A comparison of an A1 adenosine receptor agonist (CVT-510) with diltiazem for slowing of AV nodal conduction in guinea-pig.

1. The purpose of this study was to compare the pharmacological properties (i.e. the AV nodal depressant, vasodilator, and inotropic effects) of two AV nodal blocking agents belonging to different drug classes; a novel A1 adenosine receptor (A1 receptor) agonist, N-(3(R)-tetrahydrofuranyl)-6-aminopurine riboside (CVT-510), and the prototypical calcium channel blocker diltiazem. 2. In the atrial-paced isolated heart, CVT-510 was approximately 5 fold more potent to prolong the stimulus-to-His bundle (S-H interval), a measure of slowing AV nodal conduction (EC50 = 41 nM) than to increase coronary conductance (EC50 = 200 nM). At concentrations of CVT-510 (40 nM) and diltiazem (1 microM) that caused equal prolongation of S-H interval (approximately 10 ms), diltiazem, but not CVT-510, significantly reduced left ventricular developed pressure (LVP) and markedly increased coronary conductance. CVT-510 shortened atrial (EC50 = 73 nM) but not the ventricular monophasic action potentials (MAP). 3. In atrial-paced anaesthetized guinea-pigs, intravenous infusions of CVT-510 and diltiazem caused nearly equal prolongations of P-R interval. However, diltiazem, but not CVT-510, significantly reduced mean arterial blood pressure. 4. Both CVT-510 and diltiazem prolonged S-H interval, i.e., slowed AV nodal conduction. However, the A1 receptor-selective agonist CVT-510 did so without causing the negative inotropic, vasodilator, and hypotensive effects associated with diltiazem. Because CVT-510 did not affect the ventricular action potential, it is unlikely that this agonist will have a proarrythmic action in ventricular myocardium.

A2A-adenosine receptor reserve for coronary vasodilation.

BACKGROUND: Adenosine is a potent coronary vasodilator and causes an increase of coronary blood flow by activation of A2A-adenosine receptors (A2A-AdoRs). The purpose of this study was to test the hypothesis that the high potency of adenosine and adenosine analogues to cause coronary vasodilation is explained by the presence of a large A2A-AdoR reserve ("spare receptors"). METHODS AND RESULTS: A novel, irreversible antagonist of A2A-AdoRs was used to inactivate receptors and reduce the response to agonist. Agonist-induced increases of coronary conductance before and after exposure of hearts to the irreversible antagonist were compared. Three agonists were studied: 2-p-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS21680), adenosine, and 2-chloro-N6-cyclopentyladenosine (CCPA). Data were analyzed to determine agonist KA (equilibrium dissociation constant) and EC50 values. Values of KA for activation of A2A-AdoRs by CGS21680, adenosine, and CCPA were 105, 1800, and 2630 nmol/L, respectively. In contrast, values of EC50 for CGS21680, adenosine, and CCPA to increase coronary conductance were 1.5, 85, and 243 nmol/L, respectively. By use of the law of mass action, it was calculated that half-maximal responses to CGS21680, adenosine, and CCPA occurred when only 1.3%, 5%, and 9%, respectively, of A2A-AdoRs were occupied by agonist. CONCLUSIONS: Receptor reserves for 3 A2A-AdoR agonists were large. The receptor reserve for A2A-AdoRs to cause an increase of coronary conductance can explain both the high potency of adenosine to cause coronary vasodilation and the observation that an A2A-AdoR agonist can cause coronary vasodilation without systemic effects.

The A2A adenosine receptor mediates coronary vasodilation.

The coronary vasodilation caused by adenosine is due to activation of A2 adenosine receptors (A2AdoRs), but the subtype or subtypes of A2AdoR (A2A and/or A2B) that mediate this action are uncertain. The purpose of this study was to test the hypothesis that A2AAdoRs mediate coronary vasodilation caused by exogenous or endogenous adenosine in the guinea pig isolated perfused heart. The newly described A2AAdoR antagonist SCH58261 was used to selectively block A2AAdoRs. Attenuations by SCH58261 of increases in coronary conductance (A2 response) and of atrioventricular nodal conduction time (A1 response) caused by exogenous and endogenous adenosine and by agonists with relative selectivity for A2A and A1AdoRs were measured. The CGS21680-induced increase of coronary conductance was antagonized by SCH58261 in a concentration-dependent and competitive manner with a KB value of 5.01 nm. Also reversed by SCH58261 (60 nmol/L) were the increases in coronary conductance caused by the relatively selective A1AdoR agonists CCPA (70 nM), and (R)-(-)N(b)-(2-phenyl-isopropyl)adenosine (60 nM) but not those caused by sodium nitroprusside (1.2 microM) and diltiazem (0.4 microM). SCH58261 (< or = 100 nM) did not attenuate the A1AdoR-mediated prolongations of S-H interval caused by either adenosine or CCPA. SCH58261 attenuated the coronary vasodilation caused by 50 nM adenosine with an IC50 value of 6.8 +/- 0.6 nM. The coronary vasodilations caused by the nucleoside uptake inhibitor draflazine and the adenosine kinase inhibitor iodotubercidin were completely reversed by 60 nM SCH58261 or adenosine deaminase (7 U/ml). Thus, the A2AAdoR plays a major role as mediator of coronary vasodilation caused by exogenous and endogenous adenosine and by AdoR agonists.

Inflammatory mediators stimulate arginine transport and arginine-derived nitric oxide production in a murine breast cancer cell line.

Inflammatory mediators stimulate arginine-derived nitric oxide (NO) production in a variety of cells. The purpose of this study was to determine if the inflammatory mediators, endotoxin (LPS) and interferon gamma (IFN), stimulate arginine transport and nitric oxide production in a murine breast cancer cell line. We also investigated the effect of the nitric oxide synthase (NOS) inhibitors, omega-nitro-L-arginine methyl ester (LNAME) and aminoguanidine (AG), as well as the effect of varying the concentration of L-arginine in the cellular media, on arginine transport and NO production in these tumors cells. Confluent EMT-6 murine breast cancer cells were incubated with LPS (10 microgram/ml) and IFN (50 units/ml) in the presence or absence of the NOS inhibitors, L-NAME (2 mM) or AG (1 mM), and arginine transport (using L-[3H]arginine) and NO production (the stable end-product nitrite was assayed using the Greiss reagent) were measured at various time points. In addition, the effect of varying the concentration of L-arginine (0, 10, 100, 1000, 10,000 mM) in the cellular media on stimulated L-arginine transport and nitrite accumulation was assessed. Incubation of EMT-6 with LPS and IFN stimulated arginine transport approximately 70% over control levels at 12 hr and transport returned to basal levels at 24 hr. LPS/IFN-stimulated EMT-6 cells produced 25 microM nitrite at 24 hr and reached a plateau of 55 microM nitrite at 48 hr. The NO synthase inhibitors, L-NAME and AG, failed to inhibit basal and stimulated levels of arginine transport, but significantly inhibited nitrite accumulation, which was restored by 10 mM L-arginine. Finally, L-arginine was necessary in the media for nitrite accumulation by LPS/IFN-stimulated cells, with maximal accumulation at 1 mM L-arginine. In summary, LPS/IFN stimulate arginine transport and NO production in the EMT-6 breast cancer cell line. L-NAME and AG do not inhibit basal or stimulated arginine transport in this tumor cell line and extracellular L-arginine is required for NO synthesis in these cells. LPS/IFN stimulation of arginine transport may represent an adaptive response to provide increased substrate for enhanced tumor cell NO production.