Bovine Intrapulmonary Artery Research Articles

The shy Angeli and his elusive creature: the HNO route to vasodilation

what we now call Angeli's salt (AS) was discovered more than a century ago by the Italian chemist Angelo Angeli (Fig. 1) . He found that “salts of nitroxylaminic acid are readily resolved into the corresponding nitrites and the unsaturated residue nitroxyl” (HNO), proposing their use as suppliers of HNO (3). However, by analogy with the genial but very reclusive personality of the discoverer (17), the physiological properties of HNO didn't draw too much attention until early 1990s. In their seminal work, Fukuto and colleagues (19) reported that HNO elicits vasorelaxation in both the rabbit aorta and bovine intrapulmonary artery by a soluble guanylate cyclase (sGC)- dependent pathway. Later, it was evident that the thiol-donating agent l-cysteine can discriminate the vasodilatative profile of HNO from that of nitric oxide (NO·) or nitrosothiols: AS/HNO action can be blocked by this agent, whereas the NO· effect is potentiated (36). In combination with the suggestion that NO· is the only species able to directly activate sGC (9), these data led to the conclusion that HNO-induced vasorelaxation is due to its intracellular conversion to NO· with subsequent activation of sGC. For several years, this idea has relegated HNO to a mere precursor of the presence and biological activity of NO·, de facto negating to HNO its own right to exist.

Breaking bad news

The objective of the present study was to ascertain whether cyanide shares the properties of methylene blue as a selective inhibitor of vascular smooth muscle relaxation elicited by agents that stimulate the formation of cyclic GMP. Experiments were performed with endothelium-intact rings prepared from bovine intrapulmonary artery. Methylene blue, a good inhibitor of soluble guanylate cyclase, antagonized both arterial relaxation and cyclic GMP accumulation in response to sodium nitroprusside, glyceryl trinitrate, S-nitroso-N-acetylpenicillamine and acetylcholine. In contrast, cyanide inhibited only the responses to sodium nitroprusside. Increasing concentrations of methylene blue depressed resting arterial levels of cyclic GMP and caused slowly developing but marked contractions whereas cyanide was without effect. Contractile responses to phenylephrine, potassium and U46619 were potentiated by methylene blue but not by cyanide. Preincubation of dilute solutions of cyanide containing sodium nitroprusside in oxygenated Krebs9 buffer at 37 degrees C for 15 min before addition to bath chambers depressed relaxation and cyclic GMP accumulation caused by sodium nitroprusside markedly. Similar treatment of glyceryl trinitrate, however, failed to alter its effects in arterial rings. A chemical inactivation of sodium nitroprusside by cyanide appears to account for the specific inhibitory action of cyanide on arterial responses to sodium nitroprusside. This study indicates clearly that cyanide does not share the properties of methylene blue as an inhibitor of arterial relaxation elicited by vasodilators that stimulate cyclic GMP formation.

Erosion of inlet cannula of left ventricular assist device manifested as innocuous bleeding in stable patient: Lessons learned in prevention of catastrophic consequences

commonly used as a diagnostic agent for the determination of plasma volume. 5 The reason for inhibition of the sodium nitroprusside response by gentian violet is unclear. Gentian violet is metabolized to a reactive intermediate or free radical that could act at the vascular smooth muscle to inhibit the response. However, this is unlikely inasmuch as this mechanism would also interfere with the acetylcholinemediated vasodilation. A more plausible explanation would be the formation of a reactive intermediate that interacts directly with the sodium nitroprusside molecule. In conclusion, although methylene blue has been used as a marker dye, our studies suggest that it is inappropriate for marking vascular grafts. Evans blue did not adversely affect vasodilation to any agent and thus could be used as an alternative to methylene blue without fear of affecting graft reactivity. Gentian violet also appears to be a better alternate to methylene blue. However, we did observe a decrease in endothelium-independent vasodilation, a factor that should be considered when this agent is used. R E F E R E N C E S 1. Barber DA, Rubin JW, Zumbro GL, Tackett RL. The use of methylene blue as an extravascular surgical marker impairs vascular responses of human saphenous veins. J Thorac Cardiovasc Surg 1995;109:21-9. 2. Ignarro LJ, Burke TM, Wood KS, Wolin MS, Kadowitz PJ. Association between cyclic GMP accumulation and acetylcholine-elicited relaxation in bovine intrapulmonary artery. J Pharmacol Exp Ther 1984;228:682-90. 3. Marshall JJ, Wei EP, Lontos HA. Independent blockade of cerebral vasodilation from acetylcholine and nitric oxide. Am J Physiol 1988;255:H847-55. 4. Wakelin LPG, Adams A, Hunter C, Waring MJ. Interactions of crystal violet with nucleic acids. Biochemistry 1981;20:577987. 5. Windholz M, Budavari S, Blumetti RF, Otterbein ES, editors. Evans blue. In The Merck index, 10th ed. Rahway (NJ): Merck & Co, 1983:3855.

