Isolated Rat Carotid Artery Research Articles

Methylglyoxal impairs ATP- and UTP-induced relaxation in the rat carotid arteries

Although methylglyoxal (MGO), a highly reactive dicarbonyl compound, influences the functioning of the vasculature, modulating its effects on vascular reactivity to various substances remains unclear, especially purinoceptor ligands. Therefore, we sought to investigate the direct effects of MGO on relaxation induced by adenosine 5′-triphosphate (ATP) and uridine 5′-triphosphate (UTP) in isolated rat carotid arteries. When carotid arteries were exposed to MGO (420 μM for 1 h), relaxation induced by acetylcholine or sodium nitroprusside was not affected by MGO. However, ATP- and UTP-induced relaxation was impaired by MGO compared with the control. In both ATP- and UTP-induced relaxation, endothelial denudation, incubation with the nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine or the selective P2Y purinoceptor 2 (P2Y2) receptor antagonist AR-C118925XX reduced relaxation in both the control and MGO groups, while the differences between the control and MGO groups were eliminated. The cyclooxygenase (COX) inhibitor indomethacin inhibited the differences in ATP/UTP-mediated relaxations between the control and MGO groups. Moreover, N-acetyl-L-cysteine (NAC), an antioxidant, could augment carotid arterial relaxation induced by ATP/UTP in the presence of MGO. MGO increased arachidonic acid-induced contraction, which was suppressed by NAC. Following both ATP/UTP stimulation, MGO increased the release of prostanoids. These results suggest that MGO impaired ATP- and UTP-induced relaxation in carotid arteries, which was caused by suppressed P2Y2 receptor-mediated signaling and reductions in endothelial NO. Moreover, MGO partially contributed to COX-derived vasoconstrictor prostanoids through increased oxidative stress.

Requirement for sphingosine kinase 1 in mediating phase 1 of the hypotensive response to anandamide in the anaesthetised mouse

In the isolated rat carotid artery, the endocannabinoid anandamide induces endothelium-dependent relaxation via activation of the enzyme sphingosine kinase (SK). This generates sphingosine-1-phosphate (S1P) which can be released from the cell and activates S1P receptors on the endothelium. In anaesthetised mice, anandamide has a well-characterised triphasic effect on blood pressure but the contribution of SK and S1P receptors in mediating changes in blood pressure has never been studied. Therefore, we assessed this in the current study.The peak hypotensive response to 1 and 10 mg/kg anandamide was measured in control C57BL/6 mice and in mice pretreated with selective inhibitors of SK1 (BML-258, also known as SK1-I) or SK2 ((R)-FTY720 methylether (ROMe), a dual SK1/2 inhibitor (SKi) or an S1P1 receptor antagonist (W146). Vasodilator responses to S1P were also studied in isolated mouse aortic rings.The hypotensive response to anandamide was significantly attenuated by BML-258 but not by ROMe. Antagonising S1P1 receptors with W146 completely blocked the fall in systolic but not diastolic blood pressure in response to anandamide. S1P induced vasodilation in denuded aortic rings was blocked by W146 but caused no vasodilation in endothelium-intact rings.This study provides evidence that the SK1/S1P regulatory-axis is necessary for the rapid hypotension induced by anandamide. Generation of S1P in response to anandamide likely activates S1P1 to reduce total peripheral resistance and lower mean arterial pressure. These findings have important implications in our understanding of the hypotensive and cardiovascular actions of cannabinoids.

Functional Effects of Alagebrium (ALT-711)-Isolated Rat Carotid Artery.

In our study, the effects of glycosylated protein cross-link breaker, alagebrium was investigated on isolated rat carotid artery using myography. Alagebrium showed vasodilator effect on carotid artery rings; particularly, this effect was significantly increased in endothelium-intact rings. To clarify the vasodilator mechanism of alagebrium, different antagonists such as N(G)-Nitro-L-arginine methyl ester (L-NAME), glibenclamide, indomethacin, metoprolol, propranolol, tetraethylammonium, and calcium channel activator BAYK-8644 were used to reverse this effect. Relaxation% responses to alagebrium were more significantly increased in intact endothelium than in denuded arteries. Blocking vasodilation related to channels (K-ATP, PGI2, BKca) and receptors (ß1, ß2) did not reverse the relaxation response to alagebrium. Vasodilator response to alagebrium was only slightly decreased after L-NAME incubation and significantly decreased after BAYK-8644 incubation. Results of present study suggest that the mechanism of alagebrium-induced vasodilator effect may include the blockage of L-type calcium channels and partially of the nitric oxide synthase enzyme.

