thromboxane-a2 and alpha-beta-methyleneadenosine-5--triphosphate

ATP and ADP are simultaneously released from activated platelets in equimolar concentrations. Micromolar concentrations of ATP inhibit platelet aggregation by both competitive and non-competitive mechanisms. The current studies addressed the question of how platelets respond to agonists in the presence of nanomolar and micromolar concentrations of ATP and ADP alone or in combination. This is a significant issue since the concentration of ATP +/- ADP may vary widely within a microenvironment depending upon the source and cause for the release of the nucleotides. ATP (1-10 nM) was found to significantly enhance the thromboxane A2 analog, U44619-, collagen- and thrombin-induced platelet aggregations. Conversely, ATP at 1-100 microM inhibited these same reactions. ADP, in general, behaved exactly opposite to ATP. When equal amounts of ATP and ADP were added together the ADP response appeared to predominate. The observed ATP-induced response was not due to a hydrolytic product as evidenced by an unaltered response to ATP in the presence of adenosine deaminase or the ATP generating system, creatine phosphate plus creatine phosphokinase. Adenosine (1-10 nM), like ADP, inhibited agonist-induced platelet aggregation. The stimulation of agonist-induced platelet aggregation by 1-10 nM extracellular ATP appears to depend upon the phosphorylation of platelet membrane ecto proteins. The ATP analog, beta gamma-methylene ATP, that is incapable of serving as a phosphate donor for protein kinases, inhibited rather than stimulated agonist-induced platelet aggregation. The dual response of platelets to low and high concentrations of extracellular ATP +/- ADP may play a physiological role in hemostasis and thrombosis under normal and pathological conditions.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenosine Diphosphate; Adenosine Triphosphate; Chromatography, High Pressure Liquid; Collagen; Dose-Response Relationship, Drug; Humans; Platelet Aggregation; Platelet Aggregation Inhibitors; Prostaglandin Endoperoxides, Synthetic; Thrombin; Thromboxane A2

1. The potentiating effect of NG-nitro-L-arginine methyl ester, (L-NAME) a nitric oxide synthesis inhibitor, on responses of the rat pulmonary vascular pressure (PVP) to purinoceptor agonists was examined. 2. At a constant flow of 23 ml min-1 the PVP was 22.4 +/- 2.5 mmHg (n = 15), and treatment with 100 microM L-NAME for 15 min was without effect on the PVP. After the tone was raised with 28 nmol 9,11-dideoxy-11 alpha, 9 alpha-epoxymethano-prostaglandin F2 alpha (U-46619), the PVP was 29.4 +/- 3.3 mmHg and treatment with 100 microM L-NAME was still without effect on the PVP. It appears that there is a graded release of nitric oxide in response to different levels of steady shear stress and in our experimental model the threshold for detection was not reached under basal conditions. 3. In contrast, when the circulation was challenged with 30 s step, additive increases in flow between 11 and 50 ml min-1 (n = 8), treatment with 100 microM L-NAME produced a significant (P < 0.05) increase in PVP suggesting that changes in flow-derived forces evoke the release of nitric oxide. This was evident for flow rates above 30 ml min-1. 4. In preparations in which tone was raised with U-46619, a dose of 1 x 10(-8) mol ATP or 2-meSATP evoked a drop in PVP while alpha,beta-meATP produced an increase in PVP under constant flow of 23 ml min-1. After treatment with 100 microM L-NAME, all three purinoceptor agonists evoked an increase in PVP. The increase in Pvp evoked by alpha, beta-meATP was not affected by L-NAME. These results suggest that P2Y-purinoceptor stimulation evokes the release of nitric oxide to produce vasodilatation.5. Under conditions of constant flow and basal pressure, 100 microM L-NAME significantly (P<0.05)potentiated the increase in Pvp evoked by 1 x 10-6 mol ATP, although the increase evoked by 1 x 10-8 mol (alpha,beta-meATP, which was of similar magnitude, was not affected. These results indicate that a blockade of evoked nitric oxide release is responsible for the potentiation of the increase in Pvp evoked by ATP.6. This study shows that, while nitric oxide does not appear to be released in the pulmonary circulation of the rat under constant flow conditions, nitric oxide release evoked by purinoceptor agonists attenuates increases in pulmonary vascular pressure.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenosine Triphosphate; Animals; Arginine; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Prostaglandin Endoperoxides, Synthetic; Pulmonary Circulation; Pulmonary Wedge Pressure; Rats; Rats, Wistar; Regression Analysis; Thromboxane A2; Vascular Resistance; Vasoconstrictor Agents

It has been reported recently that adenosine and ATP produce dose- and tone-dependent responses in the feline pulmonary vascular (PV) bed. The present study was undertaken to investigate the mechanisms mediating vasoconstrictor (VC) responses to adenosine and ATP in the intact-chest, spontaneously breathing cat under conditions of controlled blood flow and constant left atrial pressure. The order of potency of adenosine receptor agonists to produce VC in the PV bed was the selective adenosine A1 receptor agonist R-phenylisopropyladenosine greater than the mixed A1, A2 receptor agonist, adenosine greater than the selective adenosine A2 receptor agonist, 2-phenylaminoadenosine. The dose-related increase in lobar arterial pressure in response to adenosine was blocked by an adenosine (P1) receptor antagonist, BWA1433U, the cyclooxygenase inhibitor, meclofenamate, and the thromboxane A2 receptor antagonist, SQ29548. The order of potency of ATP analogs to produce VC in the PV bed was alpha,beta-methylene ATP (alpha,beta-meATP) much greater than beta,tau-methylene ATP greater than ATP. BWA1433U inhibited VC responses to ATP without affecting responses to its degradation-resistant analogs beta,tau-methylene ATP and alpha,beta-meATP. In the presence of BWA1433U and a continuous intralobar infusion of the selective 5'-nucleotidase inhibitor, alpha,beta-methyleneadenosine-5'-diphosphate, ATP VC responses are significantly enhanced compared to those after BWA1433U. alpha,beta-Methyleneadenosine-5'-diphosphate had no effect on the VC response to U44069 after BWA1433U. Meclofenamate significantly inhibited the vasoconstrictor responses to ATP but not to alpha,beta-meATP.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine; Adenosine Diphosphate; Adenosine Triphosphate; Animals; Bridged Bicyclo Compounds, Heterocyclic; Cats; Cyclooxygenase Inhibitors; Fatty Acids, Unsaturated; Hydrazines; Lung; Meclofenamic Acid; Phenylisopropyladenosine; Purinergic Antagonists; Receptors, Prostaglandin; Receptors, Purinergic; Receptors, Thromboxane; Thromboxane A2; Vasoconstriction; Vasodilator Agents; Xanthines

Both alpha, beta-methylene ATP and beta, gamma-methylene ATP (P2X selective agonists) were shown to induce transient contraction in intact and endothelium-removed preparations of canine basilar arteries. 2-Methylthio ATP (a P2Y selective agonist) caused transient contraction of intact arteries and this response was nearly abolished by removal of the endothelium. In the presence of alpha, beta-methylene ATP (10(-6) M), the endothelium-independent contractions induced by alpha, beta-methylene ATP itself (10(-6) M) and by beta, gamma-methylene ATP (10(-5) M) were both abolished. The endothelium-dependent contraction induced by 2-methylthio ATP (10(-7) M) was not attenuated by alpha, beta-methylene ATP. The contraction induced by 2-methylthio ATP (10(-7) M) was attenuated markedly by reactive blue 2 (a P2Y antagonist) (3 x 10(-6) M), aspirin (5 x 10(-5) M), OKY-046 (thromboxane A2 synthetase inhibitor) (10(-5) M) and ONO-3708 (thromboxane A2 antagonist) (10(-8) M). However, these agents did not affect the endothelium-independent contraction induced by alpha, beta-methylene ATP (10(-6) M). Neither TMK-777 (a 5-lipoxygenase inhibitor) (10(-7) M) nor superoxide dismutase (100 U/ml) plus catalase (1,000 U/ml) affected either contraction. The present experiments demonstrate that P2X-purinoceptors mediate endothelium-dependent contraction in the canine basilar artery, and that the endothelium-derived contracting factor in this system is probably thromboxane A2.

Topics: Adenosine Triphosphate; Animals; Aspirin; Basilar Artery; Calcium; Dogs; Endothelium, Vascular; Female; Male; Methacrylates; Nifedipine; Receptors, Purinergic; Thromboxane A2; Vasoconstriction

ATP, ADP, AMP and adenosine at 10(-8) induced a slight relaxation, and at concentrations greater than 10(-7) M caused a biphasic response consisting of an initial relaxation followed, after a brief period, by a transient contraction in canine basilar artery. ATP (10(-6) and 10(-5) M) caused a triphasic response consisting of a rapid, small contraction, a relaxation and then a second, transient contraction. The order of agonist potency for contraction was ATP greater than ADP much greater than AMP = adenosine, but for producing relaxation the agonists were equipotent. Removal of the endothelium abolished the contraction after the relaxation, but had virtually no effect on the relaxation and the rapid contraction induced by ATP (10(-6) and 10(-5) M). Only the relaxation in response to ATP (10(-6) M) was attenuated by removal of the endothelium. Aspirin (a cyclooxygenase inhibitor) (5 X 10(-5) M), OKY-046 (a thromboxane A2 synthetase inhibitor) (10(-5) M) and ONO-3708 (a thromboxane A2 antagonist) (5 X 10(-9) M) attenuated markedly the endothelium-dependent contraction induced by ATP (10(-5) M) and ADP (10(-5) M), but did not affect the relaxation. Phentolamine (10(-6) M) and atropine (10(-6) M) did not affect either the contraction or the relaxation. The relaxations induced by both ATP and adenosine in both endothelium-intact preparations and endothelium-removed preparations were attenuated by 8-phenyltheophylline (P1 antagonist) (10(-6) M).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenine Nucleotides; Adenosine; Adenosine Triphosphate; Animals; Basilar Artery; Dogs; Endothelium, Vascular; Female; In Vitro Techniques; Male; Theophylline; Thromboxane A2; Vasoconstriction; Vasodilation