Dog Coronary Arteries Research Articles

Evidence That Contractions of Isolated Arteries by l-NMMA and NOLA Are Not Due to Inhibition of Basal EDRF Release

Summary: We have tested the hypothesis that endothelium-dependent contractions to two l-arginine analogues modified in either of the guanidino nitrogens [e.g., NG-monomethyl-l-arginine (l-NMMA) and Nω-nitro-l-arginine (NOLA)] are due to inhibition of basally released EDRF. We found that these contractions in the dog isolated coronary artery were maximal at 10 μM for each compound, as increasing the concentration 10-fold further gave no significant increase in developed force. Over the same concentration range, NOLA was more potent than l-NMMA in blocking relaxations to the endothelium-dependent agonist, acetylcholine. Thus, for l-NMMA, only the highest concentrations (30-100 μM) caused a small but significant depression of the maximum response with no effect on sensitivity (i.e., EC50), whereas for NOLA (10-100 μM), there was a progressive, concentration-dependent decrease in both sensitivity and the maximum response. Oxyhemoglobin (Hb; 1-3 μM) and FeSO4 (3 mM) caused a 4- to 10-fold rightwards displacement of the relaxation curve to sodium nitroprusside and blocked totally the response to nitric oxide (generated from acidified sodium nitrite), respectively. Under these conditions, the contractions to l-NMMA were virtually unaffected. These data suggest that contractions to both l-NMMA and NOLA in the dog coronary artery were not caused by the removal of effects of basally released EDRF.

Streptokinase Inhibits Acute Platelet Thrombus Formation in Stenosed Dog Coronary Arteries

Previous evidence has suggested that plasmin, in addition to its proteolytic action on fibrin, may affect platelet function. To test the effects of plasmin generated in vivo by the thrombolytic agent streptokinase (SK) on platelet-dependent vascular occlusion, we have used a well-established canine model of experimental coronary artery stenosis which produces platelet aggregate-dependent cyclical variations in coronary blood flow. Infusion of SK into 22 dogs at doses sufficient to cause a systemic lytic state led to complete abolition of cyclical blood flow reductions (CFR's) at sites of coronary artery injury. Inhibition of coronary platelet occlusion was associated with marked prolongations of the bleeding time (from 3.2 ± 0.6 min before to 14 ± 5 min after SK infusion, mean ± SD, n = 22). Despite the striking effects of SK on in vivo platelet-vessel wall interactions, only platelet aggregation in response to collagen was diminished among the ex vivo parameters of platelet function that were studied simultaneously. Platelet aggregation in response to other agonists, thromboxane A2 production, monoclonal antibody binding to platelet membrane glycoprotein (Gp) IIb-IIIa, Gp Ib-dependent botrocetin-induced platelet aggregation and platelet levels of cyclic AMP were not significantly altered. Therefore the thrombolytic agent streptokinase appears to cause important inhibitory effects on in vivo platelet reactivity with injured vascular intimal surfaces, possibly due to localised changes in platelet aggregate formation in the microenvironment of exposed collagen. These findings suggest that plasmin generated by thrombolytic agents may exhibit platelet inhibitory activity, and that this effect may be important in reestablishing blood flow in certain forms of platelet-mediated arterial thromboses.

Investigation of the interrelationship between coagulation and thrombin-induced EDRF-dependent relaxation in dog coronary artery

Little is known about the possible interrelationships between thrombin-induced EDRF-dependent vascular relaxation and coagulant activity. We have now studied the effects of the anticoagulant zymogen protein C, on EDRF-dependent relaxation in isolated canine coronary arteries. Low concentrations of activated protein C (0.1–30 ng/ml) had no significant effect, but higher concentrations caused relaxation (E max−39.2 ± 7.2%; 100 – 1000 ng/ml). To determine whether relaxation was dependent on coagulation complexes associated with endothelial cell membranes, the coumarin, brodifacoum was given three days before in vitro experiments were carried out in order to inhibit production of active vitamin K1-dependent clotting factors. Brodifacoum (10 mg/kg i.p.) increased prothrombin time from 8.5 ± 0.24 sec (control), to 46.2 ± 2.4 sec (p< 0.05), but had no effect on thrombin-induced relaxation (E max >90%; ED50 0.026 ± 0.004 units/ml control; 0.025 ± 0.004 unit/ml brodifacoum). In the final group of studies, we investigated the effects of the concomitant administration of protein C (1000 ng/ml) and thrombin in vitro. Protein C (1000 ng/ml) increased relaxant sensitivity to thrombin after partial desensitization of the relaxant response by previous thrombin administration, (−60.2 ± 7.1% thrombin alone; −77.9 ± 7.2% thrombin + protein C), but had no effect after complete desensitization of the relaxant response. In conclusion, the data appear best explained by protein C and thrombin-induced EDRF dependent relaxation being due to proteolytic actions.