Effects of the Novel High-affinity 5-HT(1B/1D)-receptor Ligand Frovatriptan on the Rat Carotid Artery

In blood vessels 5-HT stimulates sympathetic nerves, the endothelium and vascular smooth muscle cells. Triptans are specific anti-migraine drugs and they activate the serotoninergic 5HT1b/d receptors causing vasoconstriction of the cerebral vessels. To evaluate the effect of frovatriptan on isolated rat carotid artery. Contractile activity of the preparations was registered isometrically. Krebs solution (pH = 7.4) was used for washing smooth muscle (SM) preparations aerated with 95% O2 and 5% CO2 at 37°C. The 60-minute adaptation of tone level of preparations was taken as a starting tone and the changes such as contraction or relaxation were calculated using it. Frovatriptan (1×10-6 mol/l - 1×10-5 mol/l) induced a contraction, but at higher concentrations it caused relaxation of the carotid artery. The L-norepinephrine contractile reaction was enhanced in the presence of frovatriptan. In the presence of 5-HT2 receptor antagonist, methysergide, frovatriptan increased the relaxation. In the presence of the specific α-1 receptor antagonist, prazosin, the frovatriptan-induced relaxation decreased. The observed contractile effect of frovatriptan is probably associated with the main effect of the drug - activation of the serotoninergic 5HT1B /1D receptors causing vasoconstriction of the cerebral vessels and their anti-migraine effect. At higher concentrations, frovatriptan, most likely via some non-specific mechanism, could activate the following intracellular chain reaction: stimulation of α1D could activate eNOS which may increase in the concentration of NO which results in the final effect of relaxation.

Pericardial fluid of cardiac patients elicits arterial constriction: role of endothelin-1.

Recently, several vasoactive molecules have been found in pericardial fluid (PF). Thus, we hypothesized that in coronary artery disease due to ischemia or ischemia-reperfusion, the level of vasoconstrictors, mainly endothelin-1 (ET-1), increases in PF, which can increase the vasomotor tone of arteries. Experiments were performed using an isometric myograph. Vasomotor effects of PF from patients undergoing coronary artery bypass graft (PFCABG, n = 14) or valve replacement (PFVR, n = 7) surgery were examined in isolated rat carotid arteries (N = 14; n = 26). Vasomotor responses to KCl (40 or 60 mmol/L) were also tested. The selective endothelin A receptor antagonist BQ123 (10(-6) mol/L) was used to elucidate the role of ET-1. Both the first and the second additions of KCl elicited increases in the isometric force of the isolated arteries (KCl1, 6.1 ± 0.2 mN; KCl2, 6.5 ± 0.9 mN). PFCABG and PFVR elicited substantial increases in the isometric force of arteries (PFCABG, 3.1 ± 0.7 mN; PFVR, 3.0 ± 0.9 mN; p > 0.05). The presence of the selective endothelin A receptor blocker significantly reduced arterial contractions to PFCABG (before BQ123, 2.6 ± 0.5 mN vs. after BQ123, 0.8 ± 0.1 mN; p < 0.05). This study is the first to demonstrate that PFs of patients elicit substantial arterial constrictions, which is mediated primarily by ET-1. Interfering with the vasoconstrictor action of PF could be a potential therapeutic target to improve coronary blood flow in cardiac patients.

