Coronary Rings Research Articles

Standardized Rat Coronary Ring Preparation and Real-Time Recording of Dynamic Tension Changes Along Vessel Diameter

As a key event of cardiovascular system diseases, coronary artery disease (CAD) has been widely regarded as the main culprit of atherosclerosis, myocardial infarction, and angina pectoris, which seriously threaten the life and health of people all over the world. However, how to record the dynamic biomechanical characteristics of isolated blood vessels has long puzzled people. Meanwhile, precise positioning and isolation of coronary arteries to measure in vitro dynamic vascular tension changes have become a trend in CAD drug development. The present protocol describes the macroscopic identification and microscopic separation of rat coronary arteries. The contraction and dilation function of the coronary artery ring along the vessel diameter was monitored using the established multi myograph system. The standardized and programmed protocols of coronary ring tension measurement, from sampling to data acquisition, tremendously improve the repeatability of the experimental data, which ensures the authenticity of vascular tension records after physiological, pathological, and drug intervention.

Endothelium-independent vasorelaxant effect of Asphodelus tenuifolius Cav. via inhibition of myosin light chain kinase activity in the porcine coronary artery

Ethnopharmacological relevanceAsphodelus tenuifolius Cav. (Asphodelaceae), a wild, terrestrial, annual stemless herb, is widely used in traditional medicine for the treatment of hypertension, diabetes, atherosclerosis and circulatory problems. A previous research study from our laboratory revealed that A. tenuifolius has beneficial effects in reducing blood pressure and improves aortic endothelial dysfunction in chronically glucose fed rats. Despite the fact that A. tenuifolius reduces blood pressure and improves endothelial function in vivo, there are no detailed studies about its possible mechanism of action. Aim of the studyThis study was designed to provide pharmacological basis and mechanism of action for the traditional use of A. tenuifolius in hypertension and circulatory problems. We explored the vasorelaxant effect of A. tenuifolius and its underlying vasorelaxation mechanism in porcine coronary artery rings. Materials and methodsAqueous methanolic crude extract of A. tenuifolius was prepared by maceration process and then activity guided fractionation was carried out by using different polarity based solvents. Phytochemical studies were carried out using LC-DAD-MS. Segments of porcine distal coronary artery were set up in a wire myograph for isometric force measurements. Extract/fractions of A. tenuifolius seeds were tested for vasodilator activity by measurement of changes in tone after pre-contraction with the thromboxane mimetic U46619 in the presence or absence of inhibitors of intracellular signaling cascades. ResultsCrude extract/fractions of A. tenuifolius produced dose dependent endothelium independent vasorelaxant response in coronary rings, whereas, the butanol fraction of A. tenuifolius (BS-AT) produced the largest relaxation response with 100% relaxation at 1 mg/ml, therefore the mechanism of relaxation of this fraction was determined. The relaxation to BS-AT was unaffected by removal of the endothelium, pre-contraction with KCl, or the presence of the non-selective potassium channel blocker tetraethylammonium, indicating that the relaxation was endothelium-independent, and does not involve activation of potassium channels. BS-AT (1 mg/ml) inhibited the contractile response to calcium,the L-type calcium channel activator BAY K8664,and ionomycin, indicating that it inhibits calcium-induced contractions. The relaxation response to BS-AT was attenuated in the absence of extracellular calcium. However, relaxations to BS-AT were also reduced after deletion of calcium from intracellular stores with cyclopiazonic acid. Incubation with 1 mg/ml BS-AT also inhibited phosphorylation of myosin light chains in homogenates of coronary artery. ConclusionThe butanol extract of Asphodelus tenuifolius produces a large endothelium-independent relaxation of the porcine coronary artery through inhibition of calcium-induced contractions. The effect appears to be downstream of calcium influx, possibly through inhibition of myosin light chain kinase. This study supports previous studies demonstrating that A. tenuifolius reduces blood pressure. Future studies will aim to determine the active compounds underlying this response.