Effects of video frame averaging, smoothing and edge enhancement on the accuracy and precision of quantitative coronary arteriography

Digital analysis of cine film provides numerous options for altering images by frame averaging or filtering algorithms that either smooth or enhance edges. While these may subjectively enhance image quality, there is no uniformity in their use among laboratories and effects on quantitative coronary analysis may not be ideal. To determine which processing algorithms might help or hinder quantitative coronary arteriography, cine film images of precision drilled stenotic cylinders (0.83 to 1.83 nm diameter) implanted in dog coronary arteries were analyzed with and without such algorithms. Video frame averaging of 1 to 49 frames had no effect on measures of accuracy (mean differences) but precision (standard deviation of mean differences) was improved from 0.23 to 0.17 mm (p less than 0.05) with video averaging of greater than or equal to 25 frames. Edge enhancement filtering algorithms resulted in slight deterioration of accuracy and precision and smoothing filtering algorithms caused modest improvements in these parameters; however, these changes were not significantly different from unprocessed images. Using edge enhancement filtering algorithms, accuracy was significantly worse (-0.27 mm) compared to a smoothing filter enhancement algorithm (-0.08 mm, p less than 0.001). The combination of video averaging and smoothing algorithms had no additional beneficial effects. Thus, precision of quantitative coronary analysis of cine film can be optimized by appropriate video averaging. Edge enhancement filtering algorithms should be avoided whereas smoothing filter enhancement algorithms may improve accuracy.

Mechanism underlying the response to vasodilator nerve stimulation in isolated dog and monkey cerebral arteries

Relaxant responses to transmural electrical stimulation and nicotine of cerebral artery strips obtained from dogs and Japanese monkeys were abolished by tetrodotoxin and hexamethonium, respectively, and suppressed by treatment with NG-monomethyl-L-arginine (L-NMMA), a nitric oxide (NO) synthesis inhibitor. The inhibitory effect was prevented and reversed by L-arginine but not by D-arginine. The relaxations suppressed by L-NMMA were not increased by exogenously applied NO. Endothelium denudation did not alter the response to transmural stimulation and nicotine or the inhibitory effect of L-NMMA. D-NMMA did not inhibit the response to vasodilator nerve stimulation. Dog coronary artery relaxations caused by transmural stimulation were not inhibited by L-NMMA but reversed to contractions by propranolol. Relaxations caused by substance P of dog cerebral arteries treated with indomethacin were dependent on endothelium and inhibited by L-NMMA, whereas those by NO and nitroglycerin, endothelium-independent relaxations, were unaffected. It is concluded that chemical and electrical stimulation of vasodilator nerves relaxes dog and monkey cerebral arteries, possibly by a mediation of NO rather than a stimulating action of NO on the release of vasodilator transmitter. Endothelium-dependent relaxations by substance P of dog cerebral arteries appear to be mediated by NO.

Spasmolytic effect of cromakalim in the dog coronary artery

Spasmolytic effect of cromakalim in the dog coronary artery

Autoregulation by the right coronary artery in dogs with open chests; comparison with the left coronary artery

Experiments were conducted to study autoregulatory responses of the right and left coronary arteries in dogs with open chests. The right and left circumflex coronary artery were cannulated and perfused with blood from the femoral artery via a pressurized reservoir. The perfusion pressure was varied in steps over a wide range and coronary blood flow rates were measured. Both the right and left coronary arteries exhibited autoregulation but the pressure at the end of the autoregulatory range was lower in the right (39.8 +/- 9.1 mm Hg) than in the left circumflex coronary artery (57.6 +/- 14.5 mm Hg). The slope of the pressure-flow relationship in the autoregulatory range was less steep in the right than the left circumflex coronary artery. The closed-loop gain when the perfusion pressure was less than 100 mm Hg was greater in the right than in the left circumflex coronary artery. Increases in the right ventricular afterload produced by pulmonary artery constriction decreased the closed-loop gain, shifted the auto-regulatory range upward and to the right, and made the slope steeper. These results indicate that more effective autoregulation is carried out by the right than the left circumflex coronary artery.

