Activation Of A2A Receptors Research Articles

A2A adenosine receptor‐mediated increase in coronary flow is dependent on NADPH oxidase‐derived reactive oxygen species (1071.1)

Adenosine induces an increase in coronary flow (CF) mainly through the activation of both A2A and A2B adenosine receptors (ARs); however, the mechanisms responsible for the regulation of CF are not fully understood. Earlier data from our laboratory showed an interaction between adenosine, NADPH oxidase (NOX) and reactive oxygen species (ROS). We hypothesize that adenosine‐mediated increase in CF is dependent upon NOX activation through A2A but not A2B AR. Concentration response curves to adenosine and the A2A AR selective agonist (CGS‐21680) were performed in isolated perfused hearts from WT, A2A and A2B AR knockouts (A2AKO and A2BKO). Adenosine‐induced dose‐dependent increase in CF was significantly decreased (p<0.05) with pre‐treatment with gp91 ds‐tat (a NOX inhibitor; gp91) or EUK134 (a catalase/superoxide dismutase mimic ; EUK) in both WT (WT: 229.7% ± 6.5 vs. WT+gp91: 178% ± 5.9 and WT: 228.4% ± 7.2 vs. WT+EUK: 178.8% ± 5.5) and A2BKO ( A2BKO: 241.3% ± 9.1 vs. A2BKO+gp91: 191% ± 8.7 and A2BKO: 254% ± 9.3 vs. A2BKO+EUK: 194.4% ± 7.9), but not in A2AKO isolated hearts (P>0.05) (A2AKO: 271.5% ± 17.4 vs. A2AKO+gp91: 240.8% ± 13.6 and A2AKO: 271.2% ± 18.4 vs. A2AKO+EUK: 257.6% ± 15.6). Also, CGS‐21680 caused a concentration‐dependent increase in CF in both WT and A2BKO, but not in A2AKO, which was significantly reduced (P<0.05) using gp91 (WT: 226% ± 8.4 vs. WT+gp91: 191.6% ± 6.6 and A2BKO: 182.6% ± 4.4 vs. A2BKO+gp91: 159.7% ± 4.3). Our data suggest that adenosine‐induced increase in CF through the activation of A2AKO AR is dependent upon NOX activation and ROS generation.Grant Funding Source: Supported by HL 094447, HL 027339 and AI 096139

Adenosine A2a receptors activate Nuclear Factor-Kappa B (NF-κB) in rat hippocampus after exposure to different doses of MDMA

MDMA (3,4-methylenedioxy-N-methamphetamine) as an empathogenic drug causes neurotoxicity although the mechanisms have not been fully elucidated. The A2a adenosine receptor modulates the reinforcement efficacy and neurotoxicity of MDMA. A2a receptor activation inhibits Nuclear Factor-Kappa B (NF-κB) activation and nuclear translocation by a known mechanism. In the present study, we aimed to compare the NF-κB activity level in rat hippocampus between two doses of MDMA exposure (10 and 20 mg /kg) using either A2aR agonist (CGS) or A2aR antagonist (SCH). Adult male Sprague-Dawley rats were subjected to MDMA followed by intraperitoneal CGS (0.03 mg/kg) or SCH (0.03 mg/kg) injection. The hippocampi were then removed for western blot and RTPCR analyses. Administration of MDMA dose-dependently increased the expression of NF-κB both at mRNA and protein levels. We also found that administration of CGS following MDMA significantly increased the NF-κB expression especially in MDMA 20 +CGS group. By contrast, administration of the A2a-R antagonist SCH resulted in a dose-dependent decrease in NF-κB mRNA and protein. Our study results revealed that MDMA has powerful detrimental effects on expression of NF-κB in a dose-dependent manner. On the other hand, co-administration of A2a agonist (CGS) can protect against MDMA neurotoxic effects by increasing NF-κB expression levels; suggesting a potential application for protection against the neurotoxic effects observed in MDMA users.