Terminalia avicennioides causes redox-sensitive endothelium-dependent relaxation involving nitric oxide, prostacyclin, and endothelium-derived hyperpolarizing in porcine coronary artery and different conductance and resistance vessels from rats

Background: Our work is part of the enhancement of plants of Senegalese pharmacopeia with cardiovascular aim, among which Terminalia avicennioides. Aim and Objective: The aim of this study was to determine whether a hydroethanolic bark extract of T. avicennioides (TAE) is able to induce a relaxant effect in porcine coronary, rat mesenteric artery, carotid, femoral arteries, and thoracic aorta and to determine the mechanism underlying this effect. Materials and Methods: Porcine coronary, rat mesenteric, carotid, femoral arteries, and thoracic aorta rings were suspended in organ chambers for the recording of changes in isometric forces. Rings with endothelium were incubated or not with L-Nitro Arginine to block nitric oxide (NO) synthase, MnTMPyP, polyethylene glycol catalase (CAT), an inhibitors of intracellular production reactive oxygen species; CAT, an inhibitor of extracellular reactive oxygen species; Wortmannin, an inhibitor of redox-sensitive pathway PI3 kinase/Akt; PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d]pyrimidine), an inhibitor of Src kinase; apamin (APA) an inhibitor of small conductance potassium channels calcium-dependent (SKCa) and Tram-34 an inhibitor of intermediary conductance potassium channels calcium-dependent (IKCa); and indomethacin (INDO), an inhibitor of cyclooxygenase before contraction with U46619 or phenylephrine (PE) and a concentration relaxation curve to TAE. In some experiments, the endothelium was removed before contraction with U46619 or PE and concentration relaxation to TAE. The phosphorylation level of Akt and eNOS was assessed by Western blot analysis. Results: The hydroethanolic bark TAE induces a vasodilatory effect in porcine coronary artery precontracted with U46619. This effect is endothelium-dependent and is mediated by NO, prostaglandin, and endothelium-derived hyperpolarizing. TAE also induces vasorelaxant effects in superior mesenteric arteries, carotid arteries, thoracic aorta, and femoral arteries from rat. Conclusion: TAE-induced sustaining phosphorylation of Akt and eNOS in endothelial cells. T. avicennioides induces vascular relaxation which can explain the benefic effect of this plant in the treatment of high blood pressure in Senegal.

Vasoactive properties of Ceiba pentandra in porcine coronary artery and different conductance and resistance vessels from rats: A role of nitric oxide

Background: Ceiba pentandra (L.) Gaertner (Malvaceae) a plant widely spread in the world, particularly in sub-Saharan Africa where it is used to handle several pathologies like high blood pressure without scientific evidence by experimental scientific works. Aim and Objective: The purpose of this study is to show how a hydroethanolic leaves extract of Ceiba pentandra (FCP) produces relaxant effects in porcine coronary, rat mesenteric artery, carotid, femoral arteries, and thoracic aorta and to determine the mechanisms which lie beneath this effect. Materials and Methods: Porcine coronary, rat mesenteric, carotid, and femoral arteries and thoracic aorta rings were suspended in organ chambers for recording of changes in isometric forces. Rings with endothelium were incubated or not with L-nitroarginine to block nitric oxide (NO) synthase, manganese (III) tetrakis(1-methyl-4-pyridyl)porphyrin, polyethylene glycol catalase (CAT), an inhibitors of intracellular production reactive oxygen species (ROS); CAT, an inhibitor of extracellular ROS; wortmannin, an inhibitor of redoxsensitive pathway phosphoinositide 3-kinase (PI3 kinase)/Akt; (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d] pyrimidine), an inhibitor of Src kinase; apamin, an inhibitor of small conductance potassium channels calcium dependent (SKCa) and tram-34, an inhibitor of intermediary conductance potassium channels calcium dependent (IKCa); and indomethacin, an inhibitor of cyclooxygenase before contraction with U46619 or phenylephrine and a concentration-relaxation curve to FCP. Results: Some experiments show that endothelium was taken away before contraction with U46619 and concentration relaxation to FCP. The hydroethanolic leaves extract of C. pentandra produces a vasodilatory result in porcine coronary artery pre-contracted with U46619. This effect is endothelium dependent and is mediated by NO. FCP generates, as well, vasorelaxant results in superior mesenteric arteries, carotid arteries, thoracic aorta, and femoral arteries from rat. C. pentandra generates vascular relaxation which can be the explanation of the benefic effect of this plant in the treatment of high blood pressure (HBP) in Africa. Conclusion: C. pentandra holds vasorelaxants properties on pig coronary artery, thoracic aorta, the main superior mesenteric artery, and femoral and carotid artery from rat justifying its utilization in the treatment of the arterial HBP. This effect necessitates the presence of a functional endothelium and passes by the redox-sensitive pathway Src kinase and PI3-kinase/Akt.

P5508Absolute coronary blood flow and microvascular resistance: correlation with instrumented coronary artery blood flow ring measurements and coronary histology analysis in swine

P5508Absolute coronary blood flow and microvascular resistance: correlation with instrumented coronary artery blood flow ring measurements and coronary histology analysis in swine

In Endothelial Cells, the Activation or Stimulation of Soluble Guanylyl Cyclase Induces the Nitric Oxide Production by a Mechanism Dependent of Nitric Oxide Synthase Activation.