Effect of age on the contractile response of the rat carotid artery in the presence of sympathetic drugs and L-NAME

For evaluating the age-related change in noradrenaline (NA)-induced contraction of isolated rat carotid artery (CA), the effect of α and β adrenoreceptor (AR) blockers and the role of nitric oxide (NO) were investigated. Concentration-response curves to NA (10-10-10-4 M) and α1 agonist phenylephrine (PE; 10-10-10-5 M) were constructed in isolated CA rings from young and middle-aged rats. The effects of nitric oxide synthase (NOS) inhibitor (L-NAME; 100 μM), α1-AR antagonist (prazosin; 0.1 μM), α2-AR antagonist (yohimbine; 0.1 μM) and β-AR antagonist (propranolol; 1 μM) on NA-induced contraction of isolated CA rings were examined. In CA rings preconstricted with NA, the responses to α2-AR agonist (clonidine; 10-7-10-5 M), β-AR agonist (isoprenaline; 10-8-10-5 M),), sodium nitroprusside (SNP; 10-9-10-5 M) were assessed. The maximum contractile response of CA to NA and to PE was higher in younger than in middle-aged rats. Prazosin reduced the contractile response to NA in both groups, while propranolol, yohimbine and L-NAME did not affect NA-induced contraction in either of them. Clonidine, isoprenaline and SNP produced a dose-dependent vasorelaxation of CA rings, isoprenaline-induced vasodilatation was lower in middle-aged rats, while there was no difference in clonidine or SNP-induced relaxant effect between the two groups. NA-induced contraction of isolated rat CA rings is decreased in old rats, this is related to α1-AR. β-AR mediated dilatation was compromised in middle-aged rats (endothelium-dependent). α2-AR and SNP-mediated dilator effect seems to be unchanged.

In isolated vessels H2S is a less effective scavenger of exogenous superoxide than SOD

Recent studies suggest that hydrogen sulfide (H2S) exhibits potent antioxidant capacity and improves vascular and tissue functions in oxidative stress conditions. Thus we aimed to compare the direct antioxidant efficacy of H2S and superoxide dismutase (SOD).Isometric force of isolated rat carotid arteries and gracilis veins was measured with a wire myograph. The vasomotor effect of the superoxide‐generator pyrogallol (10−5M) was obtained in control conditions, and then in the presence of SOD (120 U/ml) or H2S (10−5M or 10−4M), respectively. Spectrophotometric measurements were performed to detect the effect of SOD and H2S on the auto‐oxidation of pyrogallol to superoxidePyrogallol increased the isometric force of carotid arteries (9.7±0.8mN), which was abolished by SOD (5.3±0.8mN), but was not affected by 10−5M H2S (9.1±0.5mN), whereas 10−4M H2S significantly reduced it (8.1±0.7mN). Pyrogallol significantly increased the isometric force of gracilis veins (1.3±0.2mN), which was abolished by SOD (0.9±0.2mN), whereas 10−5M (1.3±0.2mN) or 10−4M H2S (1.2±0.2mN) did not significantly affect it. Pyrogallol‐induced superoxide production was measured by a spectrophotometer (A420=0.19±0.0). SOD reduced absorbance (A420=0.02±0.0), but 10−5M H2S did not (A420=0.18±0.0), whereas 10−4M H2S slightly, but significantly reduced it (A420=0.15±0.0).This in vitro vascular bioassay study suggests that the superoxide scavenging efficacy of H2S is substantially less than that of SOD, suggesting that the beneficial effects of H2S on vascular function in conditions of oxidative stress could be due primarily to secondary mechanisms activated by H2S.

Bioassay-comparison of the antioxidant efficacy of hydrogen sulfide and superoxide dismutase in isolated arteries and veins

