels are closely associated with the progression of experimental pulmonary fibrosis and that this signaling pathway may mediate fibrosis by regulating IL-6 and IL-17 production.—Luo, F., Le, N.-B., Mills, T., Chen, N.-Y., Karmouty-Quintana, H., Molina, J. G., Davies

  2. Adenosine A2 receptors modulate haloperidol-induced catalepsy in rats.

    PubMed

    Mandhane, S N; Chopde, C T; Ghosh, A K

    1997-06-11

    The effect of adenosine A1 and A2 receptor agonists and antagonists was investigated on haloperidol-induced catalepsy in rats. Pretreatment (i.p.) with the non-selective adenosine receptor antagonist, theophylline, or the selective adenosine A2 receptor antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX), significantly reversed haloperidol-induced catalepsy, whereas the selective adenosine A1 receptor antagonists, 8-phenyltheophylline and 8-cyclopentyl-1,3-dipropylxanthine produced no effect. Similar administration of the adenosine A2 receptor agonists, 5'-(N-cyclopropyl)-carboxamidoadenosine and 5'-N-ethylcarboxamidoadenosine (NECA), and the mixed agonists with predominantly A1 site of action, N6-(2-phenylisopropyl) adenosine or 2-chloroadenosine, potentiated haloperidol-induced catalepsy. Higher doses of the adenosine agonists produced catalepsy when given alone. However, N6-cyclopentyladenosine, a highly selective adenosine A1 receptor agonist, was ineffective in these respects. The per se cataleptic effect of adenosine agonists was blocked by DMPX and the centrally acting anticholinergic agent, scopolamine. Scopolamine also attenuated the potentiation of haloperidol-induced catalepsy by adenosine agonists. Further, i.c.v. administration of NECA and DMPX produced a similar effect as that produced after their systemic administration. These findings demonstrate the differential influence of adenosine A1 and A2 receptors on haloperidol-induced catalepsy and support the hypothesis that the functional interaction between adenosine and dopamine mechanisms might occur through adenosine A2 receptors at the level of cholinergic neurons. The results suggest that adenosine A2, but not A1, receptor antagonists may be of potential use in the treatment of Parkinson's disease.

  3. Adenosine transiently modulates stimulated dopamine release in the caudate-putamen via A1 receptors.

    PubMed

    Ross, Ashley E; Venton, B Jill

    2015-01-01

    Adenosine modulates dopamine in the brain via A1 and A2A receptors, but that modulation has only been characterized on a slow time scale. Recent studies have characterized a rapid signaling mode of adenosine that suggests a possible rapid modulatory role. Here, fast-scan cyclic voltammetry was used to characterize the extent to which transient adenosine changes modulate stimulated dopamine release (5 pulses at 60 Hz) in rat caudate-putamen brain slices. Exogenous adenosine was applied and dopamine concentration monitored. Adenosine only modulated dopamine when it was applied 2 or 5 s before stimulation. Longer time intervals and bath application of 5 μM adenosine did not decrease dopamine release. Mechanical stimulation of endogenous adenosine 2 s before dopamine stimulation also decreased stimulated dopamine release by 41 ± 7%, similar to the 54 ± 6% decrease in dopamine after exogenous adenosine application. Dopamine inhibition by transient adenosine was recovered within 10 min. The A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine blocked the dopamine modulation, whereas dopamine modulation was unaffected by the A2A receptor antagonist SCH 442416. Thus, transient adenosine changes can transiently modulate phasic dopamine release via A1 receptors. These data demonstrate that adenosine has a rapid, but transient, modulatory role in the brain. Here, transient adenosine was shown to modulate phasic dopamine release on the order of seconds by acting at the A1 receptor. However, sustained increases in adenosine did not regulate phasic dopamine release. This study demonstrates for the first time a transient, neuromodulatory function of rapid adenosine to regulate rapid neurotransmitter release.

  4. Effects of caffeine on behavioral and inflammatory changes elicited by copper in zebrafish larvae: Role of adenosine receptors.

    PubMed

    Cruz, Fernanda Fernandes; Leite, Carlos Eduardo; Kist, Luiza Wilges; de Oliveira, Giovanna Medeiros; Bogo, Maurício Reis; Bonan, Carla Denise; Campos, Maria Martha; Morrone, Fernanda Bueno

    2017-04-01

    This study investigated the effects of caffeine in the behavioral and inflammatory alterations caused by copper in zebrafish larvae, attempting to correlate these changes with the modulation of adenosine receptors. To perform a survival curve, 7dpf larvae were exposed to 10μM CuSO4, combined to different concentrations of caffeine (100μM, 500μM and 1mM) for up to 24h. The treatment with copper showed lower survival rates only when combined with 500μM and 1mM of caffeine. We selected 4 and 24h as treatment time-points. The behavior evaluation was done by analyzing the traveled distance, the number of entries in the center, and the length of permanence in the center and the periphery of the well. The exposure to 10μM CuSO4 plus 500μM caffeine at 4 and 24h changed the behavioral parameters. To study the inflammatory effects of caffeine, we assessed the PGE2 levels by using UHPLC-MS/MS, and TNF, COX-2, IL-6 and IL-10 gene expression by RT-qPCR. The expression of adenosine receptors was also evaluated with RT-qPCR. When combined to copper, caffeine altered inflammatory markers depending on the time of exposure. Adenosine receptors expression was significantly increased, especially after 4h exposure to copper and caffeine together or separately. Our results demonstrated that caffeine enhances the inflammation induced by copper by decreasing animal survival, altering inflammatory markers and promoting behavioral changes in zebrafish larvae. We also conclude that alterations in adenosine receptors are related to those effects.