Antidepressant activity of the adenosine A2A receptor antagonist, istradefylline (KW-6002) on learned helplessness in rats

Istradefylline, an adenosine A2A receptor antagonist, improves motor function in animal models of Parkinson's disease (PD) and in patients with PD. In addition, some A2A antagonists exert antidepressant-like activity in rodent models of depression, such as the forced swim and the tail suspension tests. We have investigated the effect of istradefylline on depression-like behaviors using the rat learned helplessness (LH) model. Acute, as well as chronic, oral administration of istradefylline significantly improved the inescapable shock (IES)-induced escape deficit with a degree of efficacy comparable to chronic treatment with the tricyclic antidepressant desipramine and the selective serotonin (5-HT) reuptake inhibitor, fluoxetine. Both the A1/A2A receptor nonspecific antagonist theophylline and the moderately selective antagonist CGS15943, but not the A1 selective antagonist DPCPX, ameliorated the IES-induced escape deficit. The enhancement of escape response by istradefylline was reversed by a local injection of the A2A specific agonist CGS21680 either into the nucleus accumbens, the caudate-putamen, or the paraventricular nucleus of the hypothalamus, but not by the A1 specific agonist R-PIA into the nucleus accumbens. Moreover, neither the 5-HT2A/2C receptor antagonist methysergide or the adrenergic α 2 antagonist yohimbine, nor the β-adrenergic antagonist propranolol, affected the improvement of escape response induced by istradefylline. Istradefylline exerts antidepressant-like effects via modulation of A2A receptor activity which is independent of monoaminergic transmission in the brain. Istradefylline may represent a novel treatment option for depression in PD as well as for the motor symptoms.

Adenosine’s Autacoid Function in the Central Nervous System and the Behavioral State of Conservation-Withdrawal

The purine nucleoside adenosine has the critical autacoid function of directly linking cellular excitability to energy availability. The mechanism is activated whenever the rate of adenosine triphosphate (ATP) utilization exceeds the rate of synthesis. In CNS neurons, adenosine is produced by the rapid intracellular hydrolysis of purine nucleotides during neural excitation and then is extruded into extracellular space. The nucleoside is also produced by the extracellular hydrolysis of ATP by ectonucleotidases. Extracellular adenosine interacts with G-protein linked stereospecific receptors to reestablish metabolic homeostasis by exerting extraordinarily potent inhibition of neural excitation via a number of mechanisms. This autacoid mechanism is directly linked to the production of a depression like behavioral state termed conservation-withdrawal during times of physical stress or severe emotional distress. We review evidence here that adenosine produces a transition to conservation-withdrawal by activation of A2A receptors in the ventral-medial striatum.

Purinergic Signaling and Hippocampal Long-Term Potentiation

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

Adenosine as a regulator of NFκB activation

Adenosine as a regulator of NF<i>κ</i>B activation

Mechanisms involved in increased sensitivity to adenosine A2A receptor activation and hypoxia-induced vasodilatation in porcine coronary arteries

Hypoxia-induced coronary vasorelaxation is a compensatory mechanism increasing blood flow. We hypothesized that hypoxia shares pathways with adenosine and causes vasorelaxation through the adenosine A2A receptor and force suppression by increasing cAMP and phosphorylated heat shock protein (HSP)20. Adenosine receptors in porcine left anterior descending coronary arteries (LAD) were examined by RT-PCR and isometric tension recording in myographs. Vasorelaxation was induced by adenosine, 1% oxygen, or both in the absence or presence of ZM241385, an adenosine A2A receptor antagonist. cAMP was determined by ELISA and p-HSP20/HSP20 and p-MLC/MLC were determined by immunoblotting and densitometric analyses. In coronary arteries exposed to 1% oxygen, there was increased sensitivity to adenosine, the adenosine A2 selective agonist NECA, and the adenosine A2A selective receptor agonist CGS21680. ZM241385 shifted concentration–response curves for CGS21680 to the right, whereas the adenosine A1 antagonist DPCPX, the adenosine A2B receptor antagonist MRS1754 and the adenosine A3 receptor antagonist MRS1523 failed to reduce vasodilatation induced by CGS21680. 1% oxygen or adenosine increased cAMP accumulation and HSP20 phosphorylation without changing T850-MYPT1 and MLC phosphorylation. ZM241385 failed to change 1% oxygen-induced vasodilation, cAMP accumulation, HSP20 phosphorylation and MLC phosphorylation. The PKA inhibitor Rp-8-CPT-cAMPS significantly reduced vasorelaxation induced by 1% oxygen or CGS21680. Our findings suggest that the increased sensitivity to adenosine, NECA, and CGS21680 at 1% oxygen involves adenosine A2A receptors. Adenosine and 1% oxygen induce vasorelaxation in PGF2α-contracted porcine coronary arteries partly by force suppression caused by increased cAMP and phosphorylation of HSP20.

Adenosine 2A receptor promotes collagen production by human fibroblasts via pathways involving cyclic AMP and AKT but independent of Smad2/3

Activation of adenosine A2A receptor (A2AR) promotes fibrosis and collagen synthesis. However, the underlying mechanism is still unclear, not least because cAMP, its principal effector, has been found to inhibit TGFβ1-induced collagen synthesis. Here, we show that in primary normal human dermal fibroblasts, A2AR stimulation with CGS21680 elicits a modest cAMP increase (150 ± 12% of control; EC50 54.8 nM), which stimulates collagen1 (Col1) and collagen3 (Col3), but maximal cAMP resulting from direct activation of adenylyl cyclase by forskolin (15,689 ± 7038% of control; EC50 360.7 nM) inhibits Col1 and increases Col3. Similar to Col1 expression, fibroblast proliferation increased following physiological cAMP increases by CGS21680 but was inhibited by cAMP increases beyond the physiological range by forskolin. The A2AR-mediated increase of Col1 and Col3 was mediated by AKT, while Col3, but not Col1, expression was dependent on p38 and repressed by ERK. TGFβ1 induced phosphorylation of Smad2/3 and increased Col3 expression, which was prevented by Smad3 depletion. In contrast, CGS21680 did not activate Smad2/3, and Smad2/3 knockdown did not prevent CGS21680-induced Col1 or Col3 increases. Our results indicate that cAMP is a concentration-dependent switch for collagen production via noncanonical, AKT-dependent, Smad2/3-independent signaling. These observations explain the paradoxical effects of cAMP on collagen expression.

NF-κB Is Activated in CD4+ iNKT Cells by Sickle Cell Disease and Mediates Rapid Induction of Adenosine A2A Receptors

Reperfusion injury following tissue ischemia occurs as a consequence of vaso-occlusion that is initiated by activation of invariant natural killer T (iNKT) cells. Sickle cell disease (SDC) results in widely disseminated microvascular ischemia and reperfusion injury as a result of vaso-occlusion by rigid and adhesive sickle red blood cells. In mice, iNKT cell activation requires NF-κB signaling and can be inhibited by the activation of anti-inflammatory adenosine A2A receptors (A2ARs). Human iNKT cells are divided into subsets of CD4+ and CD4- cells. In this study we found that human CD4+ iNKT cells, but not CD4- cells undergo rapid NF-κB activation (phosphorylation of NF-κB on p65) and induction of A2ARs (detected with a monoclonal antibody 7F6-G5-A2) during SCD painful vaso-occlusive crises. These findings indicate that SCD primarily activates the CD4+ subset of iNKT cells. Activation of NF-κB and induction of A2ARs is concordant, i.e. only CD4+ iNKT cells with activated NF-κB expressed high levels of A2ARs. iNKT cells that are not activated during pVOC express low levels of A2AR immunoreactivity. These finding suggest that A2AR transcription may be induced in CD4+ iNKT cells as a result of NF-κB activation in SCD. In order to test this hypothesis further we examined cultured human iNKT cells. In cultured cells, blockade of NF-κB with Bay 11–7082 or IKK inhibitor VII prevented rapid induction of A2AR mRNA and protein upon iNKT activation. In conclusion, NF-κB-mediated induction of A2ARs in iNKT cells may serve as a counter-regulatory mechanism to limit the extent and duration of inflammatory immune responses. As activated iNKT cells express high levels of A2ARs following their activation, they may become highly sensitive to inhibition by A2AR agonists.

Hypoxia and P1 receptor activation regulate the high-affinity concentrative adenosine transporter CNT2 in differentiated neuronal PC12 cells

Under several adverse conditions, such as hypoxia or ischaemia, extracellular levels of adenosine are elevated because of increased energy demands and ATP metabolism. Because extracellular adenosine affects metabolism through G-protein-coupled receptors, its regulation is of high adaptive importance. CNT2 (concentrative nucleoside transporter 2) may play physiological roles beyond nucleoside salvage in brain as it does in other tissues. Even though nucleoside transport in brain has mostly been seen as being of equilibrative-type, in the present study, we prove that the rat phaeochromocytoma cell line PC12 shows a concentrative adenosine transport of CNT2-type when cells are differentiated to a neuronal phenotype by treatment with NGF (nerve growth factor). Differentiation of PC12 cells was also associated with the up-regulation of adenosine A1 receptors. Addition of adenosine receptor agonists to cell cultures increased CNT2-related activity by a mechanism consistent with A₁ and A2A receptor activation. The addition of adenosine to the culture medium also induced the phosphorylation of the intracellular regulatory kinase AMPK (AMP-activated protein kinase), with this effect being dependent upon adenosine transport. CNT2-related activity of differentiated PC12 cells was also dramatically down-regulated under hypoxic conditions. Interestingly, the analysis of nucleoside transporter expression after experimental focal ischaemia in rat brain showed that CNT2 expression was down-regulated in the infarcted tissue, with this effect somehow being restricted to other adenosine transporter proteins such as CNT3 and ENT1 (equilibrative nucleoside transporter 1). In summary, CNT2 is likely to modulate extracellular adenosine and cell energy balance in neuronal tissue.

Adenosine A2A Receptor Contributes to Ischemic Brain Damage in Newborn Piglet

Pharmacologic inactivation or genetic deletion of adenosine A2A receptors protects ischemic neurons in adult animals, but studies in neonatal hypoxia-ischemia (H-I) are inconclusive. The present study in neonatal piglets examined the hypothesis that A2A receptor signaling after reoxygenation from global H-I contributes to injury in highly vulnerable striatal neurons where A2A receptors are enriched. A2A receptor immunoreactivity was detected in striatopallidal neurons. In nonischemic piglets, direct infusion of the selective A2A receptor agonist CGS 21680 through microdialysis probes into putamen increased phosphorylation of N-methyl-D-aspartic acid (NMDA) receptor NR1 subunit and Na(+),K(+)-ATPase selectively at protein kinase A (PKA)-sensitive sites. In ischemic piglets, posttreatment with SCH 58261, a selective A2A receptor antagonist, improved early neurologic recovery and preferentially protected striatopallidal neurons. SCH 58261 selectively inhibited the ischemia-induced phosphorylation of NR1, Na(+),K(+)-ATPase, and cAMP-regulated phosphoprotein 32 KDa (DARPP32) at PKA-sensitive sites at 3 hours of recovery and improved Na(+),K(+)-ATPase activity. SCH 58261 also suppressed ischemia-induced protein nitration and oxidation. Thus, A2A receptor activation during reoxygenation contributes to the loss of a subpopulation of neonatal putamen neurons after H-I. Its toxic signaling may be related to DARPP32-dependent phosphorylation of PKA-sensitive sites on NR1 and Na(+),K(+)-ATPase, thereby augmenting excitotoxicity-induced oxidative stress after reoxygenation.

Beneficial effects of a novel agonist of the adenosine A2A receptor on monocrotaline‐induced pulmonary hypertension in rats

Pulmonary arterial hypertension (PAH) is characterized by enhanced pulmonary vascular resistance, right ventricular hypertrophy and increased right ventricular systolic pressure. Here, we investigated the effects of a N-acylhydrazone derivative, 3,4-dimethoxyphenyl-N-methyl-benzoylhydrazide (LASSBio-1359), on monocrotaline (MCT)-induced pulmonary hypertension in rats. PAH was induced in male Wistar rats by a single i.p. injection of MCT (60 mg·kg(-1)) and 2 weeks later, oral LASSBio-1359 (50 mg·kg(-1)) or vehicle was given once daily for 14 days. Echocardiography was used to measure cardiac function and pulmonary artery dimensions, with histological assay of vascular collagen. Studies of binding to human recombinant adenosine receptors (A1, A2A, A3) and of docking with A2A receptors were also performed. MCT administration induced changes in vascular and ventricular structure and function, characteristic of PAH. These changes were reversed by treatment with LASSBio-1359. MCT also induced endothelial dysfunction in pulmonary artery, as measured by diminished relaxation of pre-contracted arterial rings, and this dysfunction was reversed by LASSBio-1359. In pulmonary artery rings from normal Wistar rats, LASSBio-1359 induced relaxation, which was decreased by the adenosine A2A receptor antagonist, ZM 241385. In adenosine receptor binding studies, LASSBio-1359 showed most affinity for the A2A receptor and in the docking analyses, binding modes of LASSBio-1359 and the A2A receptor agonist, CGS21680, were very similar. In rats with MCT-induced PAH, structural and functional changes in heart and pulmonary artery were reversed by treatment with oral LASSBio-1359, most probably through the activation of adenosine A2A receptors.

