Nicorandil-induced Relaxation Research Articles

In vitro effect of nicorandil on the carbachol-induced contraction of the lower esophageal sphincter of the rat

The lower esophageal sphincter (LES) is a specialized region of the esophageal smooth muscle that allows the passage of a swallowed bolus into the stomach. Nitric oxide (NO) plays a major role in LES relaxation. Nicorandil possesses dual properties of a NO donor and an ATP-sensitive potassium channel (KATP channel) agonist, and is expected to reduce LES tone. This study investigated the mechanisms underlying the effects of nicorandil on the LES. Rat LES tissues were placed in an organ bath, and activities were recorded using an isometric force transducer. Carbachol-induced LES contraction was significantly inhibited by KATP channel agonists in a concentration-dependent manner; pinacidil >> nicorandil ≈ diazoxide. Nicorandil-induced relaxation of the LES was prevented by pretreatment with glibenclamide, whereas NG-nitro-l-arginine methyl ester (l-NAME), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and iberiotoxin were ineffective at preventing nicorandil-induced LES relaxation. Furthermore, nicorandil did not affect high K+-induced LES contraction. Reverse-transcription polymerase chain reaction analysis and immunohistochemistry revealed expression of KCNJ8 (Kir6.1), KCNJ11 (Kir6.2), ABCC8 (SUR1) and ABCC9 (SUR2) subunits of the KATP channel in the rat lower esophagus. These findings indicate that nicorandil causes LES relaxation chiefly by activating the KATP channel, and that it may provide an additional pharmacological tool for the treatment of spastic esophageal motility disorders.

Nicorandil directly and cyclic GMP-dependently opens K+ channels in human bypass grafts

As we previously demonstrated the role of different K+ channels in the action of nicorandil on human saphenous vein (HSV) and human internal mammary artery (HIMA), this study aimed to analyse the contribution of the cGMP pathway in nicorandil-induced vasorelaxation and to determine the involvement of cGMP in the K+ channel-activating effect of nicorandil. An inhibitor of soluble guanylate cyclase (GC), ODQ, significantly inhibited nicorandil-induced relaxation, while ODQ plus glibenclamide, a selective ATP-sensitive K+ (KATP) channel inhibitor, produced a further inhibition of both vessels. In HSV, ODQ in combination with 4-aminopyridine, a blocker of voltage-gated K+ (KV) channels, did not modify the concentration-response to nicorandil compared with ODQ, whereas in HIMA, ODQ plus iberiotoxin, a selective blocker of large-conductance Ca2+-activated K+ (BKCa) channels, produced greater inhibition than ODQ alone. We showed that the cGMP pathway plays a significant role in the vasorelaxant effect of nicorandil on HSV and HIMA. It seems that nicorandil directly opens KATP channels in both vessels and BKCa channels in HIMA, although it is possible that stimulation of GC contributes to KATP channels activation in HIMA. Contrary, the activation of KV channels in HSV is probably due to GC activation and increased levels of cGMP.

PP098—Nicorandil induced endothelium-independent relaxation of arterial bypass graft

