15-hydroxy-11-alpha-9-alpha-(epoxymethano)prosta-5-13-dienoic-acid and 9-(tetrahydro-2-furyl)-adenine

Amarogentin, an active principle of Gentiana lutea, possess antitumorigenic, antidiabetic, and antioxidative properties. Activation of platelets is associated with intravascular thrombosis and cardiovascular diseases. The present study examined the effects of amarogentin on platelet activation. Amarogentin treatment (15~60  μM) inhibited platelet aggregation induced by collagen, but not thrombin, arachidonic acid, and U46619. Amarogentin inhibited collagen-induced phosphorylation of phospholipase C (PLC) γ2, protein kinase C (PKC), and mitogen-activated protein kinases (MAPKs). It also inhibits in vivo thrombus formation in mice. In addition, neither the guanylate cyclase inhibitor ODQ nor the adenylate cyclase inhibitor SQ22536 affected the amarogentin-mediated inhibition of platelet aggregation, which suggests that amarogentin does not regulate the levels of cyclic AMP and cyclic GMP. In conclusion, amarogentin prevents platelet activation through the inhibition of PLC γ2-PKC cascade and MAPK pathway. Our findings suggest that amarogentin may offer therapeutic potential for preventing or treating thromboembolic disorders.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenine; Adenosine Triphosphate; Animals; Arachidonic Acid; Collagen; Cyclic GMP; Guanylate Cyclase; Humans; Iridoids; MAP Kinase Signaling System; Mice; Oxadiazoles; Phospholipase C gamma; Plant Extracts; Platelet Activation; Protein Kinase C; Quinoxalines; Thrombin; Thromboembolism; Thrombosis

The Na+/Ca2+ exchanger (NCX) may be an important modulator of Ca2+ entry and exit. The present study investigated whether NCX was affected by prostacyclin and nitric oxide (NO) released from the vascular endothelium, as NCX contains phosphorylation sites for PKA and PKG.. Rat aortic rings were set up in organ baths. Tension was measured across the ring with a force transducer.. Lowering extracellular [Na+] ([Na+]o) to 1.18 mM induced vasoconstriction in rat endothelium-denuded aortic rings. This effect was blocked by the NCX inhibitor KB-R7943 (2-2-[4-(4-nitrobenzyloxy)phenyl] ethyl isothiourea methanesulphonate; 1 microM). In endothelium-intact aortic rings, decreasing [Na+]o did not constrict the aortic rings significantly, but after treatment with the guanylate cyclase inhibitor ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; 1 microM) or the NOS inhibitor L-NAME (N(omega)-nitro-L-arginine methyl ester; 50 microM), a vasoconstriction that was similar in size to that in endothelium-denuded preparations was evident. The vasorelaxation induced by the NO donor sodium nitroprusside sodium nitroprusside dihydrate (30 nM) was the same in the endothelium-denuded aortic rings preconstricted with either low Na+ (1.18 mM), the thromboxane A2 agonist U46619 (9,11-dideoxy-9alpha, 11alpha-methanoepoxy prostaglandin F(2alpha); 0.1 microM) or high K+ (80 mM).. The results suggest that the endothelium inhibits NCX operation via guanylate cyclase/NO. This is stronger than for other constrictors such as phenylephrine and may relate to concomitant NCX-stimulated NO release from the endothelium. This finding may be important where NCX operates in reverse mode, such as during ischaemia, and highlights a new mechanism whereby the endothelium modulates Ca2+ homoeostasis in vascular smooth muscle.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenine; Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Animals; Aorta, Thoracic; Dinoprost; Endothelium, Vascular; Enzyme Inhibitors; Guanylate Cyclase; In Vitro Techniques; Indomethacin; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitroprusside; Oxadiazoles; Potassium; Prostaglandin-Endoperoxide Synthases; Quinoxalines; Rats; Rats, Sprague-Dawley; Sodium; Sodium-Calcium Exchanger; Thiourea; Vasoconstriction; Vasoconstrictor Agents; Vasodilator Agents

