Role of endogenous hydrogen peroxide in the development of nitrate tolerance

The present study was designed to test the hypothesis that hydrogen peroxide plays a role in the development of nitrate tolerance. Isolated rat aortic rings were suspended in organ chambers for isometric tension recording. The rings were incubated with (tolerant) and without (control) nitroglycerin (10(-4) M) for 90 min, followed by repeated rinsing for 1 h. Hydrogen peroxide release in control and tolerant tissues was measured fluorimetrically using amplex red. Nitroglycerin (10(-9)-10(-4) M) caused concentration-dependent relaxations in control (-logEC50=7.15+/-0.1) and tolerant rings (-logEC50=5.83+/-0.1) contracted with norepinephrine. Nitrate tolerance was evident by a >20-fold rightward shift in the nitroglycerin concentration-response curve in tissues exposed previously to nitroglycerin for 90 min. Incubation of the rings with the superoxide dismutase (SOD)-mimetic, tempol (10(-4) M), during the 90-min exposure period to nitroglycerin caused a leftward shift in the nitroglycerin concentration-response curve in tolerant rings (-logEC50=6.84+/-0.2), but had no effect on the response to nitroglycerin in control rings. Treatment of the rings with catalase (1200 U/ml) or ebselen (1.5x10(-5) M), a glutathione peroxidase-mimetic, during the 90-min exposure period to nitroglycerin resulted in a further rightward shift in the nitroglycerin concentration-response curve in tolerant rings (-logEC50=5.41+/-0.1 and 4.98+/-0.1; catalase and ebselen respectively), without altering the response to nitroglycerin in control rings. In the presence of catalase, the effect of tempol on nitrate tolerance was abolished (-logEC50=5.46+/-0.1). Hydrogen peroxide release was reduced by approximately 64% in nitrate tolerant tissues when compared to control. The decrease in hydrogen peroxide release was completely reversed by treatment with tempol, whereas treatment with ebselen caused a further decrease in hydrogen peroxide release in nitrate tolerant tissues. Addition of hydrogen peroxide (3x10(-5) M) to nitrate tolerant rings caused a leftward shift in the nitroglycerin concentration-response curve in tolerant rings (-logEC50=7.18+/-0.3), but had no effect on the response to nitroglycerin in control rings. These results suggest that nitrate tolerance is associated with decreased endogenous formation of hydrogen peroxide, which attenuates nitrate tolerance development. SOD-mimetics may reduce nitrate tolerance, in part, by increasing the formation of hydrogen peroxide.     

Effect of resveratrol on nitrate tolerance in isolated human internal mammary artery

The present study aims to examine whether resveratrol, a natural antioxidant present in red wine, restores the tolerance to nitroglycerin (GTN) on isolated human internal mammary artery (IMA), using an in vitro model of nitrate tolerance. IMA rings were obtained from 53 male patients undergoing coronary bypass operation. Nitrate tolerance was induced by incubating the artery ring with 100 microM GTN for 90 minutes. Concentration-response curves to GTN (10(-9) to 10(-4) M) were obtained on IMA rings precontracted with noradrenaline. A low concentration (5 microM) of lucigenin was used as a tool to measure superoxide production in IMA segments. GTN produced concentration-dependent relaxation in isolated human IMA rings. Preexposure of artery rings to GTN reduced the relaxations to GTN [E(max) values: 105 +/- 2% and 76 +/- 3%, n = 10 to 12, P < 0.05; EC(50) values (-log M): 6.72 +/- 0.05 and 4.95 +/- 0.06, P < 0.05, respectively]. Relaxation to sodium nitroprusside remained unchanged. Diminished relaxation to GTN is partially restored after removing endothelium or L(G)-nitro-L-arginine (L-NOARG, 10 M) or superoxide dismutase (20 and 200 U/mL) or catalase (200 U/mL) pretreatments. Pretreatments with resveratrol (1, 10, and 20 microM) for 20 minutes relatively improved the reduced relaxation to GTN in tolerant IMA rings. Coadministration of L-NOARG with resveratrol did not abolish the beneficial effect of resveratrol on nitrate tolerance. The inhibitory effect of resveratrol on GTN-induced tolerance was not abolished in arterial rings without endothelium. Exposure to GTN increased superoxide production in IMA segments with endothelium. Endothelium denudation, L-NOARG, or superoxide dismutase pretreatments markedly inhibited the increased superoxide production in tolerant arteries. Resveratrol (1 and 10 microM) almost completely abolished basal or NAD(P)H-stimulated superoxide production in tolerant and nontolerant arteries. Vascular tolerance to GTN, in in vitro tolerant human IMA rings, can be induced by endothelial superoxide anions. Resveratrol partially restored the reduced relaxation to GTN by inhibiting NAD(P)-derived superoxide production in endothelium.     