  5. Modulation of vasodilatation to levcromakalim by adenosine analogues in the rabbit ear: an explanation for hypoxic augmentation.

    PubMed Central

    Randall, M. D.; Ujiie, H.; Griffith, T. M.

    1994-01-01

    1. We have used a rabbit isolated ear, buffered-perfused preparation to investigate the effects of adenosine analogues on the vasodilatation to the potassium channel opener, levcromakalim (the active (-)-enantiomer of cromakalim). We have examined the effects of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), a selective adenosine A1 antagonist, on vasodilatation to levcromakalim under hypoxic conditions and also following inhibition of nitric oxide synthesis. 2. Levcromakalim relaxed preconstricted preparations with an EC50 = 369 +/- 48 nM and maximum relaxation of tone (Rmax) = 81.0 +/- 3.2%. In the presence of 1 microM N6-cyclohexyladenosine (CHA) a selective adenosine A1 agonist, there was a significant (P < 0.01) leftward shift in the concentration-response curve with an EC50 = 194 +/- 54 nM and Rmax = 93.2 +/- 2.0%. Conversely, the presence of CHA did not influence vasodilatation to either pinacidil or sodium nitroprusside. 3. Hypoxia also significantly (P < 0.001) increased the vasodilator potency of levcromakalim (EC50 = 134 +/- 22 nM), and this enhancement was completely reversed (EC50 = 380 +/- 107 nM, P < 0.01) by pretreatment of the preparations with 5 microM DPCPX, a selective A1 adenosine antagonist. However, under normoxic conditions DPCPX did not influence vasodilatation to levcromakalim. 4. Inhibition of nitric oxide synthesis with 100 microM NG-nitro-L-arginine methyl ester (L-NAME) caused a significant (P < 0.001) leftward shift in the concentration-response curve to levcromakalim (EC50 = 73.0 +/- 7.6 nM). Pretreatment of preparations with DPCPX partially reversed the increase in potency found in the absence of nitric oxide synthesis (EC50 = 153 +/- 18 nM, P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8032661

  6. Properties of enzyme fraction A from Chlorella and copurification of 3' (2'), 5'-biphosphonucleoside 3' (2')-phosphohydrolase, adenosine 5'phosphosulfate sulfohydrolase and adenosine-5'-phosphosulfate cyclase activities.

    PubMed

    Lik-Shing Tsang, M; Schiff, J A

    1976-05-17

    Enzyme fraction A from Chlorella which catalyzes the formation of adenosine 5'-phosphosulfate from adenosine 3'-phosphate 5'-phosphosulfate is further characterized. Fraction A is found to contain an Mg2+ -activated and Ca2+ -inhibited 3' (2')-nucleotidase specific for 3' (2'), 5'-biphosphonucleosides. This activity has been named 3' (2), 5'-biphosphonucleoside 3' (2')-phosphohydrolase. The A fraction is also found to contain an activity which catalyzes the formation of adenosine 3':5'-monophosphate (cyclic AMP) from adenosine 5'-phosphosulfate (adenosine 5'-phosphosulfate cyclase). Under the same conditions of assay, 5'-ATP and 5'-ADP are not substrated for cyclic AMP formation. Unlike the 3' (2'), 5'-biphosphonucleoside 3' (2')-phosphohydrolase activity, the adenosine 5'-phosphosulfate cyclase activity does not require Mg2+, requires NH+4 or Na+, and is not inhibited by Ca2+. The A fraction also contains an adenosine 5'-phospho sulfate sulfohydrolase activity which forms 5'-AMP and sulfate. The three activities remain together during purification and acrylamide gel electrophoresis of the purified preparation yields a pattern where only one protein band has all three activities. The phosphohydrolase can be separated from the other two activities by affinity chromatography on agarose-hexyl-adenosine 3'n5'-bisphosphate yielding a phosphohydrolase preparation showing a single band on gel electrophoresis. The adenosine 5'-phosphosulfate cyclase may provide an alternate route of cyclic AMP formation from sulfate via ATP sulfurylase, but its regulatory significance in Chlorella, if any, remains to be demonstrated. In sulfate reduction, the phosphohydrolase may serve to provide a readily utilized pool of adenosine 5'-phosphosulfate as needed by the adenosine 5'-phosphosulfate sulfotransferase. The cyclase and sulfohydrolase activities would be regarded as side reactions incidental to this pathway, but may be of importance in other metabolic and regulatory reactions.

  7. Spinal serotonin 5-HT7 and adenosine A1 receptors, as well as peripheral adenosine A1 receptors, are involved in antinociception by systemically administered amitriptyline.

    PubMed

    Liu, Jean; Reid, Allison R; Sawynok, Jana

    2013-01-05

    The present study explored a link between spinal 5-HT(7) and