N ω-Hydroxy-L-arginine: A novel arginine analog capable of causing vasorelaxation in bovine intrapulmonary artery

This study describes the effects of N ω-hydroxy-L-arginine (NOHA) on endothelium dependent and endothelium independent relaxation of bovine pulmonary artery. These results are consistent with the hypothesis that NOHA is a biosynthetic intermediate in the production of nitric oxide from arginine. N ω-Hydroxy-L-arginine causes both endothelium dependent and endothelium independent vasorelaxation, similar to that of arginine. This NOHA elicited relaxation was also inhibitable by N-methylarginine, N-nitroarginine and N-aminoarginine.

Reduction of biological effluents in purge and trap micro reaction vessels and detection of endothelium-derived nitric oxide (edno) by chemiluminescence

Various analytical approaches have been used to measure endothelium-derived nitric oxide (NO) [ 1, 3, 4, 7, 10, 12]. We have detected NO in perfusates with a sample size as low as 2 ml after acidification with 4 N HCl to pH < 2 at 25°C by using a Nitric Oxide Analyser (Sievers, Colorado) [ 7]. This procedure had the advantage that the detectable level of NO was enhanced by the self-decomposition of HNO 2 when the PH < pK a of NHO 2 (pKa = 3.15) and also the reaction temperature of 25°C substantially increased the half-line of NO [ 2]. Palmer, et al., [ 10] measured NO released by cultured porcine endothelial cells by chemiluminescence after passing cell effluents continyously at a rate of 5 ml/min into 75 ml of 1% sodium iodide in glacial acetic acid. The larger volumes involved in this method for continuous refluxing, made it less desirable for the detection of endothelium-derived nitric oxide. Feelisch et al. [ 1] utilized the activation of soluble guanylate cyclase, as well as, the quantitative oxidation of oxyhemoglobin to methemoglobin in aqueous solutions by NO as a means of measuring nitric oxide. We describe here a modification of our earlier micromethod which now enables us to detect NO after complete reduction with glacial acetic acid and sodium iodide. A comparison of the two procedures indicate that while freshly prepared NO standard solutions gave identical chemiluminescence response with and without reduction, effluents from bovine intrapulmonary artery under basal conditions gave substantially higher values upon reduction.

A model to study physiological activation of phospholipase A 2 and vasorelaxation by lysophosphatidycholine

Earlier we demonstrated that micellar solutions of LPC caused endothelium-dependent relaxation of rabbit thoracic aorta and bovine intrapulmonary artery and vein through a cyclic GMP-dependent mechanism. The availability of LPC for vasorelaxation depends on its production by deacylation of PC by PLA 2. We assessed the possible activation of PLA 2 by commonly used vasorelaxants such as acetylcholine, bradykinin, calcium ionophore A23187 and thrombin and vasoconstrictors like histamine and phenylephrine in the presence of indomethacin in a model system where 14C PC was incorporated into bovine intrapulmonary arterial segments. Taking the ratio of 14C PC:LPC formed by exogenous PLA 2 as an index of deacylation, we found that while all the agents relaxed the strips in an endothelium-dependent manner, only thrombin caused relaxation followed by an increase in 14C LPC and a concomittant decrease in 14C PC indicating activation of PLA 2. Our data show that PC/PLA 2 system can be activated to generate LPC for vascular relaxation under specific physiological conditions. This model system can be used to monitor PLA 2 activity and LPC production to compensate flow and pressure induced changes in arteries.