Recent studies suggest that hydrogen sulfide (H2S) exhibits potent antioxidant capacity and improves vascular and tissue functions. Thus we aimed to compare the antioxidant efficacy of H2S to that of superoxide dismutase (SOD).Isometric force of isolated rat carotid arteries and gracilis veins was measured with a myograph. The vasomotor effect of the superoxide-generator pyrogallol (10-5M) was obtained in control conditions, and then in the presence of SOD (120 U/ml) or H2S (10-5M or 10-4M), respectively. Spectrophotometric measurements were performed to detect the effect of SOD and H2S on the auto-oxidation of pyrogallol.Pyrogallol increased the isometric force of carotid arteries (9.7 ± 0.8 mN), which was abolished by SOD (5.3 ± 0.8 mN), was not affected by 10-5M H2S (9.1 ± 0.5 mN), whereas 10-4M H2S slightly, but significantly reduced it (8.1 ± 0.7 mN). Pyrogallol significantly increased the isometric force of gracilis veins (1.3 ± 0.2 mN), which was abolished by SOD (0.9 ± 0.2 mN), whereas 10-5M (1.3 ± 0.2 mN), or 10-4M H2S (1.2 ± 0.2 mN) did not affect it. Pyrogallol-induced superoxide production was measured by a spectrophotometer (A420 = 0.19 ± 0.0). SOD reduced absorbance (A420 = 0.02 ± 0.0), whereas 10-5M H2S did not (A420 = 0.18 ± 0.0) and 10-4M H2S slightly reduced it (A420 = 0.15 ± 0.0).These data suggest that H2S is a less effective vascular antioxidant than SOD. We propose that the previously described beneficial effects of H2S are unlikely to be related to its direct effect on superoxide.

Leptin Augments Cerebral Hemodynamic Reserve after Three-Vessel Occlusion: Distinct Effects on Cerebrovascular Tone and Proliferation in a Nonlethal Model of Hypoperfused Rat Brain

The adipocytokine leptin has distinct functions regulating vascular tone, inflammation, and collateral artery growth. Arteriogenesis is an inflammatory process and provides a mechanism to overcome the effects of vascular obstruction. We, therefore, tested the effects of leptin in hypoperfused rat brain (three-vessel occlusion). Systemic leptin administration for 1 week after occlusion surgery increased cerebral hemodynamic reserve similar to granulocyte-macrophage colony-stimulating factor (GM-CSF), as indicated by improved CO(2) reactivity (vehicle 0.53%±0.26% versus leptin 1.05%±0.6% per mm Hg arterial pCO(2), P<0.05). Infusion of microspheres under maximal vasodilation failed to show a positive effect of leptin on cerebral perfusion (vehicle 64.9%±4.5% versus leptin 66.3%±7.0%, occluded/nonoccluded hemisphere). Acute treatment with GM-CSF led to a significant increased CO(2) reactivity and cerebral perfusion (79.2%±8.1% versus 64.9%±4.5%, P<0.05). Vasoconstrictive response of isolated rat carotid artery rings, after phenylephrine was attenuated at 24 hours following preincubation with leptin, was unaffected by removal of endothelium but abrogated by coculture with N-(omega)-nitro-L-arginine methylester, pointing toward an inducible nitric oxide synthase-mediated mechanism. In chronic cerebral hypoperfusion, acute leptin treatment restored the hemodynamic reserve of the cerebral vasculature through its effects on vascular tone, while leaving vascular outward remodeling unaffected. Our results, for the first time, reveal a protective role of leptin on vascular function in hemodynamically compromised brain tissue.

Inhibition of TRPC1/TRPC3 by PKG contributes to NO-mediated vasorelaxation

Nitric oxide (NO) inhibits transient receptor potential channel 3 (TRPC3) channels via a PKG-dependent mechanism. We sought to determine 1) whether NO inhibition of TRPC3 occurs in freshly isolated smooth muscle cells (SMC); and 2) whether NO inhibition of TRPC3 channels contributes to NO-mediated vasorelaxation. We tested these hypotheses in freshly isolated rat carotid artery (CA) SMC using patch clamp and in intact CA by vessel myograph. We demonstrated TRPC3 expression in whole CA (mRNA and protein) that was localized to the smooth muscle layers. TRPC1 protein was also expressed and coimmunoprecipitated with TRPC3. Whole cell patch clamp demonstrated nonselective cation channel currents that were activated by UTP (60 microM) and completely inhibited by a TRPC channel inhibitor, La(3+) (100 microM). The UTP-stimulated current (I(UTP)) was also inhibited by intracellular application of anti-TRPC3 or anti-TRPC1 antibody, but not by anti-TRPC6 or anti-TRPC4 control antibodies. We next evaluated the NO signaling pathway on I(UTP). Exogenous NO [(Z)-1-{N-methyl-N-[6(N-methylammoniohexyl)amino]}diazen-1-ium-1,2-diolate (MAHMA NONOate)] or a cell-permeable cGMP analog (8-bromo-cGMP) significantly inhibited I(UTP). Preapplication of a PKG inhibitor (KT5823) reversed the inhibition of MAHMA NONOate or 8-bromo-cGMP, demonstrating the critical role of PKG in NO inhibition of TRPC1/TRPC3. Intact CA segments were contracted with UTP (100 microM) in the presence or absence of La(3+) (100 microM) and then evaluated for relaxation to an NO donor, sodium nitroprusside (1 nM to 1 microM). Relaxation to sodium nitroprusside was significantly reduced in the La(3+) treatment group. We conclude that freshly isolated SMC express TRPC1/TRPC3 channels and that these channels are inhibited by NO/cGMP/PKG. Furthermore, NO contributes to vasorelaxation by inhibition of La(3+)-sensitive channels consistent with TRPC1/TRPC3.