Nitroglycerin-induced release of calcitonin gene-related peptide from sensory nerves attenuates the development of nitrate tolerance

The present study was designed to determine if endogenous calcitonin gene-related peptide (CGRP) affects the process of nitrate tolerance development in blood vessels. Rat aortic rings were suspended in organ chambers and relaxations to nitroglycerin (10(-9) -10(-6) M) were obtained in nitrate tolerant and nontolerant rings contracted with norepinephrine (10(-7) M). Tolerance was induced by incubating the rings with (tolerant) or without (nontolerant) nitroglycerin (10(-4) M) for 90 minutes, followed by repeated rinsing for 1 hour. Some rings were treated with CGRP8-37 (10(-6) M), glyburide (10(-6) M), or iberiotoxin (10(-7) M) during the 90-minute desensitization period with nitroglycerin (10(-4) M), and were then washed out during the 1-hour rinsing period. Other rings were treated with capsaicin (10(-5) M) prior to the 90-minute desensitization period. Calcitonin gene-related peptide release was measured by radioimmunoassay. Relaxation to nitroglycerin was markedly reduced in tolerant rings, as compared with nontolerant. Incubation with CGRP8-37 (10(-6) M) specifically during the 90-minute desensitization period with nitroglycerin resulted in even greater impairment in the response to nitroglycerin in tolerant rings, even though the calcitonin gene-related peptide antagonist had been washed out before completion of the nitroglycerin dose-response curve. Similar results were obtained following depletion of calcitonin gene-related peptide stores in sensory nerves by treatment with capsaicin (10(-5) M) prior to the 90-minute desensitization period with nitroglycerin. Prior treatment with CGRP8-37 or capsaicin had no effect on the response to nitroglycerin in nontolerant rings. Incubation with glyburide (10(-6) M), but not iberiotoxin (10(-7) M), specifically during the 90-minute desensitization period, mimicked the effect of CGRP8-37 and capsaicin in tolerant rings, suggesting a role for KATP channels in the effect of calcitonin gene-related peptide. Nitroglycerin (10(-4) M) caused a greater than twofold increase over basal levels in calcitonin gene-related peptide release in nontolerant rings, which was abolished in rings treated with capsaicin and in nitrate tolerant rings. These results suggest that nitroglycerin releases calcitonin gene-related peptide from sensory nerves during the process of desensitization to nitrovasodilators, and that interference with either the release or action of endogenous calcitonin gene-related peptide during this period enhances the extent to which nitrate tolerance occurs. The finding that nitroglycerin-induced release of calcitonin gene-related peptide from sensory nerves attenuates the desensitizing effect of nitroglycerin represents a heretofore unknown event in the development of nitrate tolerance, and demonstrates a novel role for calcitonin gene-related peptide in the vasculature.     

Role of cGMP-dependent protein kinase in development of tolerance to nitroglycerine in porcine coronary arteries

BACKGROUND AND PURPOSE: The cGMP-dependent protein kinase (PKG) is a key enzyme for nitrovasodilator-induced vasodilation. The present study was to determine its role in nitrate tolerance. EXPERIMENTAL APPROACH: isolated porcine coronary arteries were incubated for 24 h with nitroglycerin (NTG) and their relaxant responses were determined. PKG activity was assayed by measuring the incorporation of (32)P into BPDEtide. PKG protein was determined by Western blotting and PKG mRNA by real-time PCR. KEY RESULTS: A 24 h incubation with NTG attenuated relaxation of coronary arteries to NTG, which was associated with decreased PKG activity. The nitrate tolerance induced with NTG at 10(-7) M was affected by a scavenger of reactive oxygen species and the tolerance induced with NTG at 10(-6) and 10(-5) M showed cross-tolerance to DETA NONOate and 8-Br-cGMP (a cell permeable cGMP analogue). PKG protein and mRNA were down-regulated by a 24 h incubation with NTG at 10(-5) M but not at 10(-7) M. Acute exposure to exogenous superoxide inhibited PKG activity stimulated by NTG at 10(-7) M but not at 10(-5) M. Superoxide had no effect on PKG activity stimulated with exogenous cGMP. CONCLUSIONS AND IMPLICATIONS: Nitrate tolerance induced by NTG at low concentrations may result from an increased production of reactive oxygen species acting on sites upstream of PKG. The tolerance induced by NTG at higher concentrations may be in part due to suppression of PKG expression resulting from sustained activation of the enzyme. These distinct mechanisms of nitrate tolerance may be of clinical significance.     