Alteration of cyclic GMP metabolism by CD-349, a novel calcium antagonist, and by sodium nitroprusside in bovine intrapulmonary artery and vein

The effects of 2-nitratopropyl 3-nitratopropyl 2,6-dimethyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate (CD-349) and sodium nitroprusside (NP) on cyclic GMP (cGMP) metabolism in bovine intrapulmonary artery (BPA) and vein (BPV) were examined. CD-349 inhibited cGMP phosphodiesterase (PDE) activity in BPA and BPV. In the latter, about 40% of the cGMP PDE activity was Ca 2+ dependent. The inhibition of cGMP PDE activity by CD-349 also depended on Ca 2+. The inhibitory effect of CD-349 was more potent than that of nicardipine or nifedipine. The conversion of cGMP from GTP in the homogenates of BPA and BPV was stimulated by NP in a concentration-dependent manner. The NP-induced cGMP formation was stimulated further by CD-349. This effect of CD-349 depended on Ca 2+ in the BPV but not in the BPA. The NP-induced elevation of cGMP levels in the tissue preparations of BPA and BPV was also potentiated by CD-349. These results suggest that CD-349 inhibited Ca 2+-dependent cGMP PDE activity and that the levels of cGMP were elevated in vascular smooth muscle, particularly when guanylate cyclase was activated.

Basic polyamino acids rich in arginine, lysine, or ornithine cause both enhancement of and refractoriness to formation of endothelium-derived nitric oxide in pulmonary artery and vein.

The objective of this study was to elucidate the mechanism by which polyamino acids containing L-arginine, L-lysine or L-ornithine cause endothelium-dependent relaxation of bovine intrapulmonary artery and vein. Basic but not acidic or neutral polypeptides ranging in average molecular weights from 17 to 225 kDa elicited time- and concentration-dependent relaxation and cyclic GMP accumulation in precontracted rings of artery and vein by endothelium-dependent mechanisms. Vascular responses were markedly inhibited by oxyhemoglobin, methylene blue, or potassium. The basic polyamino acids stimulated the formation and/or release of an endothelium-derived relaxing factor (EDRF) identified as nitric oxide (NO) in perfused segments of both artery and vein as assessed by bioassay. The polyamino acids and A23187 released a similar endothelium-derived NO (EDNO) from artery and vein, as assessed by the similar half-life (3-5 seconds), antagonism by superoxide anion or oxyhemoglobin, enhancement by superoxide dismutase, and lack of influence by indomethacin. The basic polyamino acids elicited potent relaxant responses with EC50 values ranging from 3 x 10(-9) to 2 x 10(-7) M, and a direct correlation was obtained between molecular weight and relaxation potency irrespective of the basic amino acid incorporated. Prolonged contact of arterial or venous rings with basic polyamino acids resulted in the rapid development of marked refractoriness to relaxation and cyclic GMP formation on addition of polyamino acid. Moreover, refractoriness developed to the vascular responses of other endothelium-dependent vasodilators but not to glyceryl trinitrate or isoproterenol. The mechanism of refractory responses was attributed to interference with EDNO formation and release as assessed by bioassay and chemical assay. The hypothesis is forwarded that the basic polyamino acids serve as partial substrates for the enzyme system that catalyzes the conversion of L-arginine to NO. Prolonged contact between substrate and enzyme results in enzyme desensitization and the development of refractoriness or a form of tolerance to vasodilators whose action is mediated by EDNO.

Uptake of CD-349 and nitrendipine into bovine intrapulmonary artery and vein.

The uptake of CD-349 into the bovine intrapulmonary artery (BPA) and vein (BPV) was examined and characteristics of this newly synthesized 1,4-dihydropyridine Ca2+ antagonist were compared with those of nitrendipine. One hundred minutes after initiation of incubation of the vessels, the uptake of [3H]CD-349 and [3H]nitrendipine reached a plateau. The concentrations of CD-349 inside the muscle cell of the BPA and BPV were 11.1 and 6.8 times higher than that of the incubation medium, respectively, and concentrations of nitrendipine in the BPA and BPV were 8.6 and 6.1 times higher, respectively. The degree of uptake was not influenced by the lowering the incubation temperature nor by adding 2,4-dinitrophenol. The partition coefficient of CD-349 measured in the corn oil/Ringer and octanol/Ringer systems was greater than the partition coefficient of nitrendipine, independent of the concentration tested. These results suggest that CD-349 permeates smooth muscle membrane by passive diffusion and accumulates inside the muscle cells, much in the same manner as seen with nitrendipine. The lipid solubility of DC-349 presumably accounts for these events.