Adenosine enhances the relaxing influence of retinal tissue

Retinal tissue from different species continuously releases an as yet unidentified retinal relaxing factor (RRF) lowering tone of isolated arteries. The potential influence of adenosine on this relaxing influence was investigated using isometric tension recording of different isolated arteries. The presence of bovine retinal tissue or rat retinal tissue enhanced the vasorelaxing effect of adenosine on isolated bovine retinal artery. In isolated rat carotid artery adenosine elicited no relaxation. However, a small relaxation is observed in the presence of rat retinal tissue, but not in the presence of porcine retina. The fact that adenosine potentiates the effect of rat retinal tissue but not that of a similar piece of porcine retinal tissue indicates species differences. Neither a NO-synthase inhibitor (nitro- l-arginine, 0.1 mM), a cyclooxygenase inhibitor (indomethacin, 10 μM) or an epoxygenase inhibitor (miconazole, 10 μM) influenced the enhanced vasodilating effect of adenosine on bovine retinal arteries in the presence of bovine retinal tissue. On the other hand, when the retinal arteries were contracted with 120 mM K +, adenosine no longer induced relaxation of the preparation with bovine retinal tissue. This is in line with the concept that adenosine enhances the influence of RRF. Also, the fact that rat carotid artery is less sensitive to RRF than bovine retinal artery – corresponding with a less enhanced adenosine response in rat carotid artery – is in line with the potential involvement of the RRF in the enhanced adenosine response. However, experiments using a bioassay setup for RRF gave no evidence for an increased RRF-release from the retina, nor for an increased RRF-sensitivity of the retinal artery in the presence of adenosine. In conclusion, our findings indicate that adenosine potentiates the relaxing influence of bovine and rat retinal tissue. This effect is species dependent as it is not seen with porcine retinal tissue. Neither NO, cyclooxygenase metabolites or epoxyeicosatrienoic acids seem to be involved in this enhanced vasorelaxing response. The involvement of the RRF cannot be excluded.

Inhibition of sphingosine kinase induces vasoconstriction in isolated carotid arteries obtained from spontaneous hypertensive but not normotensive rats

Ceramide, sphingosine and sphingosine‐1‐phosphate (S1P) are signaling molecules generated from membrane phospholipids by a concerted action of several enzymes. These sphingomyelin metabolites (SM) have growth regulating properties; while S1P is implicated in growth stimulation, ceramide and sphingosine (the precursors of S1P) induce apoptosis. In addition, SM have vasoactive properties. For instance S1P, synthesized by sphingosine kinase, can increase NO production in endothelial cells and thereby induce vasodilation. Hypertension is a disease state characterized by endothelial dysfunction (decreased NO bioavailability) and vascular growth (remodeling). Therefore we investigated whether hypertension alters the vasoactive properties of locally produced SM. Inhibition of sphingosine kinase by means of dimethylsphingosine (DMS, 10 μM) induced a biphasic and transient contraction in isolated rat carotid arteries from spontaneous hypertensive but not normotensive animals. This contraction was endothelium and eicosanoid‐dependent since removal of the endothelium and inhibition of cyclooxygenase completely prevented DMS‐induced constriction. Alterations in sphingolipid metabolism may thus contribute to a disturbed regulation of vascular tone during hypertension.This research was performed within the framework of project T2‐108 of the Dutch Top Institute Pharma.