Persistence of the response to SIN1 on isolated coronary arteries rendered tolerant to nitroglycerin in vitro or in vivo

Experiments were performed on isolated canine and human coronary arteries to provide more insight into the mechanisms responsible for the vascular tolerance to nitroglycerin that is induced under in vitro or in vivo conditions. In vitro tolerance was produced after an incubation of coronary ring segments with nitroglycerin (10 microM for 30 min at physiological pH). After elevation of tone with KCl (15 mM), dose-response curves were constructed for nitroglycerin or SIN1 (3-morpholino-syndnonimin) on control and tolerant rings. On canine tolerant rings the dose-response curve for nitroglycerin-induced relaxations was significantly (p less than 0.001) shifted to the right, and 50% of the maximal relaxation (ED50) increased from 55 +/- 9 nM to 1.2 +/- 0.2 microM. Pretreatment of tolerant rings with N-acetylcysteine (NAC, 10 microM 10 min before KCl-induced contraction) partially restored the responsiveness to nitroglycerin, with ED50 reducing to 0.56 +/- 0.03 microM (p less than 0.02). On the other hand, the dose-response curves to SIN1 were not significantly altered. Similar results were obtained on human preparations. On isolated canine coronary rings rendered tolerant in vivo by subcutaneous injections of 15 mg/kg nitroglycerin (two times daily for 4 consecutive days), ED50 for nitroglycerin was 0.67 +/- 0.08 microM (p less than 0.001 versus control rings), and NAC again partially restored the responsiveness to nitroglycerin. As for the in vitro tolerance, the relaxations to SIN1 were not significantly altered on these canine rings rendered nitrate tolerant in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparative vasorelaxing profiles of nicorandil, isosorbide dinitrate and nitroglycerin in isolated coronary arteries of the dog

The vasorelaxing effects of nicorandil (NCR), isosorbide dinitrate (ISDN) and nitroglycerin (NTG) were studied in isolated canine coronary arteries. In rings of coronary arteries precontracted with prostaglandin F2 alpha (3 x 10(-6) M) or KCl (30 mM), removal of the endothelium significantly augmented the relaxing effects of NCR, while it did not affect those of ISDN and NTG. In unrubbed rings precontracted with KCl (30 mM), methylene blue (5 x 10(-6) M) significantly inhibited vasorelaxing responses to the three drugs. The order of the inhibition was as follows: NTG greater than ISDN greater than NCR. When the unrubbed tissue was incubated with NTG (10(-5) M) or ISDN (10(-4) M) for 10 min, it developed acute tolerance in relaxing response to NTG or ISDN. Unlike NTG and ISDN, NCR did not develop any tolerance. The treatment with N-acetylcysteine (5 x 10(-5) M) tended to potentiate relaxant effects of NTG and to reduce the degree of acute tolerance to NTG. The results suggest that cGMP plays a role in the relaxation of the coronary artery induced by the drugs and furthermore that the mode of the vasorelaxing action of NCR may be somewhat different from that of NTG or ISDN.     

5-Hydroxytryptamine does not release endothelium-derived relaxing factor in rat isolated coronary arteries

Removal of the endothelium from isolated rat proximal and distal coronary artery segments shifted the 5-HT concentration-response curve to the left without affecting, the maximal contractile response. 5-HT had no relaxing effect in 10(-5) M prostaglandin F2 alpha-precontracted vessels with an intact endothelium in the presence of 10(-5) M ketanserin. The spontaneous myogenic tone increased in both proximal and distal coronary artery segments after the endothelium had been removed. Indomethacin (10(-5) M) reduced the response of the proximal coronary artery segments to 5-HT by 35% but indomethacin had no effect on the 5-HT concentration-response curve of the distal coronary artery segments. Indomethacin relaxed precontracted (40 mM potassium) proximal coronary artery segments independently of the presence of the endothelium, suggesting a non-specific relaxing effect of indomethacin in these arteries. It is concluded that rat coronary artery endothelium is unresponsive to 5-HT because it lacks 5-HT1 receptors. The increased 5-HT sensitivity and spontaneous myogenic tone of endothelium-denuded rat coronary arteries is probably due to the elimination of the relaxing stimulus mediated by spontaneously released endothelium-derived relaxing factor.     