Glibenclamide is a competitive antagonist of the thromboxane A2 receptor in dog coronary artery in vitro

1. Glibenclamide, a sulphonylurea oral hypoglycaemic agent is a widely used antagonist of cromakalim-activated K+ channels in smooth muscle. 2. In isolated ring segments of the large circumflex coronary artery from the dog, glibenclamide (1-30 microM) caused a concentration-dependent reduction in both spontaneous isometric force and contractions induced by U46619, a thromboxane A2-mimetic. 3. Glibenclamide behaved as a competitive antagonist of U46619 with an estimated pKB (-log KB) value of 6.2 by Schild regression analysis (slope 1.07). 4. Glibenclamide (30 microM) was apparently selective since it had no effect on the concentration-contraction curves to endothelin-1, noradrenaline or KCl. 5. We suggest that this additional property of glibenclamide should be considered in any smooth muscle study where active force is raised by either the exogenous application or endogenous generation of thromboxane A2.

Adenosine constricts the isolated and perfused monkey coronary artery

Adenosine constricted 37 of 58 isolated and perfused monkey coronary arteries, whereas it dilated dog coronary arteries in all cases. The vasoconstriction induced by 0.1-10 micrograms (0.37-37.4 nmol) of adenosine was examined, since adenosine in large doses produced tachyphylaxis. The adenosine-induced vasoconstriction was significantly attenuated by diltiazem (100 micrograms [0.24 mumol]), although it was not affected by aminophylline (100 micrograms [0.23 mumol]), phentolamine (100 micrograms [0.36 mumol]), or methysergide (10 micrograms [28 nmol]. Forty-minute treatment with indomethacin (5 x 10(-6) mol/l) completely prevented the adenosine-induced vasoconstriction, but it did not significantly inhibit vasoconstriction induced by KCl (3 mg [40 mumol])- or norepinephrine (NE, 0.1 micrograms [0.59 nmol]). These results suggest that: (1) adenosine constricts monkey coronary arteries by increasing Ca2+ influx across the smooth muscle cell membrane; (2) cyclooxygenase products may be involved in the adenosine-induced vasoconstriction; (3) adenosine receptors may not be involved in the vasoconstriction.

Comparison of relaxation responses of vascular and non-vascular smooth muscle to endothelium-derived relaxing factor (EDRF), acidified sodium nitrite (NO) and sodium nitroprusside

Smooth muscle relaxant activity of endothelium-derived relaxing factor (EDRF) released from columns of cultured bovine endothelial cells by bradykinin (0.1-3 nmol/l) was measured in four non-vascular preparations: guinea-pig taenia caeci, guinea-pig trachea, rat stomach (fundus) and rat anococcygeus. Each preparation was contracted to a steady level of force with a variety of agonists such that they relaxed optimally to sodium nitroprusside (SNP). The EDRF-induced relaxations in each preparation were compared with those obtained in de-endothelialized ring preparations of greyhound coronary artery by means of paired bioassays run in parallel. EDRF released from the endothelial cell columns caused 80-100% relaxation of the coronary artery, 40-80% in the guinea-pig taenia caeci, 50-70% in the rat anococcygeus, 5-8% in the guinea-pig trachea and was undetectable in the rat stomach strip. By comparison, SNP caused maximal relaxation in all tissues compared with the coronary artery. In separate organ bath experiments the sensitivity to nitric oxide (NO: generated by adding acidified solutions of NaNO2) and SNP was compared in each preparation. SNP caused maximal relaxation in all tissues with the following order of potency: dog coronary artery greater than guinea-pig trachea greater than guinea-pig taenia = rat anococcygeus greater than rat stomach strip. In contrast, the concentration of acidified NaNO2 (NO, 300 nmol/l) that caused 96 +/- 4% relaxation in the dog coronary artery caused 84 +/- 7% and 48 +/- 1% relaxation in the taenia and anococcygeus respectively. No response attributable to NO was detected in either the trachea or rat stomach strip.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholinergic prejunctional inhibition of vasodilator nerve function in bovine basilar arteries

Helical strips of bovine basilar arteries responded to transmural electrical stimulation with moderate relaxations that were abolished by treatment with tetrodotoxin and oxyhemoglobin and were significantly attenuated by quinidine, as were the relaxations previously seen in dog and monkey cerebral arteries. The relaxant response of the bovine arteries was attenuated by treatment with acetylcholine and physostigmine and was significantly potentiated by atropine. In contrast, relaxations of dog coronary arteries to transmural stimulation, mediated via beta-adrenoceptors, were not influenced by physostigmine and atropine but were attenuated by acetylcholine. Abundant fibers containing cholinesterase were histologically demonstrated in bovine basilar arteries, suggesting the presence of cholinergic innervation. Acetylcholine liberated from the cholinergic nerve appears to act on muscarinic receptors located in vasodilator nerve terminals in bovine cerebral arteries and to interfere with the release of vasodilator transmitters.