2013 e47 PP098—NICORANDIL INDUCED ENDOTHELIUMINDEPENDENT RELAXATION OF ARTERIAL BYPASS GRAFT M. Marinko; A. Novakovic; I. Stojanovic; P. Milojevic; M. Jovic; D. Nenezic; N. Ugresic; V. Kanjuh; Q. Yang; and G.-W. He Department of Pharmacology, Faculty of Pharmacy, University of Belgrade; Faculty of Medicine, University of Belgrade; Institute for Cardiovascular Diseases “Dedinje”; Academy of Sciences and Arts, Belgrade, Serbia; Department of Surgery, Chinese University of Hong Kong, Hong Kong, Hong Kong; TEDA International Cardiovascular Hospital, Medical College, Nankai University, Tianjin, China; and Providence Heart & Vascular Institute, Albert Starr Academic Center, Department of Surgery, Oregon Health and Science University, Portland, Oregon Introduction: Spasm of the human internal mammary artery (HIMA) is a rare but life-threatening complication after coronary artery bypass grafting (CABG). The reversal of this vasospasm is often challenging, and the most effective therapy is not well defined. The present study was aimed to investigate vasorelaxant effect of nicorandil, K+ channel opener, on the HIMA and to define the role of different K+ channel subtypes in nicorandil action on this blood vessel. Patients (or Materials) and Methods: Discarded segments of HIMA were collected from patients undergoing CABG and studied in organ baths. HIMA rings were precontracted with phenylephrine (10 μM) followed by cumulatively adding increasing doses of nicorandil. The endothelium was removed mechanically. Results: Nicorandil (0.001 μM–300 μM) induced a concentrationdependent relaxation of HIMA rings precontracted by phenylephrine. Glibenclamide (10 μ M), a highly selective blocker of ATP-sensitive K+ (K ATP ) channels, partially inhibited relaxation of HIMA induced by nicorandil. Tetraethylammonium (TEA, 1 mM), a nonselective blocker of Ca 2+ -activated K+ (K Ca ) channels, as well as iberiotoxin (100 nM), a most selective blocker of large-conductance K Ca (BK Ca ) channels, partly antagonized relaxation of HIMA. A nonselective blocker of voltage-gated K+ (K V ) channels, 4-aminopyridine (4-AP, 0.5 mM), as well as margatoxin (10 nM), a potent inhibitor of K V 1.3 channels, did not significantly modify the nicorandil-induced relaxation of HIMA. Conclusion: The results from our study demonstrate that, in HIMA, nicorandil has a potent vasorelaxant effect which is endotheliumindependent. It seems that mechanism of this relaxation includes KATP and 4-AP-sensitive K+ channels located in the smooth muscle of HIMA. Disclosure of Interest: None declared.

Different K+ Channels Are Involved in Relaxation of Arterial and Venous Graft Induced by Nicorandil

The drug nicorandil is a vasodilator approved for the treatment of angina. In addition to its well-known effect on the opening of ATP-sensitive K (KATP) channels, nicorandil-induced vasorelaxation also involves the opening of Ca-activated K channels. The aim of this study was to investigate the effects of nicorandil on the isolated human internal mammary artery (HIMA) and the human saphenous vein (HSV) and to define the contribution of different K channel subtypes in the nicorandil action on these arterial and venous grafts. Our results show that nicorandil induced a concentration-dependent relaxation of HSV and HIMA rings precontracted by phenylephrine. Glibenclamide, a selective KATP channels inhibitor, partially inhibited the response to nicorandil in both HSV and HIMA. Iberiotoxin, a most selective blocker of large-conductance Ca-activated K (BKCa) channels, partly antagonized relaxation of HIMA. A nonselective blocker of voltage-gated K channels, 4-aminopyridine caused partial inhibition of the nicorandil-induced relaxation of HSV but did not antagonize relaxation of HIMA induced by nicorandil. Margatoxin, a potent inhibitor of KV1.3 channels, did not abolish the effect of nicorandil on HSV and HIMA. Our results showed that nicorandil induced strong endothelium-independent relaxation of HSV and HIMA contracted by phenylephrine. It seems that KATP and 4-aminopyridine-sensitive K channels located in the smooth muscle of HSV mediated relaxation induced by nicorandil. In addition, KATP and BKCa channels are probably involved in the nicorandil action on HIMA.

Guanylate cyclase and not ATP-dependent K + channels seems temperature-dependent in smooth muscle relaxation of human umbilical arteries

