| 1. |
- Zygmunt, Peter M., et al.
(författare)
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Interactions between endothelium-derived relaxing factors in the rat hepatic artery : Focus on regulation of EDHF
- 1998
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Ingår i: British Journal of Pharmacology. - : Wiley. - 0007-1188. ; 124:5, s. 992-1000
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Tidskriftsartikel (refereegranskat)abstract
- 1. In rat isolated hepatic arteries contracted with phenylephrine, acetylcholine and the calcium ionophore A23187 each elicit endothelium-dependent relaxations, which involve both nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF). However, the contribution of prostanoids to these responses, and the potential interaction between EDHF and other endothelium-derived relaxing factors have not been examined. 2. In the presence of the NO synthase inhibitor N(G)-nitro-L-arginine (L-NOARG, 0.3 mM) and a mixture of charybdotoxin (0.3 μM) and apamin (0.3 μM), inhibitors of the target potassium (K) channel(s) for EDHF, acetylcholine and A23187 each induced a concentration-dependent and almost complete relaxation, which was abolished in the additional presence of indomethacin (10 μM). Thus, in addition to EDHF and NO, a relaxing factor(s) generated by cyclo-oxygenase (COX) contributes to endothelium-dependent relaxation in the rat hepatic artery. 3. The resting membrane potentials of endothelium-intact and endothelium-denuded vascular segments were -57 mV and -52 mV, respectively (P > 0.05). In intact arteries, the resting membrane potential was not affected by L-NOARG plus indomethacin, but reduced to -47 mV in the presence of charybdotoxin plus apamin. Acetylcholine and A23187 (10 μM each) elicited a hyperpolarization of 13 mV and 15 mV, respectively. The hyperpolarization induced by these agents was not affected by L-NOARG plus indomethacin (12 mV and 14 mV, respectively), but reduced in the presence of charybdotoxin plus apamin (7 mV and 10 mV, respectively), and abolished in the combined presence of charybdotoxin, apamin and indomethacin. 4. The NO donor 3-morpholino-sydnonimine (SIN-1) induced a concentration-dependent relaxation, which was unaffected by charybdotoxin plus apamin, but abolished by the selective soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ, 10 μM). SIN-1 (10 μM) did not alter the resting membrane potential in endothelium-denuded vascular segments. 5. The COX-dependent relaxation induced by acetylcholine was abolished following exposure to 30 mM KCl, but unaffected by glibenclamide (10 μM). The prostacyclin analogue iloprost induced a concentration-dependent relaxation, which was also abolished in 30 mM KCl and unaffected by the combined treatment with glibenclamide, charybdotoxin and apamin. Iloprost (10 μM) induced a glibenclamide-resistant hyperpolarization (8 mV with and 9 mV without glibenclamide) in endothelium-denuded vascular segments. 6. Exposure to SIN-1 or iloprost did not affect the EDHF-mediated relaxation induced by acetylcholine (i.e. in the presence of L-NOARG and indomethacin). Replacement of L-NOARG with the NO scavenger oxyhaemoglobin (10 μM) or the soluble guanylate cyclase inhibitor ODQ (10 μM) or methylene blue (10 μM), which all significantly inhibited responses to endothelium-derived NO, did not affect the acetylcholine-induced relaxation in the presence of indomethacin, indicating that endogenous NO also does not suppress EDHF-mediated responses. 7. These results show that, in addition to EDHF and NO, an endothelium-derived hyperpolarizing factor(s) generated by COX contributes significantly to endothelium-dependent relaxation in the rat heptic artery. Neither this factor nor NO seems to regulate EDHF-mediated responses. Thus, EDHF does not serve simply as a 'back-up' system for NO and prostacyclin in this artery. However, whether EDHF modulates the NO and COX pathways remains to be determined.
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| 2. |
- Zygmunt, Peter M., et al.