In endothelial cells, investigate if the soluble guanylate cyclase (sGC) activation or stimulation is able to potentiate the relaxation in vessels. Aortic and coronary rings with and without endothelium were placed in a myograph and cumulative concentration-effect curves for DETA-NO or ataciguat were performed. Nitric oxide (NO) were measured by fluorescence or by selective electrode in human umbilical endothelial cells (HUVECs) in response to some treatments, including ataciguat, 8-Br-cGMP and A23187. The presence of the endothelium potentiated the relaxation induced by DETA-NO in aortic and coronary rings. In addition, in aortic rings the endothelium potentiated the relaxation induced by ataciguat. In the presence of nitric oxide synthase (NOS) inhibitor, the endothelium effect was abolished to DETA-NO or ataciguat, in both vessels. Ataciguat, 8-Br-cGMP and A23187 were able to induce NO production in HUVECs cells. In the presence of NOS inhibitor, the NO production induced by ataciguat and 8-Br-cGMP was abolished. Our results suggest that in aortic and coronary rings the endothelium potentiates the relaxation induced by activation or stimulation of sGC through a mechanism dependent of NOS activation.This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Regulation of Coronary Arterial Large Conductance Ca2+-Activated K+ Channel Protein Expression and Function by n-3 Polyunsaturated Fatty Acids in Diabetic Rats

Aim: The objective of this study was to examine the effects of n-3 polyunsaturated fatty acids (n-3 PUFAs) on coronary arterial large conductance Ca²⁺-activated K⁺ (BK) channel function in coronary smooth muscle cells (SMCs) of streptozotocin-induced diabetic rats. Methods: The effects of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) on coronary BK channel open probabilities were determined using the patch clamp technique. The mRNA and protein expressions of BK channel subunits were measured using qRT-PCR and Western blots. The coronary artery tension and coronary SMC Ca²⁺ concentrations were measured using a myograph system and fluorescence Ca²⁺ indicator. Results: Compared to nondiabetic control rats, the BK channel function was impaired with a reduced response to EPA and DHA in freshly isolated SMCs of diabetic rats. Oral administration of n-3 PUFAs had no effects on protein expressions of BK channel subunits in nondiabetic rats, but significantly enhanced those of BK-β₁ in diabetic rats without altering BK-α protein levels. Moreover, coronary ring tension induced by iberiotoxin (a specific BK channel blocker) was increased and cytosolic Ca²⁺ concentrations in coronary SMCs were decreased in diabetic rats, but no changes were found in nondiabetic rats. Conclusions: n-3 PUFAs protect the coronary BK channel function and coronary vasoreactivity in diabetic rats as a result of not only increasing BK-β₁ protein expressions, but also decreasing coronary artery tension and coronary smooth muscle cytosolic Ca²⁺ concentrations.

Bidirectional Regulatory Effects of Dexmedetomidine on Porcine Coronary Tone In Vitro.

BackgroundStudies in vivo have shown that dexmedetomidine (DEX) could protect the myocardium and modulate the coronary blood flow. This study aimed to investigate the direct and concentration-dependent effects of DEX on the tone of porcine coronary artery in vitro and the underlying mechanisms.Material/MethodsDistal branches of the porcine anterior descending coronary arteries were dissected and cut into 3–5 mm rings. The tones of coronary rings in response to cumulative DEX were measured using the PowerLab system. Coronary rings were divided into three groups: 1) endothelium-intact coronary rings without drug pretreatment (control); 2) endothelium-intact coronary rings pretreated with either yohimbine, tetraethylamine (TEA) or NG-nitro-L-arginine methyl ester (L-NAME); and 3) endothelium-denuded coronary rings pretreated with either yohimbine or TEA.ResultsDEX induced coronary ring relaxation at lower concentrations (10−9 to 10−7 M) followed by constriction at higher concentrations (10−6 to 10−5 M). The coronary constrictive effect of higher DEX (10−5 M) was greater in the endothelium-denuded rings than in the endothelium-intact rings. Yohimbine reduced the coronary constrictive effect of DEX at higher concentrations (10−6 to 10−5 M). TEA and L-NAME significantly reduced the coronary relaxing effect of DEX at lower concentrations (10−9 to 10−7 M) in endothelium-intact rings. TEA attenuated the coronary relaxation induced by DEX in endothelium-denuded rings.ConclusionsDEX exerts bidirectional effects on porcine coronary tone. The coronary relaxing effect of DEX at lower concentrations is likely associated with endothelium integrity, NO synthesis and BKCa channel activation, while the coronary constrictive effect of DEX at higher concentrations is mediated by α2 adrenoceptors in the coronary smooth muscle cells.

