Exogenous Adenosine Triphosphate Research Articles

The two-hit hypothesis for neuroinflammation: role of exogenous ATP in modulating inflammation in the brain.

Brain inflammation is a common occurrence following responses to varied insults such as bacterial infections, stroke, traumatic brain injury and neurodegenerative disorders. A common mediator for these varied inflammatory responses is prostaglandin E2 (PGE2), produced by the enzymatic activity of cyclooxygenases (COX) 1 and 2. Previous attempts to reduce neuronal inflammation through COX inhibition, by use of nonsteroidal anti-inflammatory drugs (NSAIDs), have met with limited success. We are proposing the two-hit model for neuronal injury—an initial localized inflammation mediated by PGE2 (first hit) and the simultaneous release of adenosine triphosphate (ATP) by injured cells (second hit), which significantly enhances the inflammatory response through increased synthesis of PGE2. Several evidences on the role of exogenous ATP in inflammation have been reported, including contrary instances where extracellular ATP reduces inflammatory events. In this review, we will examine the current literature on the role of P2 receptors, to which ATP binds, in modulating inflammatory reactions during neurodegeneration. Targeting the P2 receptors, therefore, provides a therapeutic alternative to reduce inflammation in the brain. P2 receptor-based anti-inflammatory drugs (PBAIDs) will retain the activities of essential COX enzymes, yet will significantly reduce neuroinflammation by decreasing the enhanced production of PGE2 by extracellular ATP.

Human CD4+ CD39+ regulatory T cells produce adenosine upon co-expression of surface CD73 or contact with CD73+ exosomes or CD73+ cells.

While murine CD4(+) CD39(+) regulatory T cells (T(reg)) co-express CD73 and hydrolyze exogenous (e) adenosine triphosphate (ATP) to immunosuppressive adenosine (ADO), surface co-expression of CD73 on human circulating CD4(+) CD39(+) T(reg) is rare. Therefore, the ability of human T(reg) to produce and utilize ADO for suppression remains unclear. Using mass spectrometry, we measured nucleoside production by subsets of human CD4(+) CD39(+) and CD4(+) CD39(-)CD73(+) T cells or CD19(+) B cells isolated from blood of 30 volunteers and 14 cancer patients. CD39 and CD73 expression was evaluated by flow cytometry, Western blots, confocal microscopy or reverse transcription-polymerase chain reaction (RT-PCR). Circulating CD4(+) CD39(+) T(reg) which hydrolyzed eATP to 5'-AMP contained few intracytoplasmic granules and had low CD73 mRNA levels. Only ∼1% of these T(reg) were CD39(+) CD73(+) . In contrast, CD4(+) CD39(neg) CD73(+) T cells contained numerous CD73(+) granules in the cytoplasm and strongly expressed surface CD73. In vitro-generated T(reg) (Tr1) and most B cells were CD39(+) CD73(+) . All these CD73(+) T cell subsets and B cells hydrolyzed 5'-AMP to ADO. Exosomes isolated from plasma of normal control (NC) or cancer patients carried enzymatically active CD39 and CD73(+) and, when supplied with eATP, hydrolyzed it to ADO. Only CD4(+) CD39(+) T(reg) co-incubated with CD4(+) CD73(+) T cells, B cells or CD39(+) CD73(+) exosomes produced ADO. Thus, contact with membrane-tethered CD73 was sufficient for ADO production by CD4(+) CD39(+) T(reg). In microenvironments containing CD4(+) CD73(+) T cells, B cells or CD39(+) CD73(+) exosomes, CD73 is readily available to CD4(+) CD39(+) CD73(neg) T(reg) for the production of immunosuppressive ADO.