Aspirin and dipyridamole and their limitations in the therapy of coronary artery disease

We have reviewed some of the voluminous literature on the effects of aspirin combined with dipyridamole on coronary thrombosis. There is clear evidence that aspirin is partially effective in preventing platelet aggregation and subsequent thrombosis in experimental constricted and damaged coronary arteries of dogs. Clinical studies show a clear reduction in myocardial infarction in male human subjects who are given aspirin as therapy for unstable angina, or as prophylaxis in asymptomatic individuals. In many studies aspirin and dipyridamole have been combined and are effective. We have not found dipyridamole to be effective in the dog with coronary artery constriction and find no substantial evidence that it is effective in preventing myocardial infarction in man. Until definitive studies show that combining dipyridamole with aspirin is more effective than aspirin alone, we do not recommend its use for prevention of coronary thrombosis.

Synthesis of lipoxygenase and epoxygenase products of arachidonic acid by normal and stenosed canine coronary arteries.

Coronary vascular injury promotes blood cell-vessel wall interactions that influence arachidonic acid metabolism and coronary blood flow patterns. Since lipoxygenase and cytochrome P-450 epoxygenase metabolites of arachidonic acid are synthesized by vascular and inflammatory cells and have a variety of important biological actions, we investigated the metabolism of arachidonic acid by these pathways in normal and stenosed, endothelially injured canine coronary arteries. We found and confirmed by gas chromatography/mass spectrometry that primarily 12- and 15-hydroxyeicosatetraenoic acids (HETEs) are synthesized by both coronary artery segments. Lesser amounts of 11-, 9-, 8-, and 5-HETEs are also produced. 15-Ketoeicosatetraenoic acid is also synthesized. The synthesis of 14C-HETEs is fivefold to 10-fold greater by the stenosed than the normal coronary artery. Specific radioimmunoassays indicated that the stenosed coronary artery synthesized 93 +/- 14 and 1,102 +/- 154 ng/g of tissue of 15- and 12-HETE, respectively, while the normal coronary artery produced 17 +/- 3 and 162 +/- 68 ng/g of tissue of 15- and 12-HETE, respectively. Products comigrating with 14,15-; 11,12-; 8,9-; and 5,6-epoxyeicosatrienoic acids (EETs) and the corresponding dihydroxyeicosatrienoic acids (DHETs) were detected predominantly in stenosed coronary arteries by high-pressure liquid chromatography. The structures of the EETs were confirmed by GC/MS. The EETs and prostaglandin I2 produced endothelium-independent, concentration-related relaxations of dog coronary artery rings. These data indicate that normal and stenotic coronary arteries metabolize arachidonic acid to HETEs, DHETs, and EETs along with prostaglandins; however, the synthesis of these metabolites is greater in the stenosed, endothelially injured vessel. The EETs may be synthesized during the development of cyclic flow variations and counteract the vasoconstrictor effects of thromboxane A2.

Problems with platelet inhibitors of thrombus formation in stenosed dog coronary arteries

Problems with platelet inhibitors of thrombus formation in stenosed dog coronary arteries

Pulmonic stenosis caused by single coronary artery in dogs: Four cases (1965-1984)

Summary Single right coronary artery (ca) associated with pulmonic stenosis was found in 3 English Bulldogs and a Boxer, suggesting a genetic predisposition for the associated anomalies. The left main coronary branch arose from the single right ca, encircled the pulmonic root over hypoplastic pulmonic valves at the level of the obstruction, and appeared to be the primary cause of underlying pulmonic stenosis. Patchgraft surgery to relieve pulmonic stenosis caused death in 1 dog when the unrecognized anomalous ca was severed during the procedure. The anomalous left ca was detectable by use of angiocardiography in all 4 dogs and was recognized before surgery in 2 of them. In 1 dog, a right ventricle-to-pulmonary trunk-bridging conduit was implanted to improve outflow from the right side of the heart.