Endothelium-dependent modulation of angiotensin II-induced contraction in blood vessels

The influence of endothelium on angiotensin II-induced contraction was investigated in rings of rat aorta, bovine coronary artery, bovine intrapulmonary artery and bovine intrapulmonary vein. Destruction of endothelium significantly enhanced angiotensin II-induced contraction in rat aorta and bovine coronary artery, but not in bovine intrapulmonary artery and bovine intrapulmonary vein. Indomethacin (10 −5 M) did not alter angiotensin II-induced contraction in rat aorta or bovine coronary artery. However, hemoglabin (10 −5 M) or methylene blue (10 −5 M) significantly enhanced angiotensin II-induced contraction in rat aorta and bovine coronary artery with, but not without, endothelium. Intimal rubbing did not affect stimulation of phosphoinositide hydrolysis by angiotensin II in rat aorta. The findings demonstrate that angiotensin II-induced contraction in vascular rings can be modulated by endothelium. However, the effect of endothelium apparently depends upon the species and vascular bed from which the vessel is isolated. Results obtained using inhibitors suggest that in rat aorta and bovine coronary artery release of endothelium-derived relaxant factor (EDRF), rather than cyclooxygenase products, is involved in mediating the inhibitory influence of endothelium. Further, similar stimulation of phosphoinositide hydrolysis in intimally rubbed and unrubbed rat aorta suggests that EDRF does not modulate angiotensin II-induced contraction in this vessel by inhibiting angiotensin II stimulation of phosphoinositide hydrolysis.

P-480 - Effects of CD-349, a new calcium antagonist, and sodium nitroprusside on cyclic GMP metabolism in the bovine intrapulmonary artery and vein

P-480 - Effects of CD-349, a new calcium antagonist, and sodium nitroprusside on cyclic GMP metabolism in the bovine intrapulmonary artery and vein

Endothelium-derived relaxing factor produced and released from artery and vein is nitric oxide.

The objective of this study was to determine whether nitric oxide (NO) is responsible for the vascular smooth muscle relaxation elicited by endothelium-derived relaxing factor (EDRF). EDRF is an unstable humoral substance released from artery and vein that mediates the action of endothelium-dependent vasodilators. NO is an unstable endothelium-independent vasodilator that is released from vasodilator drugs such as nitroprusside and glyceryl trinitrate. We have repeatedly observed that the actions of NO on vascular smooth muscle closely resemble those of EDRF. In the present study the vascular effects of EDRF released from perfused bovine intrapulmonary artery and vein were compared with the effects of NO delivered by superfusion over endothelium-denuded arterial and venous strips arranged in a cascade. EDRF was indistinguishable from NO in that both were labile (t1/2 = 3-5 sec), inactivated by pyrogallol or superoxide anion, stabilized by superoxide dismutase, and inhibited by oxyhemoglobin or potassium. Both EDRF and NO produced comparable increases in cyclic GMP accumulation in artery and vein, and this cyclic GMP accumulation was inhibited by pyrogallol, oxyhemoglobin, potassium, and methylene blue. EDRF was identified chemically as NO, or a labile nitroso species, by two procedures. First, like NO, EDRF released from freshly isolated aortic endothelial cells reacted with hemoglobin to yield nitrosylhemoglobin. Second, EDRF and NO each similarly promoted the diazotization of sulfanilic acid and yielded the same reaction product after coupling with N-(1-naphthyl)-ethylenediamine. Thus, EDRF released from artery and vein possesses identical biological and chemical properties as NO.

Mechanisms of endothelium-dependent vascular smooth muscle relaxation elicited by bradykinin and VIP.

The objective of this study was to elucidate the mechanisms by which bradykinin and vasoactive intestinal polypeptide (VIP) relax bovine intrapulmonary artery and bradykinin, but not VIP, relaxes intrapulmonary vein. Bradykinin and VIP elicited entirely endothelium-dependent relaxation of phenylephrine-precontracted arterial rings, and this was associated with arterial accumulation of both guanosine 3',5'-cyclic monophosphate (cGMP) and adenosine 3',5'-cyclic monophosphate (cAMP). Bradykinin, but not VIP, relaxed precontracted venous rings and increased cGMP, but not cAMP levels, by endothelium-dependent mechanisms. Neither arteries nor veins relaxed in response to substance P, thrombin, bombesin, arginine vasopressin, or angiotensin II. Methylene blue or indomethacin each partially antagonized, whereas both, when together, abolished arterial relaxant responses to bradykinin and VIP. Methylene blue or indomethacin, respectively, abolished arterial cGMP or cAMP accumulation elicited by bradykinin and VIP. Venous relaxation and cGMP accumulation elicited by bradykinin was abolished by methylene blue but was unaltered by indomethacin. Thus bradykinin and VIP relaxed bovine intrapulmonary artery by endothelium-dependent mechanisms involving the actions of cGMP and cAMP whose formation may be stimulated by endothelium-derived relaxing factor and prostacyclin, respectively. In contrast, bradykinin relaxed intrapulmonary vein by endothelium-dependent mechanisms involving only cGMP.