Long-Term Depression of Synaptic Kainate Receptors Reduces Excitability by Relieving Inhibition of the Slow Afterhyperpolarization

Kainate receptors (KARs) are ionotropic glutamate receptors that also activate noncanonical G-protein-coupled signaling pathways to depress the slow afterhyperpolarization (sAHP). Here we show that long-term depression of KAR-mediated synaptic transmission (KAR LTD) at rat hippocampal mossy fiber synapses relieves inhibition of the sAHP by synaptic transmission. KAR LTD is induced by high-frequency mossy fiber stimulation and natural spike patterns and requires activation of adenosine A2A receptors. Natural spike patterns also cause long-term potentiation of NMDA receptor-mediated synaptic transmission that overrides the effects of KAR LTD on the cellular response to low-frequency synaptic input. However, KAR LTD is dominant at higher frequency synaptic stimulation where it decreases the cellular response by relieving inhibition of the sAHP. Thus we describe a form of glutamate receptor plasticity induced by natural spike patterns whose primary physiological function is to regulate cellular excitability.

Smoke Extract Impairs Adenosine Wound Healing. Implications of Smoke-Generated Reactive Oxygen Species

Adenosine concentrations are elevated in the lungs of patients with asthma and chronic obstructive pulmonary disease, where it balances between tissue repair and excessive airway remodeling. We previously demonstrated that the activation of the adenosine A2A receptor promotes epithelial wound closure. However, the mechanism by which adenosine-mediated wound healing occurs after cigarette smoke exposure has not been investigated. The present study investigates whether cigarette smoke exposure alters adenosine-mediated reparative properties via its ability to induce a shift in the oxidant/antioxidant balance. Using an in vitro wounding model, bronchial epithelial cells were exposed to 5% cigarette smoke extract, were wounded, and were then stimulated with either 10 μM adenosine or the specific A2A receptor agonist, 5'-(N-cyclopropyl)-carboxamido-adenosine (CPCA; 10 μM), and assessed for wound closure. In a subset of experiments, bronchial epithelial cells were infected with adenovirus vectors encoding human superoxide dismutase and/or catalase or control vector. In the presence of 5% smoke extract, significant delay was evident in both adenosine-mediated and CPCA-mediated wound closure. However, cells pretreated with N-acetylcysteine (NAC), a nonspecific antioxidant, reversed smoke extract-mediated inhibition. We found that cells overexpressing mitochondrial catalase repealed the smoke extract inhibition of CPCA-stimulated wound closure, whereas superoxide dismutase overexpression exerted no effect. Kinase experiments revealed that smoke extract significantly reduced the A2A-mediated activation of cyclic adenosine monophosphate-dependent protein kinase. However, pretreatment with NAC reversed this effect. In conclusion, our data suggest that cigarette smoke exposure impairs A2A-stimulated wound repair via a reactive oxygen species-dependent mechanism, thereby providing a better understanding of adenosine signaling that may direct the development of pharmacological tools for the treatment of chronic inflammatory lung disorders.

The antidepressant-like effect of inosine in the FST is associated with both adenosine A1 and A 2A receptors.

Inosine is an endogenous purine nucleoside, which is formed during the breakdown of adenosine. The adenosinergic system was already described as capable of modulating mood in preclinical models; we now explored the effects of inosine in two predictive models of depression: the forced swim test (FST) and tail suspension test (TST). Mice treated with inosine displayed higher anti-immobility in the FST (5 and 50 mg/kg, intraperitoneal route (i.p.)) and in the TST (1 and 10 mg/kg, i.p.) when compared to vehicle-treated groups. These antidepressant-like effects started 30 min and lasted for 2 h after intraperitoneal administration of inosine and were not accompanied by any changes in the ambulatory activity in the open-field test. Both adenosine A1 and A2A receptor antagonists prevented the antidepressant-like effect of inosine in the FST. In addition, the administration of an adenosine deaminase inhibitor (1 and 10 mg/kg, i.p.) also caused an antidepressant-like effect in the FST. These results indicate that inosine possesses an antidepressant-like effect in the FST and TST probably through the activation of adenosine A1 and A2A receptors, further reinforcing the potential of targeting the purinergic system to the management of mood disorders.