Adventitial Hydrogen Peroxide (H2O2) Impairs Rat Carotid Artery Relaxation via Medial p38 MAPK

The vascular adventitia is a major source of H2O2. H2O2 activates smooth muscle p38 mitogen activated protein kinase (p38 MAPK). We postulated that adventitia‐derived H2O2 impairs relaxation via medial kinase activation. Isolated rat carotid arteries were bathed perivascularly (in situ) over 90 min in vehicle, angiotensin II (AngII, 500 nM), AngII + H2O2‐scavenger catalase or AngII + p38 MAPK inhibitor, then placed in myographs and preconstricted. Both acetylcholine (ACh)‐induced endothelium‐dependent and sodium nitroprusside (SNP)‐induced endothelium‐independent relaxations were impaired by AngII (p &lt; 0.05). Catalase partially reversed the AngII effect on ACh‐induced relaxation and reversed impairment of SNP‐induced relaxations (p &lt; 0.05). The p38 MAPK inhibitor SB203580 partially ameliorated the impairment of SNP‐induced relaxations and reversed the AngII effect on ACh‐induced relaxation (p &lt; 0.05). In separate pretreated vessels, the media was dissected, homogenized and Western blots run for phosphorylated vs. total p38 MAPK. AngII activated medial p38 MAPK, but catalase inhibited this. These data suggest AngII elicits release of adventitial H2O2, impairing both endothelium‐dependent and –independent relaxation via the p38 MAPK pathway in the media. The data provide evidence of a significant paracrine signaling role of vascular adventitial H2O2 on vascular function.

Functional characterization of the mechanisms underlying bradykinin-induced relaxation in the isolated rat carotid artery

This work aimed to functionally characterize the mechanisms underlying the relaxation induced by bradykinin (BK) in the rat carotid artery. Vascular reactivity experiments, using standard muscle bath procedures, showed that BK (0.1 nmol/L–3 μmol/L) induced relaxation of phenylephrine-pre-contracted rings in a concentration-dependent manner. Endothelial removal strongly attenuated BK-induced relaxation. HOE-140, the selective antagonist of bradykinin B 2 receptors concentration-dependently reduced the relaxation induced by BK. Pre-incubation of endothelium-intact rings with l-NAME (100 µmol/L), a non-selective nitric oxide synthase (NOS) inhibitor, l-NAME (100 µmol/L), a selective inhibitor of the eNOS or 7-nitroindazole (100 µmol/L), the selective inhibitor of nNOS, reduced BK-induced relaxation. Conversely, 1400 W (10 nmol/L), a selective inhibitor of iNOS, did not alter the relaxation induced by BK. Surprisingly, indomethacin (10 µmol/L) a non-selective inhibitor of cyclooxygenase (COX) increased BK-induced relaxation in endothelium-intact but not denuded rings. Neither SQ29548 (3 µmol/L), a competitive antagonist of PGH 2/TXA 2 receptors nor AH6809 (10 µmol/L), an antagonist of PGF 2α receptors significantly altered the relaxation induced by BK in endothelium-intact rings. The combination of SQ29548 and AH6809 increased BK-induced relaxation. The present study shows that the vasorelaxant action displayed by BK in the rat carotid is mediated by endothelial B 2 receptors and the activation of the NO pathway. The major finding of this work is that it demonstrated functionally that endothelial-derived vasoconstrictor prostanoids (probably PGH 2, TXA 2 and PGF 2α) counteract the vasorelaxant action displayed by BK.