Пластичность мышечной системы в условиях белковой сенсибилизации. Участие холинергических и пуринергических механизмов

The authors investigated the involvement of the adenosine triphosphate (ATF) in the mechanisms of plasticity in the striated muscle in condition of protein sensitization. Contractile function and non-quantum secretion of the acetylcholine in the endplate zone were studied in isolated skeletal muscles of mouse legs (musculus soleus and musculus extensor digitorum longus) and strips musculus diaphragm. The authors studied the soleus and diaphragm dynamics of the force vector of muscle contraction after effects of exogenous adenosine triphosphate correlated with changes non-quantum secretion of acetylcholine in all the experimental models. However, the extent of these changes in sensitized animals is less pronounced than in the controls. It is suggested that adenosine is a party change mechanisms and functional properties soleus and diaphragm at the protein sensitization. The basis of the development of resistance, stability to long as the external stress, and at the initial stages of the experimental allergic these muscles are dependent adenosine mechanisms regulating their sensitivity to acetylcholine. These processes provide a reduction in respiratory muscle fatigue in hypoxia that occurs in asthma, chronic obstructive pulmonary disease and broncho-spastic syndrome, as well in increase their performance during prolonged physical activity. The reasons for changes in the force reduction extensor digitorum longus at a protein sensitization doesn’t associated with the mechanisms of muscle excitation mediated by adenosine triphosphate.

Quality deterioration of cut carnation flowers involves in antioxidant systems and energy status

Adenosine triphosphate (ATP) has played an important role in regulating senescence of horticultural crops. The physiological mechanism of exogenous ATP to affect the senescence of cut carnation during vase life was investigated. Fresh cut carnation flowers were treated with distilled water (control), 0.1mM adenosine triphosphate (ATP) or 0.5mM 2,4-dinitrophenol (DNP, an agent for uncoupling oxidative phosphorylation) and then held at 25°C up to nine days. Exogenous ATP supply increased flower size, maintained fresh weight, extended vase life, reduced ethylene production rate and maintained membrane integrity. ATP treatment also reduced activities of phospholipase D (LD) and lipoxygenase (LOX), enhanced activities of superoxide dismutase (SOD) and catalase (CAT) and increased endogenous contents of ATP and adenosine diphosphate (ADP) and energy charge level at the later vase life. In comparison to control, exogenous application of DNP accelerated ethylene production and reduced vase life, increased membrane permeability, MDA content and activities of PLD and LOX and reduced activities of SOD and CAT. Furthermore, contents of ATP and ADP and energy charge level decreased in the DNP-treated flowers during vase life. Thus, exogenous ATP supply could maintain membrane integrity, increase antioxidant system and improve endogenous energy level, thereby suppressing ethylene production and retarding the senescence of cut carnation flower during vase life.

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

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.

Mitochondrial Uncoupling Protein 2 Induces Cell Cycle Arrest and Necrotic Cell Death

Uncoupling protein 2 (UCP2) is a mitochondrial membrane protein that regulates energy metabolism and reactive oxygen species (ROS) production. We generated mouse carboxy- and amino-terminal green fluorescent protein (GFP)-tagged UCP2 constructs to investigate the effect of UCP2 expression on cell proliferation and viability. UCP2-transfected Hepa 1-6 cells did not show reduced cellular adenosine triphosphate (ATP) but showed increased levels of glutathione. Flow cytometry analysis indicated that transfected cells were less proliferative than nontransfected controls, with most cells blocked at the G1 phase. The effect of UCP2 on cell cycle arrest could not be reversed by providing exogenous ATP or oxidant supply, and was not affected by the chemical uncoupler carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP). However, this effect of UCP2 was augmented by treatment with genistein, a tyrosine kinase inhibitor, which by itself did not affect cell proliferation on control hepatocytes. Western blotting analysis revealed decreased expression levels of CDK6 but not CDK2 and D-type cyclins. Examination of cell viability in UCP2-transfected cells with Trypan Blue and Annexin-V staining revealed that UCP2 transfection led to significantly increased cell death. However, characteristics of apoptosis were absent in UCP2-transfected Hepa 1-6 cells, including lack of oligonucleosomal fragmentation (laddering) of chromosomal DNA, release of cytochrome c from mitochondria, and cleavage of caspase-3. In conclusion, our results indicate that UCP2 induces cell cycle arrest at G1 phase and causes nonapoptotic cell death, suggesting that UCP2 may act as a powerful influence on hepatic regeneration and cell death in the steatotic liver.