Ruthenium Complex Improves the Endothelial Function in Aortic Rings From Hypertensive Rats.

BackgroundThe endothelium is a monolayer of cells that extends on the vascular inner surface, responsible for the modulation of vascular tone. By means of the release of nitric oxide (NO), the endothelium has an important protective function against cardiovascular diseases.ObjectiveVerify if cis- [Ru(bpy)2(NO2)(NO)](PF6)2 (BPY) improves endothelial function and the sensibility of conductance (aorta) and resistance (coronary) to vascular relaxation induced by BPY.MethodsNormotensive (2K) and hypertensive (2K-1C) Wistar rats were used. For vascular reactivity study, thoracic aortas were isolated, rings with intact endothelium were incubated with: BPY(0.01 to10 µM) and concentration effect curves to acetylcholine were performed. In addition, cumulative concentration curves were performed to BPY (1.0 nM to 0.1 µM) in aortic and coronary rings, with intact and denuded endothelium.ResultsIn aorta from 2K-1C animals, the treatment with BPY 0.1µM increased the potency of acetylcholine-induced relaxation and it was able to revert the endothelial dysfunction. The presence of the endothelium did not modify the effect of BPY in inducing the relaxation in aortas from 2K and 2K-1C rats. In coronary, the endothelium potentiated the vasodilator effect of BPY in vessels from 2K and 2K-1C rats.ConclusionOur results suggest that 0.1 µM of BPY is able to normalize the relaxation endothelium dependent in hypertensive rats, and the compound BPY induces relaxation in aortic from normotensive and hypertensive rats with the same potency. The endothelium potentiate the relaxation effect induced by BPY in coronary from normotensive and hypertensive rats, with lower effect on coronary from hypertensive rats.

Effects of acrylic resin monomers on porcine coronary artery reactivity

The purpose of the present investigation was to assess the reactivity of porcine coronary arteries under in vitro conditions following their exposure to methyl methacrylate (MMA) and hydroxyethyl methacrylate (HEMA) monomers. Confirming previous studies using rat aortas, both MMA and HEMA induced acute/direct relaxation of coronary ring preparations, which was partly dependent on the endothelium. With prolonged tissue exposure, both monomers caused time- and concentration-dependent inhibition of receptor-mediated contraction of the vascular smooth muscle caused by prostaglandin F2∝ (PGF2∝), with HEMA causing more inhibition than MMA. Hydroxyethyl methacrylate, but not MMA, also produced impairment of non-receptor-mediated contraction of the coronary smooth muscle induced by KCl. On the other hand, neither HEMA nor MMA altered relaxation of the smooth muscle produced by the direct-acting pharmacological agent, sodium nitroprusside (SNP). While exposure to HEMA impaired endothelium-dependent vasorelaxation caused by bradykinin (BK), MMA markedly enhanced this endothelial-mediated response of the arteries. The enhanced endothelial response produced by MMA was linked to nitric oxide (NO) release. In conclusion, with prolonged tissue exposure, MMA causes less pronounced effects/adverse consequences on coronary smooth muscle function relative to the effect of HEMA, while enhancing vasorelaxation associated with release of NO from the endothelium. Accordingly, MMA-containing resin materials appear to be safer for human applications than materials containing HEMA.

Clinical experience of penile elongation: a comparison of four different operative approaches

To investigate the curative effect of penile elongation with four differentoperative approaches. Through four different operative approaches (the coronary sulcus ringincision, Y or Z shaped incision or Z shaped incision combined with coronary sulcus ring incision), thepenile skin and fascia were degloved until the penile root. Then the superficial and deep dorsal penilesuspensory ligament were cut off. After electric coagulation of the residue ends, the two-side tissue at thefront of the pubic symphysis was sutured. Then the penile skin and fascia were repositioned and the incisionat the inner and outer plate was closed. The increased penile static length was (2.9 ± 0.2) cmwith abdominal wall Y incision (12 cases); (3.1 ± 0.3) cm with transabdominal modified Z incision (260 cases); (3.9 ± 0.7) cm with coronary sulcus ring incision (363 cases); (3.4 ± 0.8) cm with combined incision (39 cases). The lengthening effect was significantly different between the coronary ring incision and abdominal wall Y/Z incision (P < 0.05). The postoperative follow-up period was 6 months to 5.5 years without serious complications. Only 3 cases of subcutaneous hematoma occurred with treatment of debridement and drainage. 4 cases with ischemic necrosis at distal penile skin, were treated with debridement, dressing and physiotherapy, leaving no scar. Penile lengthening surgery are safe and effective through different approaches. The coronal ring incision has the best therapeutic effect.