P-475 - Responses to a K+ channel opener cromakalim of dog coronary and mesenteric arteries with different ATP-sensitive K+ channel populations

P-475 - Responses to a K+ channel opener cromakalim of dog coronary and mesenteric arteries with different ATP-sensitive K+ channel populations

The susceptibility of various cultured cells to induction of clear cytoplasmic vacuoles by disobutamide

Cultured cells were found to be highly useful for investigating intracellular storage of amphiphilic compounds using disobutamide as a model agent. To select types of cultured cells most suitable for investigations, cells of dog coronary artery muscle, rabbit aorta muscle, rat urinary bladder carcinoma, rat basophilic leukaemia, human skin fibroblasts, bovine aorta endothelium, Chinese hamster ovary tumour and mouse fibroblasts were incubated with 0, 1, 2, 4, 6, 8 and 10 × 10 −4 m-disobutamide for 24 hr. Cultures were examined in situ by phase light microscopy for the presence of clear cytoplasmic vacuoles, cell death (cell detachment), and for drug effect on confluency/cell count. Disobutamide induced vacuoles in all cell types except rat leukaemia. The drug induced cell death and reduction in confluency or cell count in cultures of all cell types except rat carcinoma and rabbit aorta muscle. Release of lactic dehydrogenase from cells confirmed the relative resistance of the rat carcinoma and rabbit cells, and susceptibility of rat leukaemia, to drug-induced cell death. By means of electron microscopy of rat carcinoma and rabbit cells, it was established that vacuoles were membrane-bound and their content was predominantly electron-lucent.

O-106 - Two opposing effects of pinacidil on ATP-sensitive and Ca2+-activated K+ channels in K+-contracted strips of dog coronary arteries

O-106 - Two opposing effects of pinacidil on ATP-sensitive and Ca2+-activated K+ channels in K+-contracted strips of dog coronary arteries

Inhibitory effects of L-N G-nitro-arginine on the synthesis of EDRF and the cerebroarterial response to vasodilator nerve stimulation

Treatment with L-N G-nitro-arginine (L-NA) inhibited the bradykinin-induced relaxation, mediated via EDRF, in dog coronary artery strips partially contracted with prostaglandin F 2α; the inhibition was prevented by L-, but not D-, arginine. Relaxation caused by nitrogylycerin was not altered by L-NA. The release of EDRF, as assayed using dog coronary artery strips without endothelium, from perfused femoral artery segments with endothelium in response to acetylcholine and substance P was significantly reduced by treatment of the femoral artery with L-NA. The inhibitory effect was reversed by L-arginine. Relaxant responses of dog cerebral artery strips with and without endothelium to electrical stimulation of non-adrenergic, non-cholinergic nerves were suppressed by L-NA, whereas relaxation of coronary arteries with and without endothelium by the stimulation of adrenergic nerves was not influenced. The L-NA-induced inhibition was reversed by L-arginine. It is speculated that L-NA inhibits the synthesis of EDRF, as does L-N G-monomethyl arginine, and NO-like substance(s) produced plays an important role in transferring information from vasodilator nerves to smooth muscle in cerebral arteries.

Outward potassium currents in freshly isolated smooth muscle cell of dog coronary arteries.

Outward membrane currents were characterized in single coronary smooth muscle cells of adult beagle dogs. The cells averaged 96.4 x 7.1 microns and had a resting potential of -50.7 mV, an input resistance of 307.9 M omega, a capacitance of 32.3 pF, and a calculated membrane surface area of 4,037 microns2. The cells contracted in response to external application of acetylcholine or high K+. In voltage clamp by use of the suction pipette method, outward current began to appear at -50 mV and reached 15.2 nA at 50 mV with a current density of 376.5 microA/cm2. The current was reduced by external tetraethylammonium, Ba2+, and internal Cs+, and its reversal potential had a Nernst relation to external K+ concentration. Elevation of external Ca2+ (Ca2+o) from 0 to 0.3 mM increased total K+ current by up to 300%; elevation of internal Ca2+ (Ca2+i) to 5 x 10(-7) M by internal perfusion increased total outward current to a similar extent, suggesting a large difference in Ca2+ transmembrane sensitivity. Total whole-cell K+ current consisted of two components: an initial time-independent current (Ii) followed by a time-dependent current (It). Ii and It were through separate K+ channels based on differences in a) sensitivity to Ca2+09b) modulation by an inward Ca2+ current, c) current amplitudes and activation kinetics, and d) responses to pharmacological agents. It was the largest component, measuring 4.5 nA in 0 mM Ca2+o but increasing to 11.9 nA in 0.3 mM Ca2+o with a steep 2.5 power function. It activated with a biexponential time course; in Ca2+o-free solution, its time course was relatively insensitive to voltage changes but became voltage sensitive in the presence of Ca2+o. Further, such sensitivity was abolished or enhanced by Co2+ or Bay K 8644, respectively. We concluded that there are two types of Ca2+-sensitive K+ currents, Ii and It, in coronary smooth muscle cells. Via an inward Ca2+ channel Ca2+o strongly modulates It, both in amplitude and kinetics.