Adenosine A2A receptor-dependent proliferation of pulmonary endothelial cells is mediated through calcium mobilization, PI3-kinase and ERK1/2 pathways

Hypoxia and HIF-2α-dependent A2A receptor expression and activation increase proliferation of human lung microvascular endothelial cells (HLMVECs). This study was undertaken to investigate the signaling mechanisms that mediate the proliferative effects of A2A receptor. A2A receptor-mediated proliferation of HLMVECs was inhibited by intracellular calcium chelation, and by specific inhibitors of ERK1/2 and PI3-kinase (PI3K). The adenosine A2A receptor agonist CGS21680 caused intracellular calcium mobilization in controls and, to a greater extent, in A2A receptor-overexpressing HLMVECs. Adenoviral-mediated A2A receptor overexpression as well as receptor activation by CGS21680 caused increased PI3K activity and Akt phosphorylation. Cells overexpressing A2A receptor also manifested enhanced ERK1/2 phosphorylation upon CGS21680 treatment. A2A receptor activation also caused enhanced cAMP production. Likewise, treatment with 8Br-cAMP increased PI3K activity. Hence A2A receptor-mediated cAMP production and PI3K and Akt phosphorylation are potential mediators of the A2A-mediated proliferative response of HLMVECs. Cytosolic calcium mobilization and ERK1/2 phosphorylation are other critical effectors of HLMVEC proliferation and growth. These studies underscore the importance of adenosine A2A receptor in activation of survival and proliferative pathways in pulmonary endothelial cells that are mediated through PI3K/Akt and ERK1/2 pathways.

Cellular signalling pathways mediating dilation of porcine pial arterioles to adenosine A2A receptor activation

Adenosine is a potent vasodilator contributing to cerebral blood flow regulation during metabolic stress. However, the distribution of adenosine receptor subtypes and underlying signalling mechanisms for dilation of pial arterioles remain unclear. The present study aimed at addressing these issues. Isolated porcine pial arterioles were subjected to study of vasomotor function, localization of adenosine receptors, and production of nitric oxide (NO). Concentration-dependent vasodilation to adenosine was inhibited by A₂A receptor antagonist ZM241385 but not by A₁ receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine. A₂A receptors were detected in endothelium and smooth muscle of pial arterioles via immunohistochemistry. Adenosine significantly increased arteriolar production of NO, and the induced dilation was insensitive to KATP channel blocker glibenclamide but was attenuated by endothelial denudation, NO synthase inhibitor L-NAME, or guanylyl cyclase inhibitor ODQ in a similar manner. Both inward rectifier potassium (Kir) channel inhibitor barium and cAMP signalling inhibitor Rp-8-Br-cAMPS attenuated adenosine-induced dilation. In the presence of L-NAME or the absence of endothelium, addition of Rp-8-Br-cAMPS but not barium further reduced adenosine-induced responses. Barium diminished endothelium-independent vasodilation to NO donor sodium nitroprusside. Comparable to the adenosine-induced response, vasodilation to A₂A receptor agonist CGS21680 was attenuated by endothelial removal, ZM241385, L-NAME, barium, or Rp-8-Br-cAMPS, but not by glibenclamide. Adenosine evokes dilation of porcine pial arterioles via parallel activation of endothelial and smooth muscle A₂A receptors. Stimulation of endothelial NO production activates smooth muscle guanylyl cyclase for vasodilation by opening Kir channels. Adenosine also activates smooth muscle cAMP signalling leading to vasodilation.