Vanillin and vanillin analogs relax porcine coronary and basilar arteries by inhibiting L-type Ca2+ channels.

Vanillin (VA) and vanillyl alcohol (VAA), components of natural vanilla, and ethyl vanillin (EtVA; synthetic analog) are used as flavoring agents and/or as additives by the food, cosmetic, or pharmaceutic industries. VA, VAA, and EtVA possess antioxidant and anti-inflammatory properties, but their vascular effects have not been determined. Therefore, we compared in isolated porcine coronary and basilar arteries the changes in isometric tension caused by VA, VAA, and EtVA. VA and its analogs caused concentration-dependent relaxations of both preparations during contractions from U46619 (9,11-dideoxy-11α,9α-epoxymethanoprostaglandin F2α, a thromboxane A2 receptor agonist), and of coronary arteries contracted with KCl or endothelin-1. The order of potency was VAA < VA < EtVA. The relaxations were not inhibited by endothelium removal, by inhibitors of NO synthases (N(ω)-nitro-l-arginine methyl ester hydrochloride), cyclooxygenases (indomethacin), soluble guanylyl cyclase (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one [ODQ]), KCa (1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole [TRAM-34], 6,12,19,20,25,26-hexahydro-5,27:13,18:21,24-trietheno-11,7-metheno-7H-dibenzo[b,n][1,5,12,16]tetraazacyclotricosine-5,13-diium ditrifluoroacetate hydrate [UCL-1684], or iberiotoxin), by KATP (glibenclamide), by Kir (BaCl2), by transient receptor potential receptor vanilloid 3 (TRPV3) channels (ruthenium red), or by antioxidants (catalase, apocynin, tempol, N-acetylcysteine, tiron). VA and its analogs inhibited contractions induced by Ca(2+) reintroduction in coronary arteries, and by an opener of L-type Ca(2+)-channels (methyl 2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]-1,4-dihydropyridine-3-carboxylate [Bay K8644]) in coronary and basilar arteries. They inhibited contractions of coronary rings induced by the protein kinase C activator phorbol 12,13-dibutyrate to the same extent as the removal of extracellular Ca(2+) or incubation with nifedipine. Thus, in porcine arteries, relaxation from VA (and its analogs) is due to inhibition of L-type Ca(2+) channels. Hence, these compounds could be used to relieve coronary or cerebral vasospasms due to exaggerated Ca(2+) influx, but therapeutic efficacy would require exposures that far exceed the current levels obtained by the use of vanillin additives.

Increased Ca2+‐activated Ca2+ influx and impaired Ca2+ buffering in coronary smooth muscle from metabolic syndrome Ossabaw swine (1076.2)

Coronary smooth muscle (CSM) phenotype is affected by intracellular Ca2+ regulation and we have previously correlated changes in CSM Ca2+ signaling from Ossabaw swine with varying stages of coronary artery disease (CAD). Ossabaw swine are a model of metabolic syndrome (MetS) and CAD. Three year old pigs were fed an excess‐calorie, atherogenic or control diet for 6 months. CAD progression, coronary artery function, and CSM Ca2+ signaling were assessed. MetS coronaries had advanced, lipid‐laden atherosclerotic lesions containing calcium deposits, while lean had no CAD. In isometric tension studies diseased coronary rings generated significantly less tension than healthy rings in response to depolarization with 20 mM K+. CSM were enzymatically isolated and loaded with the Ca2+ indicator, fura‐2 AM. Diseased CSM showed less K+‐induced Ca2+ influx compared to healthy cells. Diseased cells recovered less from the cytosolic Ca2+ increase back to baseline, suggesting impaired Ca2+ buffering. In physiologic salt solution (PSS), there was no difference in peak Ca2+ transient elicited by caffeine‐induced intracellular Ca2+ store release. The caffeine‐induced Ca2+ transient in Ca2+‐free PSS, which represents exclusively sarcoplasmic reticulum (SR) Ca2+ release, was decreased in diseased cells. Taken together, these data suggest that lesser SR Ca2+ release elicited greater Ca2+‐activated Ca2+ influx in diseased CSM. After caffeine challenge healthy CSM buffered Ca2+ significantly below basal Ca2+ compared to diseased CSM. Overall, the data suggest impaired CSM SR Ca2+ buffering, decreased SR Ca2+ store, and increased Ca2+‐activated Ca2+ influx in CAD.Grant Funding Source: Supported by NIH HL062552, HL092245, NIH/NCATS CTSI TL1 TR000162