The effects of K + channel opener, nicorandil [ N-(2-hydroxyethyl)-nicotinamide nitrate], on isolated human umbilical arteries were investigated at two temperatures: 37°C and 25°C. The purpose of this investigation was: (1) to confirm the relaxant effect of nicorandil, (2) to elucidate the influence of endothelium and temperature on nicorandil-induced relaxation, (3) to determine which of guanylate cyclase or ATP-sensitive K + channels was implicated in temperature-induced relaxation of smooth muscles. Rings, 3-mm-wide, were suspended in organ chambers for isometric force measurement. All solutions were aerated with 95% O 2–5% CO 2 and maintained at 37°C or 25°C (cooling), pH 7.4. The presence of an intact endothelium was confirmed by immunohistochemistry. During the first set of experiments after contraction with 5-hydroxytryptamine (5-HT 10 −5 M), nicorandil (10 −9–10 −4 M) was added to the organ chambers with controls and in with rings incubated with l-arginine, N-nitro- l-arginine ( l-NNA) an inhibitor of nitric oxide (NO) synthase, [1- H-(1,2,4) oxadiazole (4,3-a) quinoxalin-1-one] (ODQ), a specific inhibitor of guanylate cyclase, or glibenclamide, an antagonist of nicorandil, all at 10 −5 M. In another set of experiments, rings were contracted with 5-HT (10 −5 M) and relaxed with 3-morpholinosydnonimine [SIN-1 (10 −9–3×10 −5 M) or cromakalim (10 −9–3×10 −5 M)]. Our results showed that nicorandil induced concentration-dependent relaxation. At 37°C, in the control, the maximum relaxation was 90±5%, and 60±8% at 25°C ( P<0.01). However, the relaxation at 37°C or 25°C remained unchanged after pretreatment with l-arginine, l-NNA, this suggests that the same concentration of drugs used in this type of vessel does not appear to modulate the relaxant effect of nicorandil. On the other hand, we observed that the relaxant effect of SIN-1 was 72±5% at 37°C and only 28±7% at 25°C ( P<0.01). However, relaxations with cromakalim were partly influenced by cooling. In the presence of ODQ, the nicorandil-induced relaxation observed at 37°C or 25°C was less than that in the control and in the rings incubated with glibenclamide. These results for human umbilical arteries indicate that cooling decreases the relaxation response of smooth muscles and that this is partly due to a decreased response to guanylate cyclase.

Role of K+ channel opening and stimulation of cyclic GMP in the vasorelaxant effects of nicorandil in isolated piglet pulmonary and mesenteric arteries: relative efficacy and interactions between both pathways.

1. The effects of the K+ channel opener levcromakalim, the guanylate cyclase stimulant nitroprusside and the dual drug nicorandil (K+ channel opener and guanylate cyclase stimulant) were analysed in piglet isolated endothelium-denuded pulmonary (PA) and mesenteric (MA) arteries stimulated by noradrenaline (NA) or by the thromboxane A2 mimetic U46619. 2. Nicorandil, levcromakalim and verapamil were less potent in PA than in MA, the efficacy of levcromakalim was also reduced in PA. The effects of nicorandil and levcromakalim were similar in arteries pre-contracted by NA and U46619, whereas verapamil was more potent in arteries pre-contracted by NA. Nitroprusside was equipotent in MA pre-contracted by either NA or U46619 and in PA pre-contracted by NA whereas in PA pre-contracted by U46619, nitroprusside showed lower potency and efficacy. 3. The relaxant effects of levcromakalim and nitroprusside were inhibited by 10(-5) M glibenclamide and 10(-6) M ODQ, respectively. Nicorandil-induced relaxation was inhibited by ODQ in all experimental conditions, whereas glibenclamide had inhibitory effects in PA and MA pre-contracted by U46619, had no effect in PA pre-contracted by NA and in MA pre-contracted by NA it was only inhibitory in the presence of ODQ. 4. No apparent interactions were found between nitroprusside and levcromakalim as indicated by the lack of effects of pretreatment with one of them (producing 20-35% relaxation) on the potency of the relaxant response to the other. However, in PA pre-contracted by U46619, where nitroprusside or levcromakalim induced only partial relaxation, the combination of both mechanisms (either by combining nitroprusside plus levcromakalim or by nicorandil) was able to induce full vasodilatation. 5. In conclusion, K+ channel opening and guanylate cyclase stimulation are independent pathways that induce additive vasorelaxation in piglet PA and MA. The mechanism of action of nicorandil is dependent on the artery and on the nature of the agonist employed to precontract the artery. The relative efficacy of K+ channel opening vs guanylate cyclase stimulation may partially explain the preferential contribution of each mechanism to the relaxant effects of nicorandil.

Analysis of relaxation and repolarization mechanisms of nicorandil in rat mesenteric artery.

1. The mechanisms by which nicorandil causes relaxation of rat isolated small mesenteric arteries mounted on a Mulvany myograph was investigated by use of a combination of putatively mechanism-specific antagonists. 2. In arteries precontracted by the thromboxane-mimetic, U46619, the EC50 for cromakalim and levcromakalim-induced relaxation curves were raised by 36 and 17 fold by glibenclamide (3 microM) while the EC50 for nicorandil-induced relaxation was unaffected by either glibenclamide or methylene blue (10 microM). A combination of these antagonists raised the EC50 for nicorandil by 8 fold. 3. In U46619-contracted arteries, nifedipine (100 nM) did not affect the cromakalim relaxation curve but it raised the EC50 for nicorandil by 5 fold. The combination of methylene blue, glibenclamide and nifedipine further inhibited the maximum relaxation to nicorandil. 4. In separate experiments, membrane potential (Em) and force responses were measured simultaneously. In arteries depolarized and contracted by U46619 both nicorandil and cromakalim repolarized (delta Em, 35 mV) and relaxed (100%) the vessels. Glibenclamide (3 microM) did not alter the relaxation-concentration curve to nicorandil but reduced the maximum repolarization to delta 10.8 mV. In contrast to Em and relaxation-response curves to cromakalim were shifted to the right by glibenclamide by 30-100 fold. 5. In unstimulated arteries, nicorandil (but not cromakalim) -induced hyperpolarization was significantly antagonized by methylene blue (10 microM) (6.6 fold rightward shift) or nifedipine (100 nM) (2.6 fold). In depolarized arteries (U46619), nifedipine but not methylene blue inhibited the nicorandil-induced hyperpolarization. 6. In arteries precontracted to 50% tissue maximum by either KCl or U46619, nifedipine (100 nM) relaxed the artery but failed to repolarize the Em. Presumably voltage-operated calcium channels (VOCC) were blocked preventing contraction but the artery remained depolarized, presumably through non VOCC mechanisms. 7. These data suggest that nicorandil may relax small arteries through 3 parallel pathways, (i) NO-donor mediated stimulation of guanylate cyclase and increase in cyclic GMP, (ii) K+ATP channel opening, and (iii) nifedipine-sensitive VOCC inhibition. Em data suggest that nicorandil-induced repolarization is caused principally through opening K+ATP channels. Blockade of this hyperpolarization by glibenclamide is not sufficient to alter the relaxation, indicating dissociation of nicorandil-induced changes in membrane potential and relaxation. 8. These results highlight the 'chameleon' actions of nicorandil where there is no apparent association of Em repolarization with relaxation, in contrast to the parallel responses for cromakalim.

Effects of Nicorandil on Human Isolated Corpus Cavernosum and Cavernous Artery

Nitric oxide (NO) released from nonadrenergic-noncholinergic (NANC) nerves seems to be a principal mediator of the relaxation of penile erectile tissue necessary for erection, and drugs acting by release of NO have been shown to produce erection when injected intracorporeally into impotent patients. By producing hyperpolarization, K+ channel openers are effective in relaxing isolated penile erectile tissue from rabbit and man, and can produce tumescence and erection when injected intracorporeally into animals. Nicorandil is classified as a K+ channel opener, but it also acts as a donor of NO. In the present study, the effects of nicorandil on isolated preparations from human corpus cavernosum (CC) and deep cavernous artery (Acc) were compared with those of cromakalim (K+ channel opener) and SIN-1 (NO donor). Nicorandil produced a concentration-dependent relaxation of CC and Acc preparations. The relaxations obtained at the highest nicorandil concentration used (10−4 M.) were 75 ± 3% and 66 ± 4% in CC preparations contracted by noradrenaline and endothelin-1, respectively. The corresponding effects in Acc preparations were 70 ± 14% and 73 ± 5%. Glibenclamide (blocking ATP-dependent K+ channels) significantly reduced the nicorandil-induced relaxation in CC, but not in Acc. Methylene blue (believed to block soluble guanylate cyclase) reduced nicorandil’s relaxant effect in CC, although statistical significance was not obtained. NG-nitro-L-arginine 10−4 M. (NO synthase inhibitor) did not significantly influence the effect of nicorandil on precontracted preparations in either tissue. In CC preparations contracted by electrical field stimulation, nicorandil and cromakalim concentration dependently inhibited the responses. This effect was significantly counteracted by glibenclamide. It is concluded that nicorandil is effective in relaxing human CC chiefly by its K+ channel opening action, and to some extent by its ability to release NO. For nicorandil’s relaxing effect on Acc, ATP dependent K+ channels seem to be of limited importance. If effective in impotent patients, the drug may represent a new, interesting approach to the treatment of erectile dysfunction.