Mucosal adenosine triphosphate mediates serotonin release from ileal but not colonic guinea pig enterochromaffin cells

Mechanical stimulation of the mucosal epithelium results in increased serotonin (5-HT) release from enterochromaffin (EC) cells. Little is known about how this process varies in different regions of the intestinal tract; however, purines are felt to play a role. We studied the relationship between mechanical stimulation, adenosine triphosphate (ATP), and 5-HT release from ileal and colonic mucosal tissue. Amperometric recordings of ATP and 5-HT were carried out using an ATP biosensor and boron-doped diamond microelectrode. Levels of extracellular ATP and 5-HT were monitored using high performance liquid chromatography. Under basal conditions, 5-HT levels were significantly decreased in the ileum (p<0.001) but not the colon in the presence of the P2 antagonist suramin (100μM). Ecto-ATPase inhibitor ARL67156 (10μM) elevated ATP levels in the ileum and colon (both p<0.001), but only 5-HT levels in the ileum (p<0.001). Exogenous ATP increased 5-HT release in the presence of tetrodotoxin in the ileum (p<0.001), but had not effect in the colon. Mechanical stimulation increased levels of 5-HT in the ileum (p<0.001) and colon (p<0.01), but levels returned to baseline in the presence of suramin and MRS2179 in the ileum. The onset of 5-HT release was delayed following mechanical stimulation. The rise time of the ATP response was quicker than that of 5-HT during mechanical stimulation. During mechanical stimulation of the mucosal epithelium, ATP mediates 5-HT release from EC cells in the ileum, but not the colon. Mucosal 5-HT signaling following mechanical stimulation is varied in different regions of the intestinal tract.

Adenosine production by human B cells and B cell–mediated suppression of activated T cells

AbstractAntibody-independent role of B cells in modulating T-cell responses is incompletely understood. Freshly isolated or cultured B cells isolated from the peripheral blood of 30 normal donors were evaluated for CD39 and CD73 coexpression, the ability to produce adenosine 5′-monophosphate (AMP) and adenosine (ADO) in the presence of exogenous adenosine triphosphate (ATP) as well as A1, A2A, A2B, and A3 adenosine receptor (ADOR) expression. Human circulating B cells coexpress ectonucleotidases CD39 and CD73, hydrolyze exogenous ATP to 5′-AMP and ADO, and express messenger RNA for A1R, A2AR, and A3R. 2-chloroadenosine inhibited B-cell proliferation and cytokine expression, and only A3R selective antagonist restored B-cell functions. This suggested that B cells use the A3R for autocrine signaling and self-regulation. Mediated effects on B-cell growth ± ADOR antagonists or agonists were tested in carboxyfluorescein diacetate succinimidyl ester assays. In cocultures, resting B cells upregulated functions of CD4+ and CD8+ T cells. However, in vitro–activated B cells downregulated CD73 expression, mainly produced 5′-AMP, and inhibited T-cell proliferation and cytokine production. These B cells acquire the ability to restrict potentially harmful effects of activated T cells. Thus, B cells emerge as a key regulatory component of T cell–B cell interactions, and their dual regulatory activity is mediated by the products of ATP hydrolysis, 5′-AMP, and ADO.

Mechanisms of muscular electrophysiological and mitochondrial dysfunction following exposure to malathion, an organophosphorus pesticide

Muscle dysfunction in acute organophosphorus (OP) poisoning is a cause of death in human. The present study was conducted to identify the mechanism of action of OP in terms of muscle mitochondrial dysfunction. Electromyography (EMG) was conducted on rats exposed to the acute oral dose of malathion (400 mg/kg) that could inhibit acetylcholinesterase activity up to 70%. The function of mitochondrial respiratory chain and the rate of production of reactive oxygen species (ROS) from intact mitochondria were measured. The bioenergetic pathways were studied by measurement of adenosine triphosphate (ATP), lactate, and glycogen. To identify mitochondrial-dependent apoptotic pathways, the messenger RNA (mRNA) expression of bax and bcl-2, protein expression of caspase-9, mitochondrial cytochrome c release, and DNA damage were measured. The EMG confirmed muscle weakness. The reduction in activity of mitochondrial complexes and muscular glycogen with an elevation of lactate was in association with impairment of cellular respiration. The reduction in mitochondrial proapoptotic stimuli is indicative of autophagic process inducing cytoprotective effects in the early stage of stress. Downregulation of apoptotic signaling may be due to reduction in ATP and ROS, and genotoxic potential of malathion. The maintenance of mitochondrial integrity by means of artificial electron donors and increasing exogenous ATP might prevent toxicity of OPs.