ASSA13-03-19 Function Role on Coronary Artery Tone of IP3 by Activating IP3R and BKCa of Porcine Coronary Artery Smooth Muscle Cells

<h3>Background and Objective</h3> The large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel mediates coronary relaxation. However, endogenous factors that regulate BK_Ca channel activities are not well defined. The present study sought to test the hypothesis that inositol trisphosphate (IP3) is an activator of BK_Ca channels in coronary smooth muscle cells (SMCs) and therefore would relax the coronary tone. <h3>Methods and Results</h3> Porcine coronary artery rings were prepared to perform the vasomotor assay. The whole-cell voltage clamp and single-channel patch clamp experiments were performed in freshly isolated coronary SMCs to test the effect of IP3 on BK_Ca channels and to figure out the underlying mechanisms. The results showed that blocking IP3 receptor (IP3R) with xestospongin-C (XeC) increased the tension of coronary rings. After pre-constricting with 0.1 μM PGF2α, the use of XeC (2 μM) induced a further increase of LAD tension by 26.10 ± 5.73% (n = 4) of that induced by PGF2α, and this effect of XeC could be abolished by pretreatment with IbTX, a specific blocker of BK_Ca. In the perforated whole-cell recordings, in which intracellular IP3 leak cannot occur, our data confirmed that IP3 had a physiological contribution to STOC generation (approximately 30% of the STOCs are XeC-sensitivive). Under traditional whole-cell recording, introduction of exogenous IP3 (0.5–50 μM) from the pipette solution into the cytoplasm increased the XeC-sensitive STOC roughly by 30% to 50% and this effect could be abolished by XeC, suggesting that exogenous IP3 stimulates the STOC via IP3R. However, it is likely that IP3 impacts the macroscopic current (MC, another component of BKCa currents) by a differential signalling pathway. Although IP3 increased the MC, this effect could be abolished neither by XeC, nor by pretreating with 1μM thapsigargin (a depletor of SR Ca2 + store), indicating an IP3R-independent way was involved. Interestingly, in the inside-out single channel recordings, application of IP3 (10–50 μM) to the cytosolic side of the membrane increased the single BK_Ca channel currents likely by enhancing the Ca²⁺ sensitivity of BK_Ca channel and as a result, changed the channel kinetics. Further, we identified that these effects were IP3R-independent. <h3>Conclusions</h3> We conclude that IP3 is an activator of BK_Ca channels in the porcine coronary SMCs and can relax coronary tone.

Effects of synthetic analogues of poly-APS on contractile response of porcine coronary arteries

APS12-2 and APS3 are synthetic analogues of polymeric alkylpyridinium salts (poly-APS) isolated from the marine sponge Reniera sarai. The aim of the present study was to determine the possible direct contractile effects of these two synthetic molecules on coronary arteries, in order partly to explain hemodynamic and cardiotoxic effects of APS12-2 previously observed in in vivo studies and to reveal possible adverse effects on the organism in the case of their clinical use. In contrast to APS3, APS12-2 caused a concentration-dependent vascular smooth muscle contraction of isolated porcine coronary ring preparations in a concentration-range from 1.36 to 13.60μM. Lanthanum chloride (5mM) and verapamil (10μM) completely abolished the APS12-2 evoked contraction of the coronary rings. Pre-incubation with indomethacin (10μM) had no effect on the contractile responses of coronary ring preparations. These results indicate that APS12-2 contracts vascular smooth muscle in a concentration-dependent manner, due to an increase of Ca2+ influx through the voltage-gated Ca2+ channels. Our data show for the first time that APS12-2 induces concentration-dependent contraction of coronary ring preparations, which may contribute to the cardiotoxic effects of APS12-2, in addition to hyperkalemia.