Endothelium-dependent modulation of cGMP levels and intrinsic smooth muscle tone in isolated bovine intrapulmonary artery and vein.

The role of the endothelium in modulating cyclic nucleotide levels and intrinsic smooth muscle tone was studied in isolated rings of bovine intrapulmonary artery and vein. Cyclic 3',5'-guanosine monophosphate (cGMP) levels were threefold to fourfold higher in unrubbed artery and vein than in vessels that had been denuded of endothelium. Cyclic 3',5'-adenosine monophosphate (cAMP) levels were twofold higher in unrubbed than in endothelium-denuded artery, but no differences were observed in veins. Methylene blue, an inhibitor of guanylate cyclase, decreased cGMP but not cAMP levels, and this was accompanied by increases in smooth muscle tone. M&B 22,948, an inhibitor of cGMP-phosphodiesterase, increased cGMP but not cAMP levels, and this was accompanied by decreases in smooth muscle tone. Unrubbed vessels were more sensitive than endothelium-denuded vessels to the actions of both methylene blue and M&B 22,948, and this may be attributed to endothelium-dependent increases in cGMP turnover. Moreover, unrubbed vessels were more sensitive than endothelium-denuded vessels to contractile responses to phenylephrine and potassium, and these responses were potentiated by methylene blue and attenuated by M&B 22,948. Although indomethacin lowered cAMP levels in unrubbed artery, no changes in tone or contractile responsiveness were observed. A consistent observation was that the smaller branches of unrubbed but not endothelium-denuded intrapulmonary artery and vein had higher levels of cGMP but not cAMP, were sensitive to endothelium-dependent vasodilators, were more sensitive to methylene blue, and would not maintain a steady level of submaximal tone to phenylephrine when compared with larger branches from a common vascular bed.(ABSTRACT TRUNCATED AT 250 WORDS)

Bradykinin-induced endothelium-dependent relaxation of bovine intrapulmonary artery and vein

Bradykinin induced relaxation and cyclic GMP accumulation in both bovine intrapulmonary artery and vein. Both the relaxant responses and the accompanying cyclic GMP accumulations were abolished or markedly reduced by intimal rubbing or pretreatment with the guanylate cyclase inhibitor, methylene blue. These findings indicate that both bovine intrapulmonary artery and vein exhibit endothelium-dependent relaxation in response to bradykinin, and that the relaxant responses in both vessels are associated with cyclic GMP accumulation.

Atriopeptin II relaxes and elevates cGMP in bovine pulmonary artery but not vein.

Atriopeptin II, a 23-amino acid synthetic peptide fragment of atrial natriuretic factor, caused an endothelium-independent relaxation of isolated precontracted rings of bovine intrapulmonary artery that was accompanied by the concomitant accumulation of guanosine 3',5'-cyclic monophosphate (cGMP) but not adenosine 3',5'-cyclic monophosphate. In contrast, rings of intrapulmonary vein were unaffected by atriopeptin II whether or not endothelium was present. Whereas methylene blue, an inhibitor of soluble guanylate cyclase, abolishes endothelium-dependent and independent arterial relaxation and cGMP accumulation in response to acetylcholine and glyceryl trinitrate, respectively, methylene blue failed to alter these responses to atriopeptin II. Similarly, the effects of atriopeptin II were unaltered by propranolol, indomethacin, or atropine. These results indicate that relaxation elicited by atriopeptin II may be selective for arterial smooth muscle receptors, does not require endothelial cells, and does not involve the soluble form of guanylate cyclase, although cGMP accumulation is stimulated.

Parenchymal and vascular strips from bovine lung respond differently to vasodilators

Strips of bovine intrapulmonary artery (IPA), intrapulmonary vein (IPV), trachealis muscle and parenchyma were mounted for recording of isometric tension and responses to several vascular smooth muscle relaxants were measured and compared. Although nitroglycerin (GTN) and sodium nitroprusside (SNP) induced concentration-related (10 −11 to 10 −5) relaxation (up to 80–100%) in IPA, IPV and trachealis muscle, 10 −5 M GTN or SNP induced only 4–5% relaxation in parenchyma. In contrast, all four tissues relaxed well in response to isoproterenol (10 −11 to 10 −5 M), forskolin (10 −5 M), 8-bromