2-(4-(2-carboxyethyl)phenethylamino)-5--n-ethylcarboxamidoadenosine and 8-(3-chlorostyryl)caffeine

In this experiment, we examined the influence of the posterior hypothalamic adenosine A(2A) receptors on the central cardiovascular regulation of blood pressure (BP) and heart rate (HR). Posterior hypothalamic injection of drugs was performed in anesthetized, artificially ventilated male Sprague-Dawley rats. Injection of CGS-21680HCl (CGS; 20 nmol), an adenosine A(2A) receptor agonist, elicited a decrease of arterial BP and HR, while injection of 8-(3-Chlorostyryl)caffeine (CSC; 10 nmol), an adenosine A(2A) receptor antagonist, blocked the depressor and bradycardiac effects of CGS (20 nmol). To examine the mechanisms of cardiovascular regulation of adenosine A(2A) receptors in the posterior hypothalamus, we applied the adenylate cyclase and guanylate cyclase inhibitors, to the posterior hypothalamus. Pretreatment with MDL-12,330 (MDL; 10 nmol), an adenylate cylase inhibitor, attenuated the depressor and bradycardiac effects of CGS. However, pretreatment with, LY-83,583 (LY; 5 nmol), a soluble guanylate cyclase inhibitor, did not alter the effects of CGS. Additionally, we examined the modification of the cardiovascular effects of adenosine A(2A) receptors through the ATP-sensitive K+ channel in the posterior hypothalamus. Posterior hypothalamic administration of glipizide (20 nmol) significantly attenuated the cardiovascular depressor actions elicited by CGS. These results suggest that adenosine A(2A) receptors in the posterior hypothalamus play an inhibitory role in central cardiovascular regulation, and that adenylate cyclase, but not guanylate cyclase, mediates the depressor and bradycardiac actions of adenosine A(2A) receptors. Furthermore, ATP-sensitive K+ channels mediate the posterior hypothalamic cardiovascular regulation of adenosine A(2A) receptors.

Topics: Adenosine; Adenosine A2 Receptor Agonists; Adenosine A2 Receptor Antagonists; Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Aminoquinolines; Anesthesia, Inhalation; Animals; Blood Pressure; Bradycardia; Caffeine; Glipizide; Guanylate Cyclase; Heart Rate; Hypothalamus, Posterior; Imines; KATP Channels; Male; Microinjections; Phenethylamines; Rats; Rats, Sprague-Dawley; Receptor, Adenosine A2A

We used the patch-clamp technique to examine the effect of adenosine on the basolateral K channels in the thick ascending limb (TAL) of the rat kidney. A 50-pS inwardly rectifying K channel was detected in the basolateral membrane, and the channel activity was decreased by hyperpolarization. Application of adenosine (10 microM) increased the activity of basolateral 50 pS K channels, defined by NP(o), from 0.21 to 0.41. The effect of adenosine on the 50 pS K channels was mimicked by cyclohexyladenosine (CHA), which increased channel activity by a dose-dependent manner. However, inhibition of the A1 adenosine receptor with 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX) failed to block the effect of CHA. In contrast, application of 8-(3-chlorostyryl) caffeine (CSC), an A2 adenosine antagonist, abolished the stimulatory effect of CHA. The possibility that the effect of adenosine and adenosine analog on the basolateral 50 pS K channel was the result of activation of the A2 adenosine receptor was also suggested by the observation that application of CGS-21680, a selected A(2A) adenosine receptor agonist, increased the channel activity. Also, inhibition of PKA with N-[2-(methylamino)ethyl]-5-isoquinoline sulfonamide-2HC1 abolished the stimulatory effect of CHA on the basolateral 50 pS K channel. Moreover, addition of the membrane-permeable cAMP analog increases the activity of 50 pS K channels. We conclude that adenosine activates the 50 pS K channel in the basolateral membrane of the TAL and the stimulatory effect is mainly mediated by a PKA-dependent pathway via the A2 adenosine receptor in the TAL.

Topics: Adenosine; Adenosine A2 Receptor Antagonists; Adenylyl Cyclases; Algorithms; Animals; Bucladesine; Caffeine; Dose-Response Relationship, Drug; Enzyme Activation; Female; Kidney; Male; Patch-Clamp Techniques; Phenethylamines; Potassium Channels; Rats; Rats, Sprague-Dawley; Receptors, Adenosine A2; Stimulation, Chemical; Xanthines

The present study was performed to clarify the mechanism of change in intraocular pressure by 2-(1-hexyn-1-yl)adenosine (2-H-Ado), a selective adenosine A2 receptor agonist, in rabbits. 2-H-Ado (0.1%, 50 microl)-induced ocular hypertension (E(max): 7.7 mm Hg) was inhibited by an adenosine A2A receptor antagonist 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine, ATP-sensitive K+ channel blocker glibenclamide or 5-hydroxydecanoic acid, but not by an adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine, an adenosine A2B receptor antagonist alloxazine or a cyclooxygenase inhibitor indomethacin. The outflow facility induced by 2-H-Ado seems to be independent of increase in intraocular pressure or ATP-sensitive K+ channel. In contrast, the recovery rate in intraocular pressure decreased by hypertonic saline was accelerated by 2-H-Ado, and this response was dependent on ATP-sensitive K+ channel. These results suggest that 2-H-Ado-induced ocular hypertension is mediated via K+ channel opening through adenosine A2A receptor, and this is probably due to aqueous formation, but independent of change in outflow facility or prostaglandin production.

Topics: Adenosine; Adenosine A2 Receptor Agonists; Adenosine A2 Receptor Antagonists; Alkynes; Animals; Antihypertensive Agents; Caffeine; Decanoic Acids; Glyburide; Hydroxy Acids; Hypotonic Solutions; Intraocular Pressure; Male; Ocular Hypertension; Phenethylamines; Pinacidil; Potassium Channel Blockers; Potassium Channels; Rabbits; Receptor, Adenosine A2A; Sodium Chloride; Time Factors; Xanthines

This study was performed to determine whether intravitreal or intravenous adenosine can alter the microcirculation in the optic nerve head (ONH) of rabbits. Capillary blood flow in the ONH was measured serially with a laser speckle tissue analyser for 2 hr after the intravitreal (0.1, 1.0 and 10 nmol) or intravenous (0.2 and 0.6 mg kg(-1)min) injections of adenosine. In addition, the effect of specific adenosine A(1) and A(2a) antagonists and an adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channel blockers on the adenosine-induced changes on the ONH blood flow was analysed. Intravitreal adenosine increased the capillary blood flow in the ONH in a dose-dependent manner, while intravenous adenosine had no effect. Co-administration of the specific adenosine A(1) receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, 10 nmol) significantly suppressed (P=0.006, ANOVA) the increase in the ONH blood flow induced by adenosine (10 nmol). The specific A(2a) receptor antagonist, 8-(3-chlorostyryl) caffeine (CSC, 10 nmol), had a weak effect in inhibiting the increase but the change was not significant (P=0.08, ANOVA). Both specific A(1) and A(2a) receptor agonists, N(6)-cyclopentyladenosine (CPA, 10 nmol) and 2-p-(2-carboxyethyl) phenethyl-amino-5'-N-ethylcarboxamidoadenosine (CGS-21680, 10 nmol), increased the ONH tissue blood flow (P<0.01, ANOVA). Glibenclamide (10 nmol), a selective K(ATP) channels antagonist, suppressed the increase of ONH blood flow induced by 10 nmol adenosine significantly (P<0.001, ANOVA). On the other hand, 10 nmol of 8-Br-cAMP, a cAMP analog, failed to enhance the capillary blood flow in the ONH. These results indicate that adenosine increases the capillary blood flow in the ONH of rabbits, and it acts through A(1) and A(2a) receptors from the ablumenal side where pericytes are located. Activation of K(ATP) channels is strongly related to the mechanism of adenosine-induced increase in ONH blood flow, while the participation of adenylate cyclase is less likely.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Adenosine; Adenosine A1 Receptor Antagonists; Adenylyl Cyclase Inhibitors; Animals; Caffeine; Capillaries; Dose-Response Relationship, Drug; Glyburide; Injections; Injections, Intravenous; Optic Disk; Phenethylamines; Potassium Channels; Purinergic P1 Receptor Agonists; Purinergic P2 Receptor Antagonists; Rabbits; Regional Blood Flow; Stimulation, Chemical; Vasodilation; Vitreous Body; Xanthines