Pharmacological interaction experiments differentiate between glibenclamide-sensitive K + channels and cyclic GMP as components of vasodilation by nicorandil

The relaxant effect of the vasodilator drug, nicorandil, was studied in circular strips of bovine coronary arteries. To differentiate between relaxation caused by cyclic GMP (cGMP) and by hyperpolarization, the influence of cGMP was blocked with methylene blue and that of hyperpolarization with the inhibitor of ATP-dependent K + channels, glibenclamide. Methylene blue and glibenclamide inhibited nicorandil-induced relaxation to similar extents. Cromakalim-induced relaxation but not that due to sodium nitroprusside (nitroprusside-Na) was inhibited by glibenclamide. Methylene blue inhibited the relaxation caused by nitroprusside-Na but not that due to cromakalim. The different modes of action of the two components of relaxation caused by nicorandil were studied in agonist-agonist interaction experiments. The interaction between nicorandil and nitroprusside-Na or 3-morpholino-sydnonimine (SIN-1) was overadditive in the absence of glibenclamide but additive, i.e. competitive, in the presence of glibenclamide. The interaction of nicorandil with cromakalim or pinacidil was overadditive in the absence of methylene blue but additive, i.e. competitive, in the presence of methylene blue. The results show that nicorandil relaxes smooth muscle through two independent mechanisms: ATP-dependent activation of K + channels and stimulation of guanylyl cyclase resulting in increases in cGMP.

Molecular mechanism of action of nicorandil.

Nicorandil relaxes coronary vascular smooth muscle by stimulating guanylyl cyclase and increasing cyclic GMP (cGMP) levels (as shown first in our laboratory) as well as by a second mechanism resulting in activation of K+ channels and hyperpolarization. Therefore, we studied the relative contributions of either mechanism to the overall response in bovine circular strips of coronary arteries by simultaneously measuring changes in length and in cGMP levels through radioimmunoassay. Blockade by 10 microM methylene blue of the cGMP increases in strips precontracted by 1 microM of the thromboxane A2 analogue U46619 reduced nicorandil-induced relaxation to 30-50%, and there were no significant changes in cGMP levels. Suppression of the hyperpolarizing component of nicorandil by 80.4 mM K+ or 1 microM glibenclamide in precontracted strips reduced nicorandil relaxation to 50% (K+) or shifted the dose response to the right by a factor of two (glibenclamide) without alteration of increases in cGMP. A quantitative separation of both mechanisms of action was obtained by comparing the correlation between increases in cGMP and relaxation under conditions of inhibited versus noninhibited hyperpolarization. The results indicate that cGMP contributes to the total relaxing effect of nicorandil by 30-40% at low concentrations and 80-90% at high concentrations of nicorandil. From the experiments with glibenclamide, it can be concluded that the probable mechanism by which nicorandil hyperpolarizes is opening glibenclamide-sensitive K+ channels in coronary vascular smooth muscle and that this latter effect mimics those of other K+ channel openers such as cromakalim or pinacidil.

Analysis of cromakalim-, pinacidil-, and nicorandil-induced relaxation of the 5-hydroxytryptamine precontracted rat isolated basilar artery

The effects of the K+ channel activators cromakalim, pinacidil, and nicorandil were investigated in endothelium intact, 5-hydroxytryptamine (5-HT) precontracted rat isolated basilar artery. Cromakalim, pinacidil, and nicorandil produced concentration-dependent relaxation of rat isolated basilar artery precontracted with 5-HT with a rank order of potency of cromakalim greater than pinacidil greater than nicorandil. All compounds produced full or nearly full relaxation. The calculated Hill coefficients for cromakalim-, pinacidil-, and nicorandil-induced relaxation of 5-HT-precontracted rat isolated basilar artery were 2.20 +/- 0.36, 1.30 +/- 0.07, and 1.00 +/- 0.01, respectively. Under conditions of increased tone produced by 50 mmol/l KCl (which inhibits cromakalim-induced relaxation) pinacidil and nicorandil produced marked reversal of spasm, with pinacidil being more potent than nicorandil. In arteries precontracted with 5-HT, preincubation with glibenclamide (0.1-1 mumol/l) produced concentration-related inhibition of relaxation with calculated mean pA2 values (and slopes of Schild regression) +/- SEM of 6.84 +/- 0.20 (1.1 +/- 0.20) against cromakalim. 6.60 +/- 0.14 (0.95 +/- 0.23) against nicorandil, and 6.57 +/- 0.26 (1.04 +/- 0.18) against pinacidil. For cromakalim, pinacidil, and nicorandil the slopes of Schild regression were not significantly different from unity. Tolbutamide 10 mumol/l was without effect against the cromakalim-, pinacidil-, or nicorandil-induced relaxation. Tetraethylammonium (TEA; 1-10 mmol/l) produced noncompetitive inhibition of the cromakalim-induced relaxation, but appeared to produce competitive inhibition of the pinacidil- and nicorandil-induced relaxations. We conclude that cromakalim, pinacidil, and nicorandil produce relaxation of the 5-HT precontracted rat basilar artery by similar mechanisms to those identified in other peripheral vascular and visceral smooth muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

P-152 - Nicorandil-Induced Relaxation of Rabbit Femoral Arterial Strips.

P-152 - Nicorandil-Induced Relaxation of Rabbit Femoral Arterial Strips.

The relaxant action of nicorandil in guinea‐pig isolated trachealis

Nicorandil (1-1000 mumol l-1) caused concentration-dependent relaxation of guinea-pig isolated trachealis. Propranolol (1 mumol l-1) did not modify the relaxant action of nicorandil but antagonized isoprenaline. Among K+-channel inhibitors tested, apamin (0.1 mumol l-1) and procaine (5 mmol l-1) did not modify the relaxant action of nicorandil. In contrast, tetraethylammonium (TEA, 8 mmol l-1) caused five fold antagonism. Trachealis exposed to K+-rich (120 mmol l-1) Krebs solution developed near-maximal tension. Nicorandil relaxed the K+-depolarized tissue though its concentration-effect curve was shifted markedly to the right. In tissues in which tone was induced by histamine, methylene blue (100 mumol l-1) antagonized nicorandil and sodium nitroprusside but did not modify the relaxant action of aminophylline. Intracellular electrophysiological recording showed that nicorandil (1 mumol l-1) could evoke some relaxation in the absence of electrical changes. Higher concentrations (10-1000 mumol l-1) reduced the amplitude and frequency of spontaneous electrical slow waves. Nicorandil also caused concentration-dependent hyperpolarization and relaxation. When the hyperpolarization was sufficiently pronounced slow wave activity was abolished. TEA (8 mmol l-1) induced slow waves which were surmounted by a spike potential. TEA slightly reduced the maximal hyperpolarization induced by nicorandil and increased the time required for nicorandil to abolish slow wave discharge. Procaine (5 mmol l-1) induced slow waves of relatively low frequency. Sometimes these were surmounted by a spike potential Procaine markedly reduced the hyperpolarization induced by nicorandil and increased the time required for abolition of slow waves. In studies of the efflux of 86Rb+ from muscle-rich strips of trachea, nicorandil (1000 mumol l-1) increased the efflux rate constant, whereas isoprenaline (1 mumol l-1) was without effect. It is concluded that nicorandil-induced relaxation does not involve the activation of beta-adrenoceptors but is partly attributable to the formation of nitric oxide from the nitrate moiety in its molecular structure. Nicorandil can evoke relaxation in the absence of membrane potential change but towards the upper end of its effective concentration range, nicorandil increases membrane K+ conductance and thereby evokes hyperpolarization of trachealis cells. The K+ channels opened by nicorandil are permeable to 86Rb, insensitive to apamin and TEA but may be inhibited by procaine.