Plasma glutamate–modulated interaction of A2AR and mGluR5 on BMDCs aggravates traumatic brain injury–induced acute lung injury

The bone marrow-derived cell (BMDC)-associated inflammatory response plays a key role in the development of acute lung injury (ALI). Activation of adenosine A2A receptor (A2AR) is generally considered to be antiinflammatory, inhibiting BMDC activities to protect against ALI. However, in the present study, we found that in a mouse model of neurogenic ALI induced by severe traumatic brain injury (TBI), BMDC A2AR exerted a proinflammatory effect, aggravating lung damage. This is in contrast to the antiinflammatory effect observed in the mouse oleic acid-induced ALI model (a nonneurogenic ALI model.) Moreover, the A2AR agonist CGS21680 aggravated, whereas the antagonist ZM241385 attenuated, the severe TBI-induced lung inflammatory damage in mice. Further investigation of white blood cells isolated from patients or mouse TBI models and of cultured human or mouse neutrophils demonstrated that elevated plasma glutamate after severe TBI induced interaction between A2AR and the metabotropic glutamate receptor 5 (mGluR5) to increase phospholipase C-protein kinase C signaling, which mediated the proinflammatory effect of A2AR. These results are in striking contrast to the well-known antiinflammatory and protective role of A2AR in nonneurogenic ALI and indicate different therapeutic strategies should be used for nonneurogenic and neurogenic ALI treatment when targeting A2AR.

Possible involvement of A2A and A3 receptors in modulation of insulin secretion and β-cell survival in mouse pancreatic islets

Adenosine A1, A2A, A2B and A3 receptor mRNAs were found to be expressed in mouse pancreatic islets and Beta-TC6 cells but their physiological or pharmacological actions are not fully clarified. We showed that adenosine (100μM) augmented insulin secretion by islets in the presence of either normal (5.5mM) or a high concentration of glucose (20mM). The augmentation of insulin secretion in the presence of high glucose was blocked by an A2A antagonist, but not by A2B and A3 antagonists, while an A1 antagonist potentiated the adenosine effect. An adenosine analogue 5′-N-ethylcarboxamidoadenosine (NECA) as well as A1, A2A and A3 receptor agonists also produced stimulation. On the other hand, an A3 agonist markedly reduced Beta-TC6 cell proliferation and the islet cell viability, while adenosine and NECA did not. The effect of A3 agonist was partially blocked by the A3 antagonist. In addition, treatment with the A3 agonist produced a small but significant extent of apoptosis in Beta-TC6 cells as judged by terminal transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay. These results combined together suggested that like the A1 receptor, activation of A2A receptors by adenosine results in augmented insulin secretion, while the A3 receptor is involved in modulation of the survival of pancreatic β-cells.

Adenosine A2A receptor activation reduces recurrence and mortality from Clostridium difficileinfection in mice following vancomycin treatment

BackgroundActivation of the A2A adenosine receptor (A2AAR) decreases production of inflammatory cytokines, prevents C. difficile toxin A-induced enteritis and, in combination with antibiotics, increases survival from sepsis in mice. We investigated whether A2AAR activation improves and A2AAR deletion worsens outcomes in a murine model of C. difficile (strain VPI10463) infection (CDI).MethodsC57BL/6 mice were pretreated with an antibiotic cocktail prior to infection and then treated with vancomycin with or without an A2AAR agonist. A2AAR-/- and littermate wild-type (WT) mice were similarly infected, and IFNγ and TNFα were measured at peak of and recovery from infection.ResultsInfected, untreated mice rapidly lost weight, developed diarrhea, and had mortality rates of 50-60%. Infected mice treated with vancomycin had less weight loss and diarrhea during antibiotic treatment but mortality increased to near 100% after discontinuation of antibiotics. Infected mice treated with both vancomycin and an A2AAR agonist, either ATL370 or ATL1222, had minimal weight loss and better long-term survival than mice treated with vancomycin alone. A2AAR KO mice were more susceptible than WT mice to death from CDI. Increases in cecal IFNγ and blood TNFα were pronounced in the absence of A2AARs.ConclusionIn a murine model of CDI, vancomycin treatment resulted in reduced weight loss and diarrhea during acute infection, but high recurrence and late-onset death, with overall mortality being worse than untreated infected controls. The administration of vancomycin plus an A2AAR agonist reduced inflammation and improved survival rates, suggesting a possible benefit of A2AAR agonists in the management of CDI to prevent recurrent disease.