Involvement of guanylate cyclase and K(+) channels in relaxation evoked by ferulate nitrate in rat aorta artery

Vasorelaxant properties of N-2-(ferulamidoethyl)-nitrate (ferulate nitrate, FLNT), a newly synthesized nitrate, were compared with those of isosorbide dinitrate, nicorandil, nitroglycerin, and 8-bromoguanosine 3,5-cyclic monophosphate (8-Br-cGMP) in rat aorta pre-contracted by phenylephrine. FLNT produced vasorelaxation in a concentration-dependent manner (0.1 - 100 μM). The degree of relaxation induced by FLNT was similar to that induced by isosorbide dinitrate. In addition, removal of endothelium did not affect the relaxant effect of FLNT. FLNT caused a rightward shift of the cumulative concentration-response curves of phenylephrine and reduced the maximal efficacy of contraction. 1H-[1,2,4]Oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 μM) and K(+)-channel blockers charybdotoxin (CHT, 0.1 μM) and BaCl(2) (1 μM) reduced the relaxant effect of FLNT in the endothelium-denuded arteries, whereas glibenclamide (1 μM) and 4-aminopyridine (1 mM) failed to influence FLNT-induced vasorelaxation. Furthermore, in the presence of ODQ, both CHT (0.1 μM) and BaCl(2) (1 μM) still significantly reduced the relaxation evoked by FLNT. Pretreatment of vessels with hydroxocobalamin, a nitric oxide scavenger, abolished the FLNT effect. These findings demonstrate that FLNT induces relaxation of the rat aorta rings endothelium-independently. Furthermore, we demonstrated that FLNT-induced vasorelaxation is related to its stimulation of soluble guanylate cyclase and activation of K(+) channels.     

The contracting and relaxing responses of human internal mammary artery grafts harvested by two different methods

Perioperative spasm of internal mammary artery is a common experience in coronary artery bypass grafting. Many techniques were described of harvesting the internal mammary artery to prevent vasospasm. We investigated the comparison of the contracting and relaxing responses of human internal mammary artery grafts harvested by two different methods. Patients were divided into two groups depending on the harvesting technique. In the first and second groups arteries were harvested by classical and carbon dioxide insufflation techniques, respectively. In both groups, endothelial function of arteries was assessed by precontracting the rings with phenylephrine (10(-5)M) and dilatating them by cumulative acetylcholine (10(-8) to 10(-5)M) concentrations. Cumulative concentration-response curves for phenylephrine (10(-8) to 10(-4)M), noradrenaline (10(-9) to 10(-4)M), and 5-hydroxytryptamine (10(-9) to 10(-4)M) were obtained in all groups. Endothelial integrity of arteries were histopathologically evaluated. In both groups, acetylcholine caused concentration-dependent relaxations in rings precontracted with phenylephrine (10(-5)M). In arteries harvested by carbon dioxide insufflation technique, acetylcholine caused significantly higher relaxations compared to the rings obtained by classical technique (p<0.05). In all rings of study groups, phenylephrine, noradrenaline and 5-hydroxytryptamine caused concentration-dependent contractions. There was not any significant difference in concentration-dependent responses of these contracting pharmacological agents between the groups. Histopathological evaluation revealed no major arterial damage in both groups. Carbon dioxide insufflation technique does seem not only to protect the integrity of the endothelium and the whole vessel, but also prevent the possible vasospasm of the internal mammary artery segments.     

The enhanced endothelin-1-induced contraction in cultured coronary arteries from mature female pigs is not antagonized by 17beta-estradiol

We have previously reported that organ cultured coronary arteries from market-age pigs (6-9 months of age) exhibit an enhanced contraction to the atherosclerotic-associated peptide, endothelin-1 (ET-1). The objective of this study was to investigate the interaction of 17beta-estradiol with ET-1 in organ cultured coronary arteries from older female pigs (3-4 years old). A cumulative concentration-response relationship (1 x 10(-9) M to 3 x 10(-7) M) was generated to ET-1, and the isometric tension measured in fresh and organ cultured (4 days at 37 degrees C) arterial rings that were each pre-incubated for 50 min in different concentrations (1 x 10(-9) M to 1 x 10(-5) M) of 17beta-estradiol. Compared to freshly used arteries, culturing induced a 2-fold increase in tension development to ET-1 (3 x 10(-7) M). Although 17beta-estradiol previously relaxed pre-constricted (with a 60 mM KCl solution) arteries, it did not affect the constrictive response to ET-1. Also, using an ET-1 ELISA we found that 17beta-estradiol did not effect ET-1 production in intact arteries. Our results indicate that 17beta-estradiol does not attenuate the production and constrictive properties of ET-1 in coronary arteries demonstrating a dedifferentiated cell phenotype.     