Adenosine is an immunosuppressive molecule that is associated with the microenvironment of solid tumors. Mouse T cells activated with anti-CD3 antibody in the presence of adenosine with or without coformycin (to prevent adenosine breakdown by adenosine deaminase) exhibited decreased tyrosine phosphorylation of some intracellular proteins and were inhibited in their ability to proliferate and synthesize interleukin (IL)-2. In addition, adenosine interfered with activation-induced expression of the co-stimulatory molecules CD2 and CD28. Activation-induced CD2 and CD28 expression was also diminished when T cells were activated in the presence of anti-IL-2 and anti-CD25 antibodies to neutralize IL-2 bioactivity. Collectively, these data suggest that CD2 and CD28 up-regulation following T cell activation is IL-2-dependent; and that adenosine inhibits activation-induced T cell expression of CD2 and CD28 by interfering with IL-2-dependent signaling. The inhibitory effect of adenosine on activation-induced CD2 and CD28 expression could not be attributed to cyclic AMP (cAMP) accumulation resulting from the stimulation of adenylyl cyclase-coupled adenosine receptors, even though cAMP at concentrations much higher than those generated following adenosine stimulation was inhibitory for both CD2 and CD28 expression. We conclude that adenosine interferes with IL-2-dependent T cell expression of co-stimulatory molecules via a mechanism that does not involve the accumulation of intracellular cAMP.

Topics: Adenosine; Animals; Antibodies, Monoclonal; Caffeine; CD2 Antigens; CD28 Antigens; CD3 Complex; Cell Cycle; Colforsin; Cyclic AMP; Female; Flavins; Interleukin-2; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Phenethylamines; Receptors, Interleukin-2; Receptors, Purinergic P1; Signal Transduction; T-Lymphocytes; Tyrosine

To assess the impact of ischemia-reperfusion (I/R) on coronary function, and the role of endogenous adenosine in modifying post-ischemic vascular function in asanguinous hearts.. Vascular function was studied in Langendorff perfused mouse hearts subjected to 20-25-min ischemia and 30-min reperfusion.. Ischemia altered the dependence of flow on work-rate observed in normoxic hearts, and inhibited reflow by mechanisms additional to diastolic compression. Coronary responses were selectively reduced: 2-chloroadenosine and ADP dilated with pEC(50)s of 8.4+/-0.1 and 7.4+/-0.1 in non-ischemic hearts versus 7.7+/-0.1 and 7.1+/-0.1 after 20-min ischemia (P<0.05). Sensitivity was further reduced after 25-min ischemia. Responses to nitroprusside were unaltered. NO-synthase antagonism (50 microM nitro-L-arginine methylester) reduced sensitivities to 2-chloroadenosine and ADP up to 10-fold, and eliminated inhibitory effects of I/R. K(ATP) blockade with 5 microM glibenclamide impaired sensitivity pre- and post-ischemia, not eliminating the inhibitory effects of I/R. A(1) adenosine receptor antagonism with 100 nM 8-cyclopentyl-1,3-dipropylxanthine worsened effects of ischemia on sensitivity. A(2A) adenosine receptor antagonism with 100 nM 8-(3-chlorostyryl)caffeine reduced post-ischemic flow by 50%, yet paradoxically enhanced post-ischemic contractile recovery.. Ischemia modifies vascular control and impairs NO- versus K(ATP)-dependent coronary dilation in an asanguinous model. Endogenous adenosine protects against vascular dysfunction via A(1) receptors, and determines coronary reflow via A(2A) receptors. However, intrinsic A(2A) activation apparently worsens contractile dysfunction.

Topics: 2-Chloroadenosine; Adenosine; Adenosine Diphosphate; Animals; Caffeine; Coronary Circulation; Heart; Male; Mice; Mice, Inbred C57BL; Models, Animal; Myocardial Contraction; Myocardial Reperfusion Injury; Nitroprusside; Perfusion; Phenethylamines; Purinergic P1 Receptor Antagonists; Vasodilator Agents; Xanthines