(författare)
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Involvement of voltage-dependent potassium channels in the EDHF-mediated relaxation of rat hepatic artery
- 1997
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Ingår i: British Journal of Pharmacology. - : Wiley. - 0007-1188. ; 121:1, s. 141-149
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Tidskriftsartikel (refereegranskat)abstract
- 1. In the rat hepatic artery, the acetylcholine-induced relaxation mediated by endothelium-derived hyperpolarizing factor (EDHF) is abolished by a combination of apamin and charybdotoxin, inhibitors of small (SK(Ca)) and large (BK(Ca)) conductance calcium-sensitive potassium (K)-channels, respectively, but not by each toxin alone. The selective BK(Ca) inhibitor iberiotoxin cannot replace charybdotoxin in this combination. Since delayed rectifier K-channels (K(V)) represent another target for charybdotoxin, we explored the possible involvement of K(V) in EDHF-mediated relaxation in this artery. 2. The K(V) inhibitors, agitoxin-2 (0.3 μM), kaliotoxin (0.3 μM), β-dendrotoxin (0.3 μM), dofetilide (1 μM) and terikalant (10 μM), each in combination with apamin (0.3 μM) had no effect on the EDHF-mediated relaxation induced by acetylcholine in the presence of N(ω)-nitro-L-arginine (0.3 mM) and indomethacin (10 μM), inhibitors of nitric oxide (NO) synthase and cyclo-oxygenase, respectively (n = 2-3). Although the K(V) inhibitor margatoxin (0.3 μM) was also without effect (n = 5), the combination of margatoxin and apamin produced a small inhibition of the response (pEC50 and E(max) values were 7.5 ± 0.0 and 95 ± 1% in the absence and 7.0 ± 0.1 and 81 ± 6% in the presence of margatoxin plus apamin, respectively; n = 6; P < 0.05). 3. Ciclazindol (10 μM) partially inhibited the EDHF-mediated relaxation by shifting the acetylcholine-concentration-response curve 12 fold to the right (n =.6; P < 0.05) and abolished the response when combined with apamin (0.3 μM; n = 6). This combination did not inhibit acetylcholine-induced relaxations mediated by endothelium-derived NO (n = 5). 4. A 4-aminopyridine-sensitive delayed rectifier current (I(K(V))) was identified in freshly-isolated single smooth muscle cells from rat hepatic artery. None of the cells displayed a rapidly-activating and -inactivating A-type current. Neither charybdotoxin (0.3 μM; n = 3) nor ciclazindol (10 μM; n = 5), alone or in combination with apamin (0.3 μM; n = 4-5), had an effect on I(K(V)). A tenfold higher concentration of ciclazindol (0.1 mM, n = 4) markedly inhibited I(K(V)), but this effect was not increased in the additional presence of apamin (0.3 μM; n = 2). 5. By use of membranes prepared from rat brain cortex, [125I]-charybdotoxin binding was consistent with an interaction at a single site with a K(D) of approximately 25 pM. [125I]-charybdotoxin binding was unaffected by iberiotoxin (0.1 μM, n = 6), but was increased by apamin in a concentration-dependent manner (E(max) 43 ± 10%, P < 0.05 and pEC50 7.1 ± 0.2; n = 7-8). Agitoxin-2 (10 nM) displaced [125I]-charybdotoxin binding by 91 ± 3% (n = 6) and prevented the effect of apamin (1 μM; n = 6). 6. It is concluded that the EDHF-mediated relaxation in the rat hepatic artery is not mediated by the opening of either K(V) or BK(Ca). Instead, the target K-channels for EDHF seem to be structurally related to both K(V) and BK(Ca). The possibility that a subtype of SK(Ca) may be the target for EDHF is discussed.
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| 3. |
- Zygmunt, Peter M., et al.
(författare)
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Vanilloid receptors on sensory nerves mediate the vasodilator action of anandamide
- 1999
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 400:6743, s. 452-457
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Tidskriftsartikel (refereegranskat)abstract
- The endogenous cannabinoid receptor agonist anandamide is a powerful vasodilator of isolated vascular preparations, but its mechanism of action is unclear. Here we show that the vasodilator response to anandamide in isolated arteries is capsaicin-sensitive and accompanied by release of calcitonin- gene-related peptide (CGRP). The selective CGRP-receptor antagonist 8-37 CGRP (ref. 5), but not the cannabinoid CB1 receptor blocker SR141716A (ref. 7), inhibited the vasodilator effect of anandamide. Other endogenous. (2- arachidonylglycerol, palmitylethanolamide) and synthetic (HU 210, WIN 55,212- 2, CP 55,940) CB1 and CB2 receptor agonists could not mimic the action of anandamide. The selective 'vanilloid receptor' antagonist capsazepine inhibited anandamide-induced vasodilation and release of CGRP. In patch-clamp experiments on cells expressing the cloned vanilloid receptor (VR1), anandamide induced a capsazepine-sensitive current in whole cells and isolated membrane patches. Our results indicate that anandamide induces vasodilation by activating vanilloid receptors on perivascular sensory nerves and causing release of CGRP. The vanilloid receptor may thus be another molecular target for endogenous anandamide, besides cannabinoid receptors, in the nervous and cardiovascular systems.
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