Sphingosine Kinase–Dependent Activation of Endothelial Nitric Oxide Synthase by Angiotensin II

In addition to their role in programmed cell death, cell survival, and cell growth, sphingolipid metabolites such as ceramide, sphingosine, and sphingosine-1-phosphate have vasoactive properties. Besides their occurrence in blood, they can also be formed locally in the vascular wall itself in response to external stimuli. This study was performed to investigate whether vasoactive compounds modulate sphingolipid metabolism in the vascular wall and how this might contribute to the vascular responses. In isolated rat carotid arteries, the contractile responses to angiotensin II are enhanced by the sphingosine kinase inhibitor dimethylsphingosine. Endothelium removal or NO synthase inhibition by N(omega)-nitro-L-arginine results in a similar enhancement. Angiotensin II concentration-dependently induces NO production in an endothelial cell line, which can be diminished by dimethylsphingosine. Using immunoblotting and intracellular calcium measurements, we demonstrate that this sphingosine kinase-dependent endothelial NO synthase activation is mediated via both phosphatidylinositol 3-kinase/Akt and calcium-dependent pathways. Angiotensin II induces a sphingosine kinase-dependent activation of endothelial NO synthase, which partially counteracts the contractile responses in isolated artery preparations. This pathway may be of importance under pathological circumstances with reduced NO bioavailability. Moreover, a disturbed sphingolipid metabolism in the vascular wall may lead to reduced NO bioavailability and endothelial dysfunction.

Vascular effects of a soy leaves (Glycine max) extract and kaempferol glycosides in isolated rat carotid arteries.

We have recently purified genistin, and six kaempferol glycosides from a soy leaves ( Glycine max L. Merr.) butanol extract. Here we report the vascular effects of the extract and purified genistin and kaempferol glycosides on contractions induced by different constricting agonists in isolated rat carotid arteries. The butanol extract relaxed artery rings preconstricted by 9,11-dideoxy-11alpha,9alpha-epoxy-methanoprostaglandin F 2 alpha (U46619) or [5 Z,9alpha,11alpha,13 E,15 S]-9,11,15-trihydroxyprosta-5,13-dienoic acid (PGF 2 alpha ) in a dose-dependent manner and this effect was independent of the presence of endothelium. The extract also inhibited the concentration-dependent contraction to U46619 with a slight reduction of the maximal response. The extract produced partial relaxation of both phenylephrine-preconstricted endothelium-intact and -denuded rings. In contrast, the extract had no effect on the contractile response to 50 mM extracellular K (+). None of the six kaempferol glycosides affected vessel tension induced by U46619. A mixture of kaempferol glycosides prepared according to their relative composition in the extract had no effect either. However, kaempferol relaxed U46619- and high K (+)-contracted rings to the same extent. Endothelium played no role in kaempferol-induced relaxation. Genistein induced concentration-dependent relaxation and this effect was attenuated in the endothelium-denuded rings. Genistin caused a smaller relaxant effect. The present results indicate that a butanol extract from soy leaves causes endothelium-independent relaxation in rat carotid artery rings. Kaempferol glycosides, accounting for approximately 48 % of the extract in weight, are not the ingredients responsible for the extract-induced relaxation. Genistein and genistin also caused relaxation, however, the dose range is beyond that of the extract causing relaxation.

Cibenzoline has an inhibitory effect on vasorelaxation mediated by adenosine triphosphate-sensitive K(+) channels in the rat carotid artery.