The A(2A)R is largely coexpressed with D(2)Rs and enkephalin mRNA in the striatum where it modulates dopaminergic activity. Activation of the A(2A)R antagonizes D(2)R-mediated behavioral and neurochemical effects in the basal ganglia through a mechanism that may involve direct A(2A)R-D(2)R interaction. However, whether the D(2)R is required for the A(2A)R to exert its neural function is an open question. In this study, we examined the role of D(2)Rs in A(2A)R-induced behavioral and cellular responses, by using genetic knockout (KO) models (mice deficient in A(2A)Rs or D(2)Rs or both). Behavioral analysis shows that the A(2A)R agonist 2-4-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine reduced spontaneous as well as amphetamine-induced locomotion in both D(2) KO and wild-type mice. Conversely, the nonselective adenosine antagonist caffeine and the A(2A)R antagonist 8-(3-chlorostyryl)caffeine produced motor stimulation in mice lacking the D(2)R, although the stimulation was significantly attenuated. At the cellular level, A(2A)R inactivation counteracted the increase in enkephalin expression in striatopallidal neurons caused by D(2)R deficiency. Consistent with the D(2) KO phenotype, A(2A)R inactivation partially reversed both acute D(2)R antagonist (haloperidol)-induced catalepsy and chronic haloperidol-induced enkephalin mRNA expression. Together, these results demonstrate that A(2A)Rs elicit behavioral and cellular responses despite either the genetic deficiency or pharmacological blockade of D(2)Rs. Thus, A(2A)R-mediated neural functions are partially independent of D(2)Rs. Moreover, endogenous adenosine acting at striatal A(2A)Rs may be most accurately viewed as a facilitative modulator of striatal neuronal activity rather than simply as an inhibitory modulator of D(2)R neurotransmission.

Topics: Adenosine; Amphetamines; Animals; Caffeine; Catalepsy; Corpus Striatum; Dopamine Antagonists; Enkephalins; Gene Expression; Haloperidol; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Activity; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Receptor, Adenosine A2A; Receptors, Dopamine D1; Receptors, Dopamine D2; Receptors, Purinergic P1; RNA, Messenger

Adenosine (ADO) exerts potent anti-inflammatory and immunosuppressive effects. In this paper we address the possibility that these effects are partly mediated by inhibition of the secretion of IL-12, a proinflammatory cytokine and a major inducer of Th1 responses. We demonstrate that 5'-N-ethylcarboxamidoadenosine (NECA), a nonspecific ADO analogue, and 2-p-(2-carbonyl-ethyl)phenylethylamino-5'-N-ethylcarboxamidoadenos ine (CGS-21680), a specific A2a receptor agonist, dose-dependently inhibited, in whole blood ex vivo and monocyte cultures, the production of human IL-12 induced by LPS and Stapholococcus aureus Cowan strain 1. However, the A1 receptor agonist 2-Chloro-N6-cyclopentyladenosine and the A3 receptor agonists N6-Benzyl-NECA and 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-be ta-d -ribofuranuronamide expressed only weak inhibitory effects. On the other hand, NECA and CGS-21680 dose-dependently potentiated the production of IL-10. The differential effect of these drugs on monocyte IL-12 and IL-10 production implies that these effects are mediated by A2a receptor signaling rather than by intracellular toxicity of ADO analogue's metabolites. Moreover, CGS-21680 inhibited IL-12 production independently of endogenous IL-10 induction, because anti-IL-10 Abs failed to prevent its effect. The selective A2a antagonist 8-(3-Chlorostyryl) caffeine prevented the inhibitory effect of CGS-21680 on IL-12 production. The phosphodiesterase inhibitor Ro 20-1724 dose-dependently potentiated the inhibitory effect of CGS-21680 and, furthermore, Rp-cAMPS, a protein kinase A inhibitor, reversed the inhibitory effect of CGS-21680, implicating a cAMP/protein kinase A pathway in its action. Thus, ligand activation of A2a receptors simultaneously inhibits IL-12 and stimulates IL-10 production by human monocytes. Through this mechanism, ADO released in excess during inflammatory and ischemic conditions, or tissue injury, may contribute to selective suppression of Th1 responses and cellular immunity.

Topics: Adenosine; Adenosine-5'-(N-ethylcarboxamide); Caffeine; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dose-Response Relationship, Immunologic; Female; Humans; Immunosuppressive Agents; Interleukin-10; Interleukin-12; Ligands; Lipopolysaccharides; Male; Monocytes; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Receptor, Adenosine A2A; Receptor, Adenosine A3; Receptors, Purinergic P1; Signal Transduction

Adenosine (Ado) is an important autocrine modulator of neutrophil functions. In this study, we determined the effects of endogenous Ado on fMet-Leu-Phe (fMLP)-induced phospholipase D (PLD) activity in neutrophils. The removal of extracellular Ado by Ado deaminase (ADA) or the blockade of its action by the A2a receptor antagonists 8-(3-chlorostyryl) caffeine (CSC) or CGS15943 markedly increased fMLP-induced PLD activation. The concentration-dependent stimulatory effects of CSC and CGS15943 were abolished by a pretreatment of neutrophil suspensionswith ADA. In contrast, the selective A2a receptor agonist CGS21680 suppressed fMLP-induced PLD activation. Furthermore, inhibition by CGS21680 of fMLP-induced PLD activity was reversed by CSC or CGS15943. The removal of Ado by ADA or the blockade of its action by CSC or CGS15943, markedly increased the membrane recruitment of cytosolic protein kinase Calpha (PKCalpha), RhoA, and ADP-ribosylation factor (ARF) in response to fMLP. As shown for PLD activity, the stimulatory effect of Ado receptor antagonists on PLD cofactors translocation was abolished by a pretreatment of the cells with ADA. Moreover, the membrane translocation of both PKCalpha, RhoA, and ARF in response to fMLP was attenuated by CGS21680 and this effect of the A2a receptor agonist was antagonized by CSC or CGS15943. These data demonstrate that Ado released by neutrophils in the extracellular milieu inhibits PLD activation by blocking membrane association of ARF, RhoA, and PKCalpha through Ado A2a receptor occupancy. (Blood. 2000;95:519-527)

Topics: Adenosine; Adenosine Deaminase; Adenosine-5'-(N-ethylcarboxamide); ADP-Ribosylation Factor 1; Adult; Caffeine; Cell Membrane; Enzyme Activation; GTP Phosphohydrolases; Humans; In Vitro Techniques; Isoenzymes; Kinetics; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Phenethylamines; Phospholipase D; Protein Kinase C; Protein Kinase C-alpha; Purinergic P1 Receptor Antagonists; Quinazolines; Receptor, Adenosine A2A; Receptors, Purinergic P1; rhoA GTP-Binding Protein; Triazoles

Cholinergic neurons were identified in rat striatal slices by their size, membrane properties, sensitivity to the NK(1) receptor agonist (Sar(9), Met(O(2))(11)) Substance P, and expression of choline acetyltransferase mRNA. A(1) receptor mRNA was detected in 60% of the neurons analysed, and A(2A) receptor mRNA in 67% (n=15). The A(1) receptor agonist R-N(6)-(2-phenylisopropyl)adenosine (R-PIA) hyperpolarized cholinergic neurons in a concentration dependent manner sensitive to the A(1) antagonist 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX, 100 nM). In dual stimulus experiments, the A(2A) receptor antagonist 8-(3-chlorostyryl)caffeine (CSC, 500 nM) decreased release of [(3)H]-acetylcholine from striatal slices (S2/S1 0.78+/-0.07 versus 0.95+/-0.05 in control), as did adenosine deaminase (S2/S1 ratio 0.69+/-0.05), whereas the A(1) receptor antagonist DPCPX (100 nM) had no effect (S2/S1 1.05+/-0.14). In the presence of adenosine deaminase the adenosine A(2A) receptor agonist 2-p-((carboxyethyl)phenylethylamino)-5'-N-ethylcarboxamidoadeno sin e (CGS21680, 10 nM) increased release (S2/S1 ratio 1.03+/-0.05 versus 0.88+/-0.05 in control), an effect blocked by the antagonist CSC (500 nM, S2/S1 0.68+/-0.05, versus 0.73+/-0.08 with CSC alone). The combined superfusion of bicuculline (10 microM), saclofen (1 microM) and naloxone (10 microM) had no effect on the stimulation by CGS21680 (S2/S1 ratio 0.99+/-0.04). The A(1) receptor agonist R-PIA (100 nM) inhibited the release of [(3)H]-acetylcholine (S2/S1 ratio 0.70+/-0.03), an effect blocked by DPCPX (S2/S1 ratio 1.06+/-0.07). It is concluded that both A(1) and A(2A) receptors are expressed on striatal cholinergic neurons where they are functionally active.

Topics: Acetylcholine; Adenosine; Adenosine Deaminase; Animals; Baclofen; Bicuculline; Caffeine; Cholinergic Fibers; Corpus Striatum; Dose-Response Relationship, Drug; GABA Antagonists; Gene Expression; Male; Membrane Potentials; Naloxone; Narcotic Antagonists; Neurons; Phenethylamines; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P1; RNA, Messenger; Tritium; Xanthines

We have examined the potency of several adenosine receptor antagonists at adenosine A1 and A2A receptors using quantitative autoradiography and have compared the results with those of previous studies using the same radioligands in membrane preparations. The agonists [3H]cyclohexyladenosine and [3H]2-[p-(2-carbonylethyl)-phenylethylamino]-5'-N-ethylcarbo xamido adenosine ([3H]CGS 21680) were used as radioligands for the two receptors. The results show that 1,3-dipropyl-8-cyclopentyl xanthine (DPCPX) is almost 1000-fold and 8-chloro-4-cyclohexyl-amino-1-(trifluoromethyl)[1,2,4]triazolo[4,3-a] quinoxaline (CP-68,247) about 300-fold more potent at adenosine A1 receptors in cortex and striatum than at striatal adenosine A2A receptors. Conversely, 5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo [1,5-c]pyrimidine (SCH 58261) is approximately 1000-fold and 4-(2-[7-amino-2-(2-furyl) [1,2,4]-triazolo[2,3-a][1,3,5]triazin-5-yl amino]ethyl)phenol (ZM 241,385) about 400-fold more potent at adenosine A2A than at A1 receptors. Caffeine and its metabolites did not show any selectivity. Other studied antagonists were non-selective or showed a modest (20- to 40-fold) adenosine A2A receptor selectivity. Thus, only a few of the antagonists show such high selectivity that it is not offset by differences in drug distribution and levels of receptor subtype expression.

Topics: Adenosine; Animals; Autoradiography; Binding, Competitive; Brain; Caffeine; In Vitro Techniques; Male; Phenethylamines; Purinergic P1 Receptor Antagonists; Pyrazines; Quinazolines; Rats; Rats, Sprague-Dawley; Receptor, Adenosine A2A; Receptors, Purinergic P1; Theophylline; Triazines; Triazoles; Tritium; Xanthines

The present experiments tested the hypothesis that adenosine A2 receptors are involved in central reward function. Adenosine receptor agonists or antagonists were administered to animals that had been trained to self-stimulate in a rate-free brain stimulation reward (BSR) task that provides current thresholds as a measure of reward. The adenosine A(2A) receptor-selective agonists 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamido adenosine hydrochloride (CGS 21680) (0.1-1.0 mg/kg) and 2-[(2-aminoethylamino)carbonylethyl phenylethylamino]-5'-N-ethylcarboxamido adenosine (APEC) (0.003-0.03 mg/kg) elevated reward thresholds without increasing response latencies, a measure of performance. Specifically, CGS 21680 had no effect on response latency, whereas APEC shortened latencies. Bilateral infusion of CGS 21680 (3, 10, and 30 ng/side), directly into the nucleus accumbens, elevated thresholds but shortened latencies. The highly selective A(2A) antagonist 8-(3-chlorostyryl)caffeine (0.01-10.0 mg/kg) and the A2-preferring antagonist 3,7-dimethyl-1-propargylxanthine (DMPX) (0.3-10.0 mg/kg) did not alter thresholds or latencies, but DMPX (1.0, 10.0 mg/kg) blocked the threshold-elevating effect of APEC (0.03 mg/kg). In another study, repeated administration of cocaine (eight cocaine injections of 15 mg/kg, i.p., administered over 9 hr) produced elevations in thresholds at 4, 8, and 12 hr after cocaine. DMPX (3 and 10 mg/kg), administered before both the 8 and 12 hr post-cocaine self-stimulation tests, reversed the threshold elevation produced by cocaine withdrawal. These results indicate that stimulating adenosine A(2A) receptors diminishes BSR without producing performance deficits, whereas blocking adenosine receptors reverses the reward impairment produced by cocaine withdrawal or by an A(2A) agonist. These findings indicate that adenosine, via A(2A) receptors, may inhibit central reward processes, particularly during the neuroadaptations associated with chronic drug-induced neuronal activation.

Topics: Adenosine; Animals; Brain; Caffeine; Cocaine; Differential Threshold; Dopamine Uptake Inhibitors; Drug Interactions; Electric Stimulation; Male; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Rats; Rats, Wistar; Reaction Time; Receptors, Purinergic P1; Reference Values; Reward; Substance Withdrawal Syndrome; Theobromine

Intrathecal (i.t.) strychnine produces localized allodynia in the rat without peripheral or central nerve injury. Intrathecal CPA (A1-selective agonist) and CGS-21680 (A2-selective agonist) dose-dependently inhibited strychnine-allodynia but with a 50-fold difference in potency (0.02-0.07 vs. 2.7-3.1 microgram, respectively). The anti-allodynic effect of CPA and CGS was completely blocked by pretreatment with the A1-selective antagonist, DPCPX (10 microgram i.t. ), but unaffected by the A2-selective antagonist, CSC (2 microgram i.t. ). The results indicate that spinal A1-, but not A2-, receptors modulate abnormal somatosensory input in the strychnine model, and suggest a difference in spinal purinergic modulation in injury vs. non-injury models of allodynia.