Calcitonin gene-related peptide (CGRP), a capsaicin-sensitive neuromodulator of splanchnic vascular tone in several animal species, remains poorly investigated in mouse models. We therefore assessed whether endogenous CGRP is a non-adrenergic/non-cholinergic (NANC) neuromodulator in the mesenteric vascular bed of the mouse.. Arterial and venous changes in perfusion pressure in response to perivascular nerve stimulation (PNS) were monitored in the mouse mesenteric bed under basal conditions or precontracted with KCl (artery) or U46619 (vein) in circuits pretreated with guanethidine, atropine, indomethacin and prazosin. Arterial responses to NANC were also characterized with a CGRP1 antagonist, halphaCGRP8-37. Finally, the PNS-induced release of arterial CGRP was measured by enzyme immunoassay.. HalphaCGRP8-37 enhanced PNS-induced arterial increases in perfusion pressure under basal tone. PNS-induced stimulation of NANC triggered an halphaCGRP8-37 or capsaicin- sensitive reduction in perfusion pressure of the pre-contracted arterial bed only. Chemical removal of the endothelium inhibited PNS- and halphaCGRP- induced reduction in perfusion pressure in the arterial mesenteric bed. Responses to NANC nerves were reduced by guanylate and adenylate cyclase inhibitors (1H-[1,2,4]oxadiazole[4,3-a] quinoxalin-1-one (ODQ)) and [9-(tetrahydro-2-furanyl)-9H-purin-6-amine] (SQ 22,536), respectively. A neuronal NOS inhibitor (7-nitroindazole; 7-NI) also enhanced the response to NANC in vessels from wild-type, eNOS KO but not iNOS KO mice. Finally, PNS enhanced the release of immunoreactive CGRP from the perfused arterial mesenteric bed.. Our study demonstrates a role for CGRP in the NANC-dependent reduction in perfusion pressure of the arterial but not venous mesenteric bed of the mouse.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenine; Animals; Calcitonin Gene-Related Peptide; Electric Stimulation; Endothelium, Vascular; Female; Male; Mesenteric Arteries; Mesenteric Veins; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Organ Culture Techniques; Oxadiazoles; Perfusion; Peripheral Nerves; Potassium Chloride; Quinoxalines; Splanchnic Circulation

The aim of the present study was to investigate the involvement of adenosine 3',5'-cyclic monophosphate (cAMP) cascade in the acute impairment of contraction by 17beta-estradiol in porcine coronary arteries, and to elucidate the signaling pathway leading to the activation of this cascade by the hormone. Isometric tension was recorded in isolated rings of porcine coronary arteries. The contraction to U46619 was reduced significantly following 30 min incubation with 1 nM 17beta-estradiol or 1 nM isoproterenol. There was no additive effect when 17beta-estradiol and isoproterenol were administered together. The effect of 17beta-estradiol was mimicked by both the cyclic AMP analogue 8-Br-cAMP and the guanosine 3',5'-cyclic monophosphate (cyclic GMP) analogue 8-Br-cGMP. In rings with and without endothelium, the modulatory effect of 17beta-estradiol was abolished by the adenylyl cyclase inhibitor, SQ 22536, but was unaffected by the guanylyl cyclase inhibitor, ODQ. Both the cAMP antagonist Rp-8-Br-cAMPS and the cGMP antagonist inhibitor Rp-8-Br-cGMPS inhibited the effect of 17beta-estradiol. The effect of 17beta-estradiol was unaffected by the protein kinase A inhibitor, KT5720, but was abolished by the protein kinase G (PKG) inhibitor, KT5823, which also abolished the effect of isoproterenol. These data support our earlier findings that 17beta-estradiol (1 nM) acutely impairs contractile responses of porcine coronary arteries in vitro. This acute effect of 17beta-estradiol involves cAMP in vascular smooth muscles and the activation of PKG.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; 8-Bromo Cyclic Adenosine Monophosphate; Adenine; Adenylyl Cyclase Inhibitors; Animals; Carbazoles; Coronary Vessels; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Drug Interactions; Estradiol; Indoles; Isometric Contraction; Isoproterenol; Muscle, Smooth, Vascular; Swine; Thionucleotides; Time Factors

Responses to human CGRP, adrenomedullin (ADM), and proadrenomedullin NH2-terminal 20 peptide (PAMP) were studied in small human thymic arteries. CGRP, ADM, and PAMP produced concentration-dependent vasodilator responses in arteries preconstricted with the thromboxane mimic U-46619. Responses to ADM and PAMP were attenuated, whereas responses to CGRP were not altered by endothelial denudation. Inhibitors of nitric oxide synthase and guanylyl cyclase attenuated responses to ADM and PAMP but not to CGRP. The CGRP1 receptor antagonist CGRP(8-37) attenuated responses to CGRP and ADM but not to PAMP. Responses to CGRP were reduced by SQ-22536 and Rp-cAMPS, inhibitors of adenylyl cyclase and PKA. These data suggest that responses to CGRP and ADM are mediated by CGRP(8-37)-sensitive receptors and that the endothelial ADM receptor induces vasodilation by a nitric oxide-guanylyl cyclase mechanism, whereas a smooth muscle CGRP receptor signals by a cAMP-dependent mechanism. A different endothelial receptor recognizes PAMP and signals by a nitric oxide-dependent mechanism.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adenine; Adenylyl Cyclase Inhibitors; Adrenomedullin; Arteries; Calcitonin Gene-Related Peptide; Cyclic AMP; Dose-Response Relationship, Drug; Endothelium, Vascular; Guanylate Cyclase; Humans; Nitric Oxide; Nitric Oxide Synthase; Peptide Fragments; Peptides; Proteins; Receptors, Calcitonin Gene-Related Peptide; Thionucleotides; Thymus Gland; Vasodilation