Studies in cardiac myocytes have shown that cibenzoline reduces adenosine triphosphate (ATP)-sensitive K(+) currents, suggesting that this class Ia antiarrhythmic drug may modify the activity of ATP-sensitive K(+) channels in these preparations. The effects of class Ia antiarrhythmic drugs on vasodilation mediated by ion channels have not been studied. Therefore, we designed this study to examine whether cibenzoline may produce changes in vasorelaxation in response to a selective ATP-sensitive K(+) channel opener, levcromakalim, in the isolated rat carotid artery. Rings of rat carotid arteries without endothelium were suspended for isometric force recording. Concentration-response curves were obtained in a cumulative fashion. During submaximal contraction to phenylephrine (3 x 10(-7) M), vasorelaxation in response to levcromakalim (10(-8) to 10(-5) M) or 1-hydroxy-2-oxo-3-(N-methyl-3-aminopropyl)-3-methyl-1-triazene (NOC-7; 10(-10) to 10(-5) M) was obtained. During contraction to phenylephrine, levcromakalim induced concentration-dependent vasorelaxation. A selective ATP-sensitive K(+) channel antagonist, glibenclamide (5 x 10(-6) M), completely abolished vasorelaxation in response to levcromakalim, whereas a selective Ca(2+)-dependent K(+) channel antagonist, iberiotoxin (5 x 10(-8) M), did not affect the relaxation. Cibenzoline (10(-6) to 10(-5) M) significantly reduced vasorelaxation to levcromakalim in a concentration-dependent fashion. In contrast, cibenzoline (10(-5) M) did not alter vasorelaxation to a nitric oxide donor, NOC-7. These results suggest that from the clinically relevant concentrations, a novel class Ia antiarrhythmic drug, cibenzoline, impairs carotid vasodilation mediated by ATP-sensitive K(+) channels. In isolated rat carotid artery, cibenzoline (10(-6) to 10(-5) M) reduced vasorelaxation to levcromakalim in a concentration-dependent fashion. These results suggest that from the clinically relevant concentrations, a novel class Ia antiarrhythmic drug, cibenzoline, impairs carotid vasodilation mediated by adenosine triphosphate-sensitive K(+) channels.

Cibenzoline Has an Inhibitory Effect on Vasorelaxation Mediated by Adenosine Triphosphate-Sensitive K+ Channels in the Rat Carotid Artery

Studies in cardiac myocytes have shown that cibenzoline reduces adenosine triphosphate (ATP)-sensitive K+ currents, suggesting that this class Ia antiarrhythmic drug may modify the activity of ATP-sensitive K+ channels in these preparations. The effects of class Ia antiarrhythmic drugs on vasodilation mediated by ion channels have not been studied. Therefore, we designed this study to examine whether cibenzoline may produce changes in vasorelaxation in response to a selective ATP-sensitive K+ channel opener, levcromakalim, in the isolated rat carotid artery. Rings of rat carotid arteries without endothelium were suspended for isometric force recording. Concentration-response curves were obtained in a cumulative fashion. During submaximal contraction to phenylephrine (3 × 10−7 M), vasorelaxation in response to levcromakalim (10−8 to 10−5 M) or 1-hydroxy-2-oxo-3-(N-methyl-3-aminopropyl)-3-methyl-1-triazene (NOC-7; 10−10 to 10−5 M) was obtained. During contraction to phenylephrine, levcromakalim induced concentration-dependent vasorelaxation. A selective ATP-sensitive K+ channel antagonist, glibenclamide (5 × 10−6 M), completely abolished vasorelaxation in response to levcromakalim, whereas a selective Ca2+-dependent K+ channel antagonist, iberiotoxin (5 × 10−8 M), did not affect the relaxation. Cibenzoline (10−6 to 10−5 M) significantly reduced vasorelaxation to levcromakalim in a concentration-dependent fashion. In contrast, cibenzoline (10−5 M) did not alter vasorelaxation to a nitric oxide donor, NOC-7. These results suggest that from the clinically relevant concentrations, a novel class Ia antiarrhythmic drug, cibenzoline, impairs carotid vasodilation mediated by ATP-sensitive K+ channels.

COCAINE-INDUCED HYPERTENSION: ROLE OF THE PERIPHERAL SYMPATHETIC SYSTEM

Cocaine causes hypertension at least in part by stimulating the sympathetic nervous system, but it is not clear if this effect is centrally or peripherally mediated. To address this issue we studied the vasoconstrictive effect of cocaine in vivo and in isolated artery segments. In vivo cocaine increased mean arterial blood pressure (MAP) by 40 mmHg within 1 min of administration. Pretreatment with prazosin blocked this response by 62%. With clonidine the pre-cocaine MAP was lower and the hypertensive effect of cocaine was blocked by 50%, indicating an important role for central α-adrenergic mechanisms. In isolated rat carotid arteries cocaine-induced vasoconstriction was completely blocked by prazosin, phentolamine, and 6-hydroxydopamine, indicating a clear role for a peripheral effect. However, the relative contribution of the central α-adrenergic mechanism to the total vasoconstrictive response of cocaine was not clarified. 1999 Academic Press@p$hr