Topics: Adenosine; Animals; Blood Pressure; Caffeine; Heart Rate; Injections, Spinal; Male; Pain; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P1; Spinal Cord; Strychnine; Xanthines

Whole-cell patch clamp experiments were carried out in rat striatal brain slices. In a subset of striatal neurons (70-80%), NMDA-induced inward currents were inhibited by the adenosine A2A receptor selective agonist CGS 21680. The non-selective adenosine receptor antagonist 8-(p-sulphophenyl)-theophylline and the A2A receptor selective antagonist 8-(3-chlorostyryl)caffeine abolished the inhibitory action of CGS 21680. Intracellular GDP-beta-S, which is known to prevent G protein-mediated reactions, also eliminated the effect of CGS 21680. Extracellular dibutyryl cAMP, a membrane permeable analogue of cAMP, and intracellular Sp-cAMPS, an activator of cAMP-dependent protein kinases (PKA), both abolished the CGS 21680-induced inhibition. By contrast, Rp-cAMPS and PKI 14-24 amide, two inhibitors of PKA had no effect. Intracellular U-73122 (a phospholipase C inhibitor) and heparin (an inositoltriphosphate antagonist) prevented the effect of CGS 21680. Finally, a more efficient buffering of intracellular Ca2+ by a substitution of EGTA (11 mM) by BAPTA (5.5 mM) acted like U-73122 or heparin. Hence, A2A receptors appear to negatively modulate NMDA receptor channel conductance via the phospholipase C/inositoltriphosphate/Ca2+ pathway rather than the adenylate cyclase/PKA pathway.

Topics: Adenosine; Adenylyl Cyclases; Animals; Caffeine; Cyclic AMP-Dependent Protein Kinases; Enzyme Inhibitors; In Vitro Techniques; Neostriatum; Neurons; Patch-Clamp Techniques; Phenethylamines; Purinergic P1 Receptor Agonists; Rats; Receptors, N-Methyl-D-Aspartate; Receptors, Purinergic P1; Signal Transduction; Theophylline

Prolonged activation of an A2A adenosine receptor significantly inhibits the cellular response to subsequent stimulation (A2A desensitization). We have reported previously that activation of phosphodiesterase (PDE) contributes to A2A desensitization in PC12 cells. In the present study, we show that a type IV PDE (PDE4)-selective inhibitor (Ro 20-1724) effectively blocks the increase in PDE activity in desensitized cells. Thus, PDE4 appears to be the PDE specifically activated during A2A desensitization in PC12 cells. Prolonged treatment of PC12 cells with an A2A-selective agonist (CGS21680) leads to increased PDE4 activity in a dose-dependent manner, which can be blocked by an A2A-selective antagonist [8-(3-chlorostyryl)caffeine]. Using two PDE4 antibodies, we were able to demonstrate that the levels of two PDE4-immunoreactive bands (72 and 79 kDa) were increased significantly during A2A desensitization. Prolonged treatment with forskolin to elevate intracellular cyclic AMP contents also resulted in increased PDE4 activity. In addition, activation of PDE4 activity during A2A desensitization could be blocked by a protein kinase A (PKA)-selective inhibitor (H89) and was not observed in a PKA-deficient PC12 cell line (A123). Taken together, activation of PDE4 via a cyclic AMP/PKA-dependent pathway plays a critical role in dampening the signal of the A2A receptor.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; 4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone; Adenosine; Adrenal Gland Neoplasms; Animals; Blotting, Western; Caffeine; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclic Nucleotide Phosphodiesterases, Type 4; Enzyme Activation; Enzyme Inhibitors; Isoquinolines; Kinetics; PC12 Cells; Phenethylamines; Pheochromocytoma; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Polymerase Chain Reaction; Purinergic P1 Receptor Agonists; Rats; Receptor, Adenosine A2A; Receptors, Purinergic P1; Recombinant Proteins; Sulfonamides

In the hippocampus, the neuromodulatory role of adenosine depends on a balance between inhibitory A1 responses and facilitatory A2A responses. Since the presynaptic effects of hippocampal inhibitory A1 adenosine receptors are mostly mediated by inhibition of Ca2+ channels, we now investigated whether presynaptic facilitatory A2A adenosine receptors would modulate calcium influx in the hippocampus. The mixed A1/A2 agonist, 2-chloroadenosine (CADO; 1 microM) inhibited veratridine (20 microM)-evoked 45Ca2+ influx into hippocampal synaptosomes of the CA1 or CA3 areas by 24.2 +/- 4.5% and 17.2 +/- 5.8%, respectively. In the presence of the A, antagonist, 1,3-dipropyl-8-cyclopentylxanthine (DPCPX; 100 nM), the inhibitory effect of CADO (1 microM) on 45Ca2+ influx was prevented in CA1 synaptosomes, but was converted into a facilitatory effect (14.2 +/- 6.7%) in CA3 synaptosomes. The A2A agonist, CGS 21680 (3-30 nM) facilitated 45Ca2+ influx in CA3 synaptosomes, with a maximum increase of 22.9 +/- 3.9% at 10 nM, and was virtually devoid of effect in CA1 synaptosomes. This facilitatory effect of CGS 21680 (10 nM) in CA3 synaptosomes was prevented by the A2A antagonist 8-(3-chlorostyryl)caffeine (CSC; 200 nM), but not by the A1 antagonist, DPCPX (20 or 100 nM). The facilitatory effect of CGS 21680 on 45Ca2+ uptake by CA3 synaptosomes was prevented by the class A calcium channel blocker, omega-agatoxin-IVA (200 nM). These results indicate that presynaptic adenosine A2A receptors facilitate calcium influx in the CA3 but not the CA1 area of the rat hippocampus through activation of class A calcium channels.

Topics: 2-Chloroadenosine; Adenosine; Animals; Caffeine; Calcium Channel Blockers; Calcium Channels; Calcium Radioisotopes; Hippocampus; Male; omega-Agatoxin IVA; Phenethylamines; Purinergic P1 Receptor Agonists; Rats; Rats, Wistar; Receptors, Purinergic P1; Spider Venoms; Synaptosomes; Veratridine; Xanthines

In the present study, we demonstrate that the Ca(2+)-inhibitable adenylyl cyclase (AC) activity in the striatum exhibits a daily oscillation with a peak occurring around 10:00 h. A circadian fluctuation of the AC activity evoked by an A2a adenosine-selective agonist was also observed. Intrastriatal injection of an A2a-selective adenosine agonist or antagonist during the interval in which the Ca(2+)-inhibitable AC activity was at its peak resulted in a more significant alteration of locomotor activity than those observed at a later interval. The marked circadian variation in the Ca(2+)-inhibitable AC activity in the striatum appears to cause a circadian fluctuation in the action of at least one neuromodulator.

Topics: Adenosine; Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Animals; Caffeine; Calcium; Circadian Rhythm; Colforsin; Corpus Striatum; Cyclic AMP; Guanosine Diphosphate; Isoenzymes; Male; Motor Activity; Phenethylamines; Purinergic P1 Receptor Agonists; Rats; Rats, Sprague-Dawley

The purpose of the present study was to quantify the in vivo potency of the selective adenosine A2a antagonist CSC [8-(3-chlorostyryl)caffeine]. Four groups of conscious, normotensive rats received a continuous i.v. infusion of 0, 6, 12 and 24 micrograms/min/kg of CSC. During a steady-state infusion of CSC, the animals received 1000 micrograms/kg of the adenosine A2a receptor agonist CGS 21680C [the sodium salt of 2-p-(2-carboxyethyl) phenylethylamino-5'-N-ethylcarboxamidoadenosine] i.v. over 15 min. During the experiment, the mean arterial pressure and the heart rate were recorded continuously and arterial blood samples were taken for the analysis of drug concentrations. For each individual rat, the CGS 21680C-provoked reduction in blood pressure was related to the blood concentration of the agonist according to the sigmoidal Emax model. The presence of CSC produced a parallel shift of the concentration-hypotensive effect curve to the right, indicating competitive interaction of the compounds. Infusion of 0, 6, 12 and 24 micrograms/min/kg of CSC resulted in steady-state concentrations of 0, 85 +/- 7, 210 +/- 20 and 400 +/- 40 ng/ml, and apparent EC50 values of CGS 21680C based on free concentrations (EC50,u) of 4.8 +/- 1.1, 7.2 +/- 0.5, 32 +/- 6 and 57 +/- 10 ng/ml, respectively (mean +/- S.E., n = 6, 6, 5 and 6). The relationship between the CSC concentration and the apparent EC50 was quantified according to a competitive pharmacodynamic interaction model.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine; Animals; Antihypertensive Agents; Blood Pressure; Caffeine; Drug Evaluation, Preclinical; Drug Interactions; Heart Rate; Male; Phenethylamines; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Rats