ATP Induces NO Production in Hippocampal Neurons by P2X7 Receptor Activation Independent of Glutamate Signaling

To assess the putative role of adenosine triphosphate (ATP) upon nitric oxide (NO) production in the hippocampus, we used as a model both rat hippocampal slices and isolated hippocampal neurons in culture, lacking glial cells. In hippocampal slices, additions of exogenous ATP or 2′(3′)-O-(4-Benzoylbenzoyl) ATP (Bz-ATP) elicited concentration-dependent NO production, which increased linearly within the first 15 min and plateaued thereafter; agonist EC50 values were 50 and 15 µM, respectively. The NO increase evoked by ATP was antagonized in a concentration-dependent manner by Coomassie brilliant blue G (BBG) or by Nω-propyl-L-arginine, suggesting the involvement of P2X7Rs and neuronal NOS, respectively. The ATP induced NO production was independent of N-methyl-D-aspartic acid (NMDA) receptor activity as effects were not alleviated by DL-2-Amino-5-phosphonopentanoic acid (APV), but antagonized by BBG. In sum, exogenous ATP elicited NO production in hippocampal neurons independently of NMDA receptor activity.

ATP‐mediated vasodilatation: all thanks to potassium?

ATP‐mediated vasodilatation: all thanks to potassium?

The role of exogenous adenosine triphosphate administration for the prevention of ischemia-reperfusion injury in skeletal muscle

Organ preservation solutions are essential for the temporary storage of tissue prior to transplantation or reimplantation. The depletion of oxygen stores occurs within a few seconds. Stored energy in the form of creatine phosphate (CP) and adenosine triphosphate (ATP) is depleted. Anaerobic glycolysis then occurs followed by an attempt to maintain normal cellular function. However, glycolysis cannot maintain tissue survival for long. Metabolic waste accumulates and the cellular pH decreases. Mitochondrial ATP generation is blocked because of a lack of molecular oxygen, and insufficient supply of NAD+ also causes the Krebs cycle to cease function.

Administration of exogenous adenosine triphosphate to ischemic skeletal muscle induces an energy-sparing effect: Role of adenosine receptors

BackgroundIschemia–reperfusion injury is a devastating complication that occurs in allotransplantation and replantation of limbs. Over the years, several preservation strategies have been used to conserve the critical levels of intracellular adenosine triphosphate (ATP) during ischemia to sustain the ion gradients across the membranes and thus the tissue viability. The administration of exogenous ATP to ischemic tissues is known to provide beneficial effects during reperfusion, but it is unclear whether it provides protection during ischemia. The purpose of the present study was to determine the effect of ATP administration on high-energy phosphate levels in ischemic skeletal muscle and to examine the role of purinergic and adenosine receptors in mediating the response to exogenous ATP. MethodsThe extensor digitorum longus muscles of Fischer rats were subjected to ischemia and treated with different concentrations of ATP with or without purinergic and adenosine receptor blockers. Phosphorus-31 nuclear magnetic resonance spectroscopy was used to measure the rate of decay of ATP, phosphocreatine (PCr), and the formation of adenosine monophosphate and acidification. Phosphorylated compounds were analyzed using a simple model of energy metabolism, and the PCr half-life was used as an index of internal depletion of ATP to distinguish between intracellular and extracellular ATP. ResultsPCr decay was rapid in all muscle groups and was followed by gradual ATP decay. The half-life of PCr was significantly longer in the ATP-treated muscles than in the vehicle controls and was maximally prolonged by treating with slow hydrolyzing adenosine 5′-O-(3-thio)triphosphate. Purinoceptor (P2X) blockade with ATP treatment significantly increased the half-life of PCr, and adenosine receptor blockers blunted the response. Administration of adenosine to ischemic muscles significantly increased the half-life of PCr compared with that in the vehicle controls. ConclusionsExogenous ATP administration to ischemic skeletal muscles appears to spare intracellular energy by acting primarily through adenosine receptors.