N-acetylcysteine does not modify nitroglycerin-induced tolerance in canine vascular rings

The mechanism whereby nitroglycerin initiates relaxation in vascular smooth muscle remains unclear. One hypothesis states that nitroglycerin oxidizes critical sulfhydryl groups in smooth muscle to initiate relaxation, and that repeated exposure to nitroglycerin results in tolerance. In the current study, N-acetylcysteine, a sulfhydryl-reducing agent, was used to explore the sulfhydryl hypothesis by assessing whether or not tolerance to nitroglycerin was reversed by N-acetylcysteine in canine dorsal pedal artery rings. Two nitroglycerin dose-response curves were performed (n = 18)--one before (1st dose-response curve, from 10(-9) to 1.1 X 10(-5) M nitroglycerin) and one after (2nd dose-response curve, from 10(-9) to 5 X 10(-7) M nitroglycerin) incubation with 10(-5) M nitroglycerin for 105 min. At 5 X 10(-7) M nitroglycerin there was 50.7 +/- 10.0% relaxation during the first dose-response curve. During the second dose-response curve, tolerance to nitroglycerin was evident, as demonstrated by a 6.8 +/- 4.8% relaxation (p less than 0.001) at 5 X 10(-7) M nitroglycerin. A 10-min treatment with 10(-3) M N-acetylcysteine (n = 10) during the second nitroglycerin dose-response curve was performed after the 5 X 10(-7) M concentration of nitroglycerin; the second dose-response curve was then completed up to 1.1 X 10(-5) M nitroglycerin. The dose of 10(-3) M N-acetylcysteine was chosen since higher concentrations (i.e., 1.3 X 10(-2) and 1.2 X 10(-1) M N-acetylcysteine) produced 20.3 +/- 8.4 and 43.6 +/- 11.6% relaxation in vascular rings (n = 5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Prevention of nitroglycerin tolerance in vitro by T0156, a selective phosphodiesterase type 5 inhibitor

The efficacy of nitroglycerin as a vasodilator is limited by tolerance, which develops shortly after treatment begins. The present study aims to examine whether T0156, a newly developed potent and selective inhibitor of phosphodiesterase type 5 (PDE5), could attenuate the tolerance to nitroglycerin on rat aortas. Rat aortic rings were suspended in organ bath for the measurement of changes in isometric tension and nitrate tolerance was acutely induced by preceding exposure for 90 min to 30 microM nitroglycerin. Concentration-response curves to nitroglycerin were obtained on aortic rings pre-contracted with phenylephrine. Pre-exposure of rings with or without endothelium to nitroglycerin reduced the relaxations to nitroglycerin. The tissue levels of cyclic GMP were measured by enzyme immunoassay kit. Treatment with T0156 inhibited and prevented the reduced relaxation and cyclic GMP levels in response to nitroglycerin in tolerant rings. In contrast, nitroglycerin-induced tolerance was unaffected by cilostazol (PDE3 inhibitor) and rolipram (PDE4 inhibitor). Finally, incubation of aortic rings with thromboxane prostanoid receptor antagonist, cyclooxygenase inhibitor, or endothelin ET(A) receptor antagonist did not inhibit the development of tolerance. The present results suggest that nitroglycerin tolerance may involve an increased activity of PDE5 but not PDE3 or PDE4 isoforms in vascular smooth muscle cells since T0156 prevents the development of tolerance. Thromboxane A(2), cyclooxygenase (COX)-dependent prostaglandins and endothelin 1 play little role in the acute induction of nitroglycerin tolerance.     

Comparison of the effects of the novel vasodilator FK409 with those of nitroglycerin in isolated coronary artery of the dog.

1. The vasorelaxant effects of FK409, a new nitrovasodilator synthesized from a microbial product, were compared with those of nitroglycerin in isolated coronary artery rings of the dog contracted with U46619 (10(-7) M). 2. FK409 (10(-11)-10(-5) M) and nitroglycerin (10(-9)-10(-4) M) each produced a concentration-dependent relaxation. Comparison of EC50 values showed that FK409 was about 25 times more potent than nitroglycerin. 3. Submaximum concentrations of nitroglycerin (10(-6) M) and FK409 (3 x 10(-8) M) elevated guanosine 3':5'-cyclic monophosphate (cyclic GMP) levels, effects associated with vasorelaxation. Adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels were unaffected. 4. The concentration-relaxation curves for nitroglycerin and FK409 were shifted to the right by methylene blue (3 x 10(-6) - 3 x 10(-5) M), an inhibitor of soluble guanylate cyclase, and to the left by M&B22,948 (3 x 10(-6) - 3 x 10(-5) M), an inhibitor of cyclic GMP phosphodiesterase. 5. After exposure of coronary arteries to the maximally-effective concentration of nitroglycerin (10(-4) M), the mean EC50 value of FK409 did not change significantly, although that of nitroglycerin increased about 60 fold. After exposure to the maximally-effective concentration of FK409 (10(-5) M), the mean EC50 value of FK409 increased about 6 fold and that of nitroglycerin about 11 fold. 6. These results suggest that the vasorelaxant effect of FK409, like that of nitroglycerin, is due to activation of soluble guanylate cyclase and a resultant increase in intracellular cyclic GMP. However, compared with nitroglycerin, there was less self-tolerance to the relaxant effects of FK409 and relatively little cross-tolerance between the two agents.     