Involvements of calcium channel and potassium channel in Danshen and Gegen decoction induced vasodilation in porcine coronary LAD artery

Danshen (Salviae Miltiorrhizae Radix) and Gegen (Puerariae Lobatae Radix) have been widely used in treating cardiovascular diseases for thousands of years in China. The present study was carried out to evaluate the effects of a Danshen and Gegen decoction (DG) on the vascular reactivity of a porcine isolated coronary artery and the underlying mechanisms involved. Porcine coronary rings were precontracted with 15nM U46619. The involvement of endothelium-dependent mechanisms was explored by removing the endothelium; the involvement of potassium channels was investigated by the pretreatment of the artery rings with various blockers, and the involvement of the calcium channels was investigated by incubating the artery rings with Ca2+-free buffer and priming them with high [K+] prior to adding CaCl2 to elicit contraction. The involvement of Ca2+ sensitization was explored by evaluating the Rho-activity expression.The results revealed that DG elicited a concentration-dependent relaxation on a U46619-precontracted coronary artery ring. These relaxation responses were not altered by the pretreatment of inhibitors of endothelium-related dilator synthases, cGMP and cAMP pathway inhibitors, potassium channel (BKCa, SKCa, KV and KATP) blockers and endothelium removal. The KIR channel blocker BaCl2 only slightly attenuated the DG-induced relaxation. However, the Ca2+-induced artery contraction was inhibited by DG. Additionally, the expression of the phosphorylated myosin light chain was inhibited by DG whereas the activity of RhoA was not affected. Therefore, DG could be a useful cardioprotective agent for vasodilation in patients who have hypertension.

Low-dose celecoxib improves coronary function after acute myocardial ischaemia in rabbits

The role of celecoxib in cardiovascular events remains contentious. The aim of the present study was to investigate the effects of celecoxib in acute myocardial ischaemia (AMI) in rabbits in comparison with those of another non-steroidal anti-inflammatory drug, namely aspirin. Male New Zealand white rabbits were divided into four groups: (i) a sham-operated group; (ii) an AMI group, in which the left anterior descending coronary arteries were occluded for 60min; (iii) the celecoxib+AMI group, pretreated with 3mg/kg celecoxib, twice a day, for 3days before AMI induction; and (iv) the aspirin+AMI group, pretreated with 12.5mg/kg aspirin, twice a day, for 3days before AMI induction. Haemodynamic parameters were monitored using a multichannel physiological recorder. Serum levels of creatine kinase (CK), malondialdehyde (MDA), cyclo-oxygenase-2 (COX-2), tumour necrosis factor (TNF)-α, total nitrate/nitrite (NO(x) ), nitric oxide synthase (NOS) and myocardial infarct size were determined. Changes in isometric tension of isolated coronary rings were recorded by a myograph system. Compared with the sham group, the AMI group had lower blood pressure, higher left ventricular (LV) end-diastolic pressure, depressed maximum dP/dt of LV pressure, a larger infarct size and higher CK and MDA levels. Celecoxib, but not aspirin, pretreatment significantly ameliorated these effects of AMI. Celecoxib reversed AMI-induced increases in COX-2 levels to a similar extent as aspirin. Pretreatment with celecoxib resulted in a significant reduction in TNF-α levels and an increase in NO(x) and NOS levels compared with the AMI group. The dysfunctional vasoconstriction and vasodilation of coronary arteries were ameliorated by celecoxib administration. 4. In conclusion, the experimental evidence suggests that celecoxib exerts its protective effects in a COX-independent manner.

Abstract P090: The Contractile Effect of Angiotensin II Type 1 Receptor Autoantibodies on Human Placental Vessels

Background: Decreased placental perfusion induced by abnormal placental vascular contraction is one of the pathological basis of preeclampsia. It has been reported that the sera titers of the autoantibody against the second extracellular loop of angiotensin II type 1 receptor (AT1-AA) were negatively correlated with placental blood flow in preeclampsia. Our previous study has found that AT1-AA could induce contraction of rat thoracic aorta and coronary rings by activing angiotensin II type 1 receptor (AT1R). However, there is no direct evidence for explaining whether AT1-AA might cause vasoconstriction on human placental blood vessels. Methods: The SD rats were immunized with the synthetic peptide corresponding to the sequence of the second extracellular loop of the human AT1 receptor (AT1R-EC II ), and anti-AT1R antibody (AT1R-Ab) was extracted. The expression of AT1R on human placental vessels was determined by immunohistochemistry. The effects of AT1R-Ab on placental vessels were measured with isolated vascular ring technique. Results: (1) AT1R was highly expressed in the human placental artery, vein and vascular endothelial cells. (2) AT1R-Ab (10 mmol/L) respectively enhanced the contraction of the placental arteries and veins (29.21% ± 3.7% vs . 21.35% ± 2.8%, P &gt;0.05), which could be completely reversed when AT1R was blocked by AT1R inhibitor. (3) AT1R-Ab (0.1, 1 and 10 mmol/L) induced placental vasoconstriction of normal human. The percentage of maximal contraction was 2.73% ± 1.11%, 4.00% ± 3.2% and 33.30% ± 5.6%, respectively. There was significantly difference between the three groups for contraction amplitude induced by different concentrations of AT1R-Ab ( P &lt;0.01). (4) AT1R-Ab (0.1, 1 and 10 mmol/L) induced placental vasoconstriction of preeclampsia. The percentage of maximal contraction was 1.74% ± 0.3%, 5.58% ± 1.41% and 3.73% ± 2.5%, respectively. There was significantly difference between the three groups for contraction amplitude induced by different concentrations of AT1R-Ab ( P &lt;0.01). Conclusion: AT1R-Ab could induce human placental vasoconstriction in a concentration-dependent manner, which suggested that AT1-AA might be involved in the pathogenesis of preeclampsia by directly contracted placental blood vessles.

Pre-conditioning with the soluble guanylate cyclase activator Cinaciguat reduces ischaemia–reperfusion injury after cardiopulmonary bypass

Activation of the nitric oxide-soluble guanylate cyclase-cyclic guanosine monophosphate (NO-sGC-cGMP) pathway can induce potent cardioprotection-like effects against ischaemia-reperfusion injury and nitro-oxidative stress. We investigated the effects of pharmacological pre-conditioning with Cinaciguat (BAY 58-2667), a novel sGC activator on peroxynitrite-induced endothelial dysfunction in vitro, as well as on myocardial and coronary vascular function during reperfusion in a canine model of cardioplegic arrest and extracorporeal circulation. Isolated coronary arterial rings exposed to peroxynitrite were investigated for vasomotor function. Vehicle- and Cinaciguat-pre-treated (8.33 μg h(-1) or 25 μg h(-1) intravenous (IV) for 30 min) anaesthetised dogs (n = 6-7 per group) underwent hypothermic cardiopulmonary bypass with 60 min of hypothermic cardioplegic arrest. Left- and right-ventricular end-systolic pressure-volume relationship (ESPVR) was measured by a pressure-volume conductance catheter at baseline and after 60 min of reperfusion. Coronary blood flow, vasodilatation to acetylcholine and myocardial level of adenosine triphosphate were determined. Pre-incubation of coronary rings with Cinaciguat improved peroxynitrite-induced endothelial dysfunction. Compared with control, pharmacological pre-conditioning with Cinaciguat (25 μg h(-1)) led to higher myocardial adenosine triphosphate content, to a better recovery of left- and right-ventricular contractility (Δ slope of left ventricular ESPVR given as percent of baseline: 102.4 ± 19.1% vs 56.0 ± 7.1%) and to a higher coronary blood flow (49.6 ± 3.5 ml min(-1) vs 28.0 ± 3.9 ml min(-1)). Endothelium-dependent vasodilatation to acetylcholine was improved in the treatment groups. Pre-conditioning with Cinaciguat improves myocardial and endothelial function after cardiopulmonary bypass with hypothermic cardiac arrest. The observed protective effects imply that pharmacological sGC activation could be a novel therapeutic option in the protection against ischaemia-reperfusion injury in cardiac surgery.

Effect of exercise‐training on EDHF‐mediated relaxation after chronic coronary occlusion

Endothelium‐derived hyperpolarizing factors (EDHF) play an important role in maintaining vasodilatory capacity when nitric oxide bioavailability is impaired such as in coronary artery disease. We hypothesized that exercise training would increase the contribution of EDHF to endothelium‐dependent relaxation in coronary arteries from a model of ischemic heart disease. Ameroid constrictors were surgically placed around the proximal left circumflex coronary artery (LCX) of adult female Yucatan miniswine. Coronary rings (~1 mm luminal diameter) were isolated from the collateral‐dependent (LCX) and control (nonoccluded left anterior descending, LAD) arteries of sedentary (SED, pen confined) and exercise trained (EX, treadmill run; 5 days/wk for 14 wks) pigs. NOS inhibition (L‐NAME) attenuated bradykinin‐mediated relaxation to a greater extent in SED compared with EX of both LAD and LCX. Combined inhibition of NOS and prostanoids (indomethacin) partially corrected the EX‐enhanced relaxation observed after NOS inhibition alone, while concurrent inhibition of large‐conductance potassium (BKca) channels (iberiotoxin) ablated the differences. Thus the contribution of endothelium‐derived vasodilators is altered by EX in the underlying setting of ischemic heart disease, with an increased contribution of an EDHF acting via BKca channel, however, the specific EDHF remains undetermined. NIH:HL064931