Phenotypic and functional characteristics of CD4+CD39+ FOXP3+ and CD4+CD39+FOXP3neg T‐cell subsets in cancer patients

Human CD4(+) CD39(+) regulatory T (Treg) cells hydrolyze exogenous adenosine triphosphate (ATP) and participate in immunosuppressive adenosine production. They contain two T-cell subsets whose role in mediating suppression is not understood. Frequencies of both CD4(+) CD39(+) subsets were evaluated in peripheral blood lymphocytes of 57 cancer patients and in tumor infiltrating lymphocytes (TILs) of 6 patients. CD4(+) CD39(+) and CD4(+) CD39(neg) T cells isolated using immunobeads and cell sorting were cultured under various conditions. Their conversion into CD39(+) FOXP3(+) CD25(+) or CD39(+) FOX(neg) CD25(neg) cells was monitored by multiparameter flow cytometry. Hydrolysis of exogenous ATP was measured in luminescence assays. Two CD4(+) CD39(+) cell subsets differing in expression of CD25, FOXP3, CTLA-4, CD121a, PD-1, latency associated peptide (LAP), glycoprotein A repetitions predominant (GARP), and the cytokine profile accumulated with equal frequencies in the blood and tumor tissues of cancer patients. The frequency of both subsets was significantly increased in cancer. CD39 expression levels correlated with the subsets' ability to hydrolyze ATP. Conventional CD4(+) CD39(neg) T cells incubated with IL-2 + TGF-β expanded to generate CD4(+) CD39(+) FOXP3(+) Treg cells, while CD4(+) CD39(+) FOXP3(neg) CD25(neg) subset cells stimulated via the TCR and IL-2 converted to FOXP3(+) CTLA4(+) CD25(+) TGF-β-expressing Treg cells. Among CD4(+) CD39(+) Treg cells, the CD4(+) CD39(+) FOXP3(neg) CD25(neg) subset serves as a reservoir of cells able to convert to Treg cells upon activation by environmental signals.

The Therapeutic Potential of Exogenous Adenosine Triphosphate (ATP) for Cartilage Tissue Engineering.

Objective:While mechanical stimuli can be used to enhance the properties of engineered cartilage, a promising alternative may be to directly harness the underlying mechanotransduction pathways responsible. Our initial studies on the adenosine triphosphate (ATP)–purinergic receptor pathway demonstrated that stimulation by exogenous ATP improved tissue growth and properties but elicited matrix turnover under high doses (250 µM) potentially due to the accumulation of extracellular inorganic pyrophosphate (ePPi). Therefore, the purpose of this study was to identify the mechanism of ATP-mediated catabolism and determine a therapeutic dose to maximize the anabolic effect.Design:Isolated bovine articular chondrocytes were seeded in high-density, 3-dimensional culture supplemented with varying doses of ATP for 4 weeks. The effects on biosynthesis, matrix metalloproteinase 13 (MMP-13) protein activity, and PPi accumulation were determined. Separate monolayer experiments were conducted to determine the effect of ePPi on MMP-13 activity.Results:High doses of ATP resulted in an increase in ePPi accumulation (by 54%) and MMP-13 activity (by 39%). Monolayer experiments confirmed a link between increased ePPi accumulation and MMP-13 activity, which appeared to require calcium and was inhibited by the MEK1/2 inhibitor U0126. Cultures supplemented with 62.5 to 125 µM ATP favored an anabolic response, which represented the therapeutic dose range.Conclusions:A therapeutic dose range of exogenous ATP to improve the properties of engineered cartilage has been identified, and a possible catabolic mechanism involving excess PPi was determined. Future research into PPi signal transduction and pathological crystal formation is necessary to maximize the beneficial effect of exogenous ATP on chondrocyte cultures.

Disarming suppressor cells to improve immunotherapy.

Human tumors can use many different mechanisms to induce dysfunction in the host immune system. Accumulations of inducible regulatory T cells (iTreg, Tr1) are commonly seen in the tumor microenvironment. These Treg express CD39 and up-regulate CD73 ectonucleotidases, hydrolyze exogenous adenosine triphosphate (ATP) to AMP and adenosine and produce prostaglandin E(2) (PGE(2)). Most tumors also express CD39/CD73 and COX-2 and thus contribute to immune suppression. Pharmacologic inhibitors can be used to eliminate adenosine/PGE(2) production by Tr1 as well as the tumor or to block binding of these factors to their receptors on Teff or to selectively block cAMP synthesis in Teff. These pharmacologic blocking strategies used alone or in combination with conventional treatments or immunotherapies could disarm Tr1, at the same time restoring antitumor functions of Teff.

Modulation of postjunctional α‐adrenergic vasoconstriction during exercise and exogenous ATP infusions in ageing humans

The ability to modulate sympathetic α-adrenergic vasoconstriction in contracting muscle is impaired with age. In young adults, adenosine triphosphate (ATP) has been shown to blunt sympathetic vasoconstrictor responsiveness similar to exercise. Therefore, we tested the hypothesis that modulation of postjunctional α-adrenergic vasoconstriction to exogenous ATP is impaired in ageing humans.We measured forearm blood flow (FBF; Doppler ultrasound) and calculated vascular conductance (FVC) to intra-arterial infusions of phenylephrine (α₁-agonist) and dexmedetomidine (α₂-agonist) during rhythmic handgrip exercise (15% MVC), a control non-exercise vasodilator condition (adenosine), and ATP infusion in seven older (64 ± 3 years) and seven young (22 ± 1 years) healthy adults. Forearm hyperaemia was matched across all vasodilatating conditions. During adenosine, forearm vasoconstrictor responses to direct α₁-stimulation were lower in older compared with young adults (ΔFVC=-25 ± 3% vs. -41 ± 5%; P <0.05), whereas the responses to α₂-stimulation were not different (-35±6% vs. -44 ± 8%; NS). During exercise, α₁-mediated vasoconstriction was significantly blunted compared with adenosine in both young (-9 ± 2% vs. -41 ± 5%) and older adults (-15 ± 2% vs. -25 ± 3%); however, the magnitude of sympatholysis was reduced in older adults (32 ± 13 vs. 74 ± 8%; P <0.05). Similarly, α₂-mediated vasoconstriction during exercise was significantly blunted in both young (-15 ± 4% vs. -44 ± 8%) and older adults (-26 ± 3% vs. -35 ± 6%), however the magnitude of sympatholysis was reduced in older adults (19 ± 8% vs. 60 ± 10%; P <0.05). During ATP, both α₁- and α₂-mediated vasoconstriction was nearly abolished in young and older adults (ΔFVC ∼ -5%), and the magnitude of sympatholysis was similar in both age groups (∼85-90%). Our findings indicate that the ability to modulate postjunctional α-adrenergic vasoconstriction during exercise is impaired with age, whereas the sympatholytic effect of exogenous ATP is preserved. Thus, if impairments in vascular control during exercise in older adults involve vasoactive ATP, we speculate that circulating ATP is reduced with advancing age.

Vasodilatory responsiveness to adenosine triphosphate in ageing humans

Endothelium-dependent vasodilatation is reduced with advancing age in humans, as evidenced by blunted vasodilator responsiveness to acetylcholine (ACh). Circulating adenosine triphosphate (ATP) has been implicated in the control of skeletal muscle vascular tone during mismatches in oxygen delivery and demand (e.g. exercise) via binding to purinergic receptors (P2Y) on the endothelium evoking subsequent vasodilatation, and ageing is typically associated with reductions in muscle blood flow under such conditions. Therefore, we tested the hypothesis that ATP-mediated vasodilatation is impaired with age in healthy humans. We measured forearm blood flow (venous occlusion plethysmography) and calculated vascular conductance (FVC) responses to local intra-arterial infusions of ACh, ATP, and sodium nitroprusside (SNP) before and during ascorbic acid (AA) infusion in 13 young and 13 older adults. The peak increase in FVC to ACh was significantly impaired in older compared with young adults (262 ± 71% vs. 618 ± 97%; P < 0.05), and this difference was abolished during AA infusion (510 ± 82% vs. 556 ± 71%; not significant, NS). In contrast, peak FVC responses were not different between older and young adults to either ATP (675 ± 105% vs. 734 ± 126%) or SNP (1116 ± 111% vs. 1138 ± 148%) and AA infusion did not alter these responses in either age group (both NS). In another group of six young and six older adults, we determined whether vasodilator responses to adenosine and ATP were influenced by P1-receptor blockade via aminophylline. The peak FVC responses to adenosine were not different in young (350 ± 65%) versus older adults (360 ± 80%), and aminophylline blunted these responses by ∼50% in both groups. The peak FVC responses to ATP were again not different in young and older adults, and aminophylline did not impact the vasodilatation in either group. Thus, in contrast to the observed impairments in ACh responses, the vasodilatory response to exogenous ATP is not reduced with age in healthy humans. Further, our data also indicate that adenosine mediated vasodilatation is not reduced with age, and that ATP-mediated vasodilatation is independent of P1-receptor stimulation in both young and older adults.

The Role of the Urothelium and ATP in Mediating Detrusor Smooth Muscle Contractility

To examine the contractility of urothelium-intact (+UE) and urothelium-denuded (-UE) rat detrusor strips under adenosine triphosphate (ATP) treatment. Purinergic signaling exists in the bladder but both the inhibitory effect of ATP on detrusor contractions and the function of urothelial ATP are not established. Detrusor strips were obtained from bladders of young adult rats. Isometric tension from both transverse and longitudinal contractions was measured using a myograph. The muscarinic agonist carbachol (CCh) was used to induce contractions, which were under the influences of different concentrations of ATP. In both +UE and -UE strips, 1 mM ATP suppressed CCh-induced contractions. In longitudinal contractions, ATP added to the inhibitory effect of urothelium on CCh responses. Removal of the urothelium, but with exogenous ATP added, recovered the CCh responses to the same level as in +UE strips with no added ATP. Transverse contractions were less susceptible to ATP in the presence of urothelium. We showed that the urothelium and ATP suppressed CCh-induced contractions to a similar extent. The findings suggest an inhibitory role of urothelial ATP in mediating detrusor smooth muscle contractility, which may be impaired in diseased bladders.

Modulation of postjunctional α‐adrenergic vasoconstriction during exercise and exogenous ATP infusions in aging humans

The ability to modulate α‐adrenergic vasoconstriction in contracting muscle is impaired with age. In young adults, adenosine triphosphate (ATP) has been shown to inhibit vasoconstrictor responsiveness similar to exercise. We tested the hypothesis that modulation of postjunctional α‐adrenergic vasoconstriction to exogenous ATP is impaired in aging humans. We measured forearm blood flow (FBF; Doppler ultrasound) and calculated vascular conductance (FVC) to intra‐arterial infusions of phenylephrine (α1‐agonist) and dexmedetomidine (α2‐agonist) during rhythmic handgrip exercise (15% MVC), a control non‐exercise vasodilator condition (adenosine), and ATP infusion in 7 young (22±1) and 7 older (64±3 yrs) adults. Forearm hyperemia was matched across all conditions. During adenosine, forearm vasoconstrictor responses to direct α1‐stimulation were lower in older adults (ΔFVC= −25±3 vs −41±5%); the responses to α2‐stimulation were not different (−38±5% vs −44±7%). The ability to blunt α1‐ and α2‐vasoconstriction was impaired in older adults during exercise (α1 = 31±9 vs 74±8%; α2 = 28±10 vs 60±10%). In contrast, ATP reduced the vasoconstrictor responses by ~85–90% in both age groups. We conclude that the sympatholytic effect of exogenous ATP is not reduced with age, and thus blunted ATP release during exercise may contribute to the impaired exercise sympatholysis in older adults.Supported by HL‐087952