Mechanism of relaxations to dendroaspis natriuretic peptide in canine coronary arteries

Experiments were designed to determine mechanisms by which Dendroaspis natriuretic peptide (DNP) causes relaxations in coronary arteries. Rings of canine left circumflex artery with and without endothelium were suspended in organ chambers filled with Krebs-Ringer bicarbonate solution (37 degrees C, bubbled with 95% O2/5% CO2). Concentration-response curves to DNP (10(-10) to 3 x 10(-7) M) were obtained in arteries contracted with prostaglandin (PGF(2alpha), 2 x 10(-6) M), either in the absence or the presence of C-ANP (10(-6) M) to inhibit natriuretic clearance receptors; indomethacin to inhibit cyclooxygenase (INDO, 10(-5) M), N(G)-monomethyl-L-arginine to inhibit production of nitric oxide (L-NMMA; 10(-4) M), HS-142-1 to inhibit particulate guanylate cyclase (10(-5) M); 1H-[1,2,4]oxadiazolo-[4,3-alpha]quinoxalin-1-one to inhibit soluble guanylate cyclase (ODQ; 10(-5) M), or tetraethylammonium to inhibit potassium channels (TEA; 10(-3) or 10(-2) M). Relaxations to DNP were greater in rings with than in those without endothelium. C-ANP significantly attenuated relaxations to DNP only in rings with endothelium. HS-146-1 but not INDO, L-NMMA, ODQ, and TEA significantly reduced relaxations to DNP in rings with and without endothelium contracted with PGF(2alpha). These results suggest that the endothelium augments inhibitory effects of DNP and that natriuretic clearance receptors mediate this component of the response to DNP in canine coronary arteries. In addition, relaxations to DNP in canine arterial smooth muscle involve activation of particulate guanylate cyclase but not hyperpolarization.     

Glyceryl trinitrate (nitroglycerin) and the organic nitrates. Choosing the method of administration

Nitrate usage worldwide is on the increase as the indications for therapy expand. Present indications for nitrate therapy include chronic stable angina pectoris, unstable angina pectoris, complications of acute myocardial infarction, and 'unloading' therapy for acute and chronic congestive heart failure. Nitrates are also being used in the operating suite by anaesthesiologists to control systolic blood pressure during various surgical procedures. New nitrate delivery systems have recently become available which provide considerable dosing flexibility, further increasing the interest in this group of compounds. The dominant action of nitrates is a direct effect on vascular smooth muscle, producing vasodilation of the veins and arteries. These drugs decrease myocardial work by lowering systolic blood pressure, systemic vascular resistance, and reducing intracardiac dimensions. In addition, nitrates have a potent effect on cardiac preload as a result of systemic venodilatation. There is also some evidence that nitrates exert direct effects on the coronary circulation (vasodilatation of coronary arteries and coronary collateral vessels, and direct atherosclerotic stenosis dilatation). These actions may play a role in relieving myocardial ischaemia. Adverse sequelae of nitrate therapy are well known and serious adverse reactions are uncommon. Headache and dizziness are the most frequent side effects. Nitrate tolerance is a definite problem - present evidence indicates that long acting formulations, high doses, or frequent dosing regimens are particularly likely to induce vascular tolerance to nitrates. Consequently, provision of a nitrate-free interval has taken on increasing significance as a strategy to avoid tolerance. Nitrate delivery systems are numerous. Although availability varies from country to country, in most countries there are a wide variety of formulations of glyceryl trinitrate (nitroglycerin) available, including sublingual and oral tablets, oral spray, topical ointment as well as discs or patches for transdermal administration, a transmucosal tablet and an intravenous formulation. Similar formulations of isosorbide dinitrate, except buccal tablets, are available in some countries. Isosorbide 5-mononitrate, a potent metabolite of isosorbide dinitrate, is achieving increasing popularity as an antianginal drug. Optimum nitrate therapy requires a good understanding of the properties of the various formulations, particularly onset and duration of action and propensity to induce tolerance.(ABSTRACT TRUNCATED AT 400 WORDS)     

鸡豆黄素A对狗体外冠状动脉血管的舒张作用及其机制

目的 以血管张力变化为指标,对鸡豆黄素A的心血管保护作用及相关机制进行探讨.方法 制备狗的冠状动脉血管环,固定于恒温浴槽内,待平衡后,加入各种药物观察血管张力的变化.结果 (1)5-羟色胺(5-HT)(10-7～10-5 mol· L-1)使冠状动脉血管环产生明显的剂量依赖性收缩,分别用40 nmol· L-1、1 μmol· L-1鸡豆黄素A温育预处理后,5-HT量效收缩明显被抑制,量效收缩曲线明显右移.(2)鸡豆黄素A可使60 mmol· L-1 KCl预收缩冠状动脉血管环产生明显剂量依赖的舒张作用,缺失内皮细胞或分别加入一氧化氮合酶抑制剂Nω-L-硝基精氨酸1×10-4 mol· L-1,吲哚美辛1×10-5 mol· L-1,甲烯蓝1×10-5mol·L-1温育预处理,鸡豆黄素A对60 mmol· L-1 KCl预收缩冠状动脉血管环产生的量效舒张作用无明显改变(P＞0.05).结论 (1)鸡豆黄素A使血管环对5-HT的敏感性和最大收缩明显降低.(2)鸡豆黄素A对冠状动脉血管的舒张作用与内皮细胞及其释放的一氧化氮无关,与前列腺素类物质亦无关.     

Flunarizine inhibits endothelium-dependent hypoxic facilitation in canine coronary arteries through an action on vascular smooth muscle.

1. Hypoxia augments contractile responses to several vasoactive agents in canine isolated coronary arteries with intact endothelium. Calcium antagonists inhibit the further increases in tension caused by hypoxia. The present experiments were designed to determine whether the calcium-antagonist flunarizine would inhibit hypoxic contractions in isolated blood vessels through an action on the endothelium or on the vascular smooth muscle. 2. Rings of canine coronary arteries, with or without endothelium, were suspended at optimal length for isometric tension recording in organ chambers filled with modified Krebs-Ringer bicarbonate solution. 3. Hypoxia (95% N2 and 5% CO2) augmented contractile responses to prostaglandin F2 alpha (2 x 10(-6) M); removal of the endothelium abolished this hypoxic facilitation. 4. Flunarizine (5 x 10(-5)-5 x 10(-7) M) exerted a long-lasting inhibition of the hypoxic facilitation in a concentration-dependent manner. Flunarizine did not inhibit the response to prostaglandin F2 alpha. 5. To differentiate between the response of smooth muscle and the endothelium, strips of coronary arteries without endothelium were layered with strips with or without endothelium. Hypoxia augmented contractions only in layered preparations with endothelium. Flunarizine prevented the hypoxic contractions in layered preparations in which only the smooth muscle was treated with flunarizine. In contrast, when only the endothelium was treated, no or minimal inhibition of the hypoxic contraction occurred with flunarizine (10(-5) and 5 x 10(-5) M, respectively). 6. These experiments indicate that the calcium antagonist flunarizine inhibits endothelium-dependent hypoxic facilitation in canine coronary arteries primarily through its action on vascular smooth muscle.     

The mode of vasorelaxing action of melatonin in rabbit aorta.

1. The vasorelaxing effect of melatonin on the contractile response to 5-hydroxytryptamine (5-HT) was investigated in rabbit isolated aorta. 2. Melatonin (10(-5)-10(-3) M) caused relaxation of the 5-HT (10 M) response in a concentration-dependent manner. Nifedipine (10(-6) M) did not affect the relaxing action of melatonin. 3. Pretreatment with methylene blue (10(-5) M) or nitroglycerin (3 x 10(-8) M) inhibited or potentiated, respectively, the relaxing action of melatonin. 4. Pretreatment with melatonin (10(-3) M) or M&B 22.948 (10(-3) M) potentiated the relaxing effect of nitroglycerin (10(-9)-10(-5) M) on the contraction induced by PGF2 alpha (4 x 10(-6) M). The effect of a combined treatment with melatonin and M&B 22.948 was not significantly different from that of a single treatment with M&B 22.948. 5. Melatonin (10(-5)-10(-3) M) inhibited the activity of cGMP-phosphodiesterase, in a concentration-dependent manner. 6. These results suggest that the vasorelaxing action of melatonin may be due to an increase in the level of cGMP.     

Endothelium modulation of the effects of nitroglycerin on blood vessels from dogs with pacing-induced heart failure.

1. The relaxant actions of nitroglycerin (previously considered to be an endothelium-independent relaxing agent) and acetylcholine (an endothelium-dependent relaxing agent) were compared on 4 vascular preparations (dorsal pedal artery, saphenous vein, left anterior descending coronary artery and circumflex coronary artery) from dogs with and without pacing-induced congestive heart failure (CHF). 2. Responses of the coronary arteries to acetylcholine were unaltered in endothelium-intact rings from dogs with and without heart failure. Similarly no such changes were observed in the peripheral vessels. The maximum relaxation produced by acetylcholine was always greater in the coronary vessels compared to the peripheral vessels. 3. Before heart failure, the coronary vessels were more sensitive and reactive to nitroglycerin compared to the peripheral vessels. 4. Removal of the endothelium in both the control (dogs without CHF) and experimental (dogs with CHF) rings enhanced the relaxant effects of nitroglycerin, such that the EC50 for nitroglycerin became significantly lower in all denuded rings, with the exception of the saphenous vein and the left anterior descending coronary artery, before the development of CHF. 5. When CHF was maximally developed, vascular sensitivity to nitroglycerin was increased in peripheral vessels with an intact endothelium, but not in the coronary vessels. 6. These findings indicate that relaxation produced by nitroglycerin cannot be considered as entirely endothelium-independent but should be considered endothelium-modulated.     

Vasorelaxant and antiproliferative effects of berberine

The present study was intended to examine the relaxant effects of berberine in rat isolated mesenteric arteries. Berberine produced a rightward shift of the concentration-response curve to phenylephrine and significantly reduced the maximal contractile response to phenylephrine. Berberine (10(-7)-3x10(-5) M) also relaxed the phenylephrine- and 9,11-dideoxy-11alpha, 9alpha-epoxy-methanoprostaglandin F(2alpha)-precontracted arteries with respective IC(50) values of 1.48+/-0.16x10(-6) and 2.23+/-0. 22x10(-6) M. Removal of a functional endothelium significantly attenuated the berberine-induced relaxation (IC(50): 4.73+/-0. 32x10(-6) M) without affecting the maximum relaxant response. Pretreatment with N(G)-nitro-L-arginine methyl ester (L-NAME) or methylene blue reduced the relaxant effect of berberine, and L-arginine (10(-3) M) partially antagonized the effect of L-NAME. In contrast, pretreatment with 10(-6) M glibenclamide or 10(-5) M indomethacin had no effect. Berberine (10(-5) M) reduced over by 50% the transient contraction induced by caffeine or phenylephrine in endothelium-denuded rings bathed in Ca(2+)-free Krebs solution. Pretreatment with putative K(+) channel blockers, such as tetrapentylammonium ions (1-3x10(-6) M), 4-aminopyridine (10(-3) M), or Ba(2+) (3x10(-4) M), significantly attenuated the berberine-induced relaxation in endothelium-denuded arteries. In contrast, tetraethylammonium ions (3x10(-3) M), charybdotoxin (10(-7) M) or glibenclamide (10(-6) M) were without effect. Berberine reduced the high-K(+)-induced sustained contraction and the relaxant response to berberine was greater in rings with endothelium (IC(50): 4.41+/-0.47x10(-6) M) than in those without endothelium (IC(50): 8.73+/-0.74x10(-6) M). However, berberine (10(-6)-10(-4) M) did not affect the high-K(+)-induced increase of intracellular [Ca(2+)] in cultured aortic smooth muscle cells. Berberine did not affect active phorbol ester-induced contraction in Ca(2+)-free Krebs solution. In addition, berberine inhibited proliferation of cultured rat aortic smooth muscle cells with an IC(50) of 2.3+/-0.43x10(-5) M. These findings suggest that berberine could act at both endothelium and the underlying vascular smooth muscle to induce relaxation. Nitric oxide from endothelium may account primarily for the berberine-induced endothelium-dependent relaxation, while activation of tetrapentylammonium-, 4-aminopyridine- and Ba(2+)-sensitive K(+) channels, inhibition of intracellular Ca(2+) release from caffeine-sensitive pools, or a direct relaxant effect, is likely responsible for the berberine-induced endothelium-independent relaxation. Mechanisms related to either Ca(2+) influx or protein kinase C activation may not be involved. Both vasorelaxant and antiproliferative effects may contribute to a long-term benefit of berberine in the vascular system.