thromboxane-a2 and linsidomine

We compared the effects of L-arginine (L-ARG), the precursor of endogenous NO, on platelet aggregation and thromboxane A2 formation in vivo and in vitro. Human platelet-rich plasma (PRP) was anticoagulated with citrate (which decreases extracellular Ca2+) or with recombinant hirudin (which does not affect extracellular Ca2+). Two groups of 10 healthy male volunteers received intravenous infusions of L-ARG (30 g or 6 g, 30 min) or placebo. Blood was collected immediately before and at the end of the infusions for aggregation by ADP or collagen. Infusion of L-ARG inhibited ADP-induced aggregation in PRP anticoagulated with citrate by 37.5+/-6.3% (P < 0.05). In PRP anticoagulated with hirudin, aggregation was inhibited by 33.6+/-16.0% (P < 0.05). L-ARG infusion also inhibited platelet TXB2 formation and slightly, but not significantly decreased the urinary excretion rate of 2,3-dinor-TXB2; cGMP concentrations in PRP were significantly elevated during L-arginine infusion. In vitro preincubation with L-ARG (10 microM-2.5 mM) inhibited platelet aggregation in PRP anticoagulated with rhirudin, but not citrate. This effect was stereospecific for L-arginine, as D-arginine had no effect. It was dependent upon NO synthase activity, as indicated by increased cGMP levels in PRP. Moreover, both the NOS inhibitor L-NMMA and the inhibitor of soluble guanylyl cyclase ODQ antagonized the effects of L-ARG. Haemoglobin, an extracellular scavenger of NO, partly antagonized the antiplatelet effects of L-ARG. 8-Br-cyclic GMP and the exogenous NO donor linsidomine inhibited aggregation in PRP anticoagulated with citrate or r-hirudin. The inhibitory effects of L-ARG on platelet aggregation in vitro were paralleled by increased cyclic GMP levels; L-ARG also inhibited platelet TXB2 formation in PRP anticoagulated with r-hirudin, but not citrate. We conclude that the L-arginine/NO pathway is present in human platelets as a Ca2+-dependent anti-aggregatory pathway. In vivo the formation of NO from L-ARG by endothelial cells may contribute to the platelet-inhibitory effects of L-ARG. NO-releasing compounds like linsidomine inhibit platelet aggregation in vitro independent of extracellular Ca2+.

Topics: Adult; Antithrombins; Arginine; Blood Platelets; Cyclic GMP; Enzyme Inhibitors; Humans; In Vitro Techniques; Infusions, Intravenous; Male; Molsidomine; Nitric Oxide Synthase; Platelet Aggregation; Platelet Aggregation Inhibitors; Thromboxane A2; Thromboxane B2; Thromboxanes

The central effect of 3-morpholinosydnonimine, a nitric oxide donor, on the sympatho-adrenomedullary system was investigated in urethane-anesthetized rats. Intracerebroventricular administration of 3-morpholinosydnonimine (100, 250 and 500 microg/animal) induced a marked elevation of adrenaline levels and a slight elevation of noradrenaline levels in the plasma. These 3-morpholinosydnonimine (250 microg/animal)-induced elevations of catecholamines were abolished by intracerebroventricular treatments with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-l-oxyl 3-oxide (750 microg/animal), a nitric oxide scavenger, and indomethacin (500 microg/animal), a cyclo-oxygenase inhibitor, but not with superoxide dismutase (250 units/animal), a superoxide anion scavenger. Furthermore, the 3-morpholinosydnonimine (250 microg/animal)-induced elevation of plasma adrenaline levels was abolished by intracerebroventricular treatments with thromboxane A2 synthase inhibitors [furegrelate (100, 250 and 1000 microg/animal) and carboxyheptyl imidazole (500 microg/animal)], and also with thromboxane A2 receptor blockers [(+)-S-145 (100, 250 and 1000microg/animal) and SQ29548 (8microg/animal)]. The elevation of noradrenaline levels was, however, not attenuated by these thromboxane A2-related test agents. The present results indicate that nitric oxide but not peroxynitrite markedly activates central adrenomedullary outflow. Thromboxane A2 in the brain is probably involved in this central activation of adrenomedullary outflow.

Topics: Animals; Benzoates; Cyclooxygenase Inhibitors; Epinephrine; Imidazoles; Indomethacin; Injections, Intraventricular; Male; Medulla Oblongata; Molsidomine; Nitric Oxide; Norepinephrine; Rats; Rats, Wistar; Receptors, Thromboxane; Superoxide Dismutase; Thromboxane A2; Thromboxane-A Synthase

We have recently reported that halothane (Hal) anesthesia attenuates the pulmonary vasodilator responses to both bradykinin (BK) and sodium nitroprusside (SNP) compared with responses measured in the conscious state. These agonists have been classically used to activate endothelium-dependent and -independent vasodilator pathways, respectively. Our present goal was to assess the effect of isoflurane (Iso) anesthesia on pulmonary vasodilation activated via these pathways. Left pulmonary vascular pressure-flow (P-Q) plots were used to measure the pulmonary vascular responses to cumulative intravenous doses of BK, SNP, and 3-morpholinosydonimine-N-ethylcarbamide (SIN-1), a nitric oxide donor, in chronically instrumented dogs in the conscious state and during Iso anesthesia after matched preconstriction with the thromboxane analogue U-46619. Iso attenuated the vasodilator response to BK (P < 0.05). However, Iso had a differential effect on the responses to SIN-1 and SNP. Iso potentiated the vasodilator response to SIN-1 (P < 0.05), whereas Iso attenuated the response to SNP (P < 0.05). The vasodilator response to SIN-1 was unchanged during Hal anesthesia. The ATP-sensitive potassium (KATP)-channel inhibitor glibenclamide attenuated the vasodilator response to SNP (P < 0.05) but not to SIN-1. Thus Iso and Hal selectively attenuate the endothelium-dependent pulmonary vasodilator response to BK. Both anesthetics attenuate vasodilation induced by SNP but not by SIN-1. Moreover, a component of SNP-induced vasodilation involves KATP-channel activation.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Anesthesia; Animals; Blood Pressure; Bradykinin; Dogs; Dose-Response Relationship, Drug; Endothelium, Vascular; Gases; Halothane; Hemodynamics; Homeostasis; Isoflurane; Male; Molsidomine; Nitroprusside; Prostaglandin Endoperoxides, Synthetic; Pulmonary Circulation; Thromboxane A2; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents

In extracts of human platelets, three isoenzymes of cyclic nucleotide phosphodiesterase (PDE), namely, PDE2, PDE3, and PDE5, were identified; activities of PDE1 and PDE4 were not detected. In human platelets, the cGMP-hydrolytic activity was about six times higher than the cAMP-hydrolytic activity, and PDE5 and PDE3 are the major phosphodiesterase isoenzymes that hydrolyze cGMP and cAMP, respectively. PDE5 exhibited organ-specific expression in humans, and platelets were among the tissues richest in PDE5. A novel inhibitor of PDE5, sodium 1-[6-chloro-4-(3,4-methylenedioxybenzyl)aminoquinazolin-2-yl ] piperidine-4-carboxylate sesquihydrate (E4021), was a potent and highly selective inhibitor of human platelet PDE5. However, E4021 (up to 10 microM) did not inhibit 9,11-epithio-11,12-methano-thromboxane A2-induced platelet aggregation, in vitro. E4021 plus SIN-1 (3-morpholino-sydnonimine), at concentrations that had little effect individually, inhibited aggregation. These results suggest the unique distribution of phosphodiesterase isoenzymes in human platelets and the PDE5 inhibitors might be useful as a new class of antiplatelet drugs.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Blood Platelets; Cyclic Nucleotide Phosphodiesterases, Type 5; Humans; In Vitro Techniques; Isoenzymes; Molsidomine; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Phthalazines; Piperidines; Platelet Aggregation; Platelet Aggregation Inhibitors; Quinazolines; Signal Transduction; Thromboxane A2; Tissue Distribution

When whole blood is stirred there is a "spontaneous" platelet aggregation (SPA) which is presumed to be caused by proaggregatory factors released from platelets and other blood cells. Adding streptokinase (SK) to stirred whole blood frequently increases the rate and extent of the platelet aggregation that occurs; this is likely to be via immune complex formation between SK and natural anti-SK antibodies leading to increased release of pro-aggregatory factors. In this investigation we have examined the effects of several inhibitors and antagonists in an attempt to identify the proaggregatory factors that contribute to both SPA and SK-induced aggregation (SKA) and to evaluate different means of inhibiting both processes. The effects of the inhibitors/antagonists were determined in vitro after adding them to citrated whole blood obtained from healthy volunteers. Platelet aggregation was measured using a platelet counting technique. Inhibition of both SPA and SKA by apyrase and by FPL 66096 (a P2T receptor antagonist) demonstrated the involvement of ADP in both processes. Inhibition by chlorpromazine indicated that the most likely source of the ADP is red cells. The effects of sulotroban (a TXA2 antagonist) indicated involvement of TXA2 in SKA but not in SPA. The lack of effect of specific antagonists at S2, alpha 2 and PAF receptors suggested lack of involvement of serotonin, catecholamines and platelet-activating factor in either SPA or SKA. Both SPA and SKA were potently inhibited by low concentrations of iloprost (a PGI2 analogue), but a high concentration of SIN-1 (a NO donor) was much less effective.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Diphosphate; Amino Acid Sequence; Hirudins; Humans; Iloprost; Magnesium Chloride; Molecular Sequence Data; Molsidomine; Platelet Aggregation; Platelet Aggregation Inhibitors; Reference Values; Streptokinase; Thromboxane A2

The metabolite of molsidomine, 3-morpholino-syndnonimine (SIN-1) is a vasodilator and an inhibitor of platelet activation induced by adenosine-5'-diphosphate (ADP), arachidonic acid (AA) and thrombin. We present the results of SIN-1 on platelet aggregation induced by paf-acether and on the biosynthesis of this mediator by washed rabbit platelets. SIN-1 inhibits submaximal platelet aggregation induced by 50 pM paf-acether. This inhibition is dose-dependent (SIN-1 IC50: 37 +/- 10 nM, n = 3). SIN-1 also inhibits submaximal aggregations induced by ADP (IC50: 13 +/- 11 nM) and AA (IC50: 26 +/- 12 nM) (n = 3-5). Finally, SIN-1 inhibits dose-dependently the formation of lyso paf-acether (the precursor of paf-acether) and of thromboxane A2, and AA freeing by thrombin-stimulated platelets. Thus SIN-1 has a wide spectrum activity on platelets. He is both capable of inhibiting platelet aggregation induced by all used agonists and to inhibit the formation of paf-acether and thromboxane A2, two powerful mediators of platelet aggregation.

Topics: Animals; Blood Platelets; Molsidomine; Platelet Activating Factor; Platelet Aggregation; Rabbits; Thromboxane A2

N-Carboxy-3-morpholinosydnone imine ethyl ester (molsidomine) and its main metabolite 3-morpholinosydnone imine (SIN-1) were investigated in rabbit platelet-rich plasma (PRP) for antiaggregatory activity and inhibition of thromboxane A2 (TXA2) generation (arachidonic acid (AA)-induced) as well as in human PRP regarding prostaglandin endoperoxide analogue (U-46 619)- and AA-induced aggregation and TXB2 formation. The results were compared with the effects of nictindole. In rabbit PRP the inhibitory effects of molsidomine on aggregation and TXA2 generation were 20fold lower than that of SIN-1. The IC50-values of SIN-1, which is the pharmacologically active biotransformation product of molsidomine, were about 1 mumol/l in the experimental models used. The inhibitory effects of nictindole were about 50 times higher than those of SIN-1. In human PRP the inhibitory potency of SIN-1 decreased in the following sequence: U-46 619-induced aggregation (IC50 = 0.9 mumol/l) greater than AA-induced aggregation (IC50 = 1.4 mumol/l) greater than TXB2 formation (IC50 = 2.9 mumol/l). Molsidomine was nearly without effect in human PRP. Our own preliminary results and findings obtained by other authors seem to exclude a direct effect of SIN-1 on cyclooxygenase and thromboxane synthetase. The possible clinical significance of our findings in different types of angina pectoris and for myocardial infarction is discussed.

Topics: Animals; Arachidonic Acids; Blood Platelets; Drug Interactions; Female; Humans; Indoles; Male; Molsidomine; Oxadiazoles; Platelet Aggregation; Prostaglandin Endoperoxides, Synthetic; Pyridines; Rabbits; Sydnones; Thromboxane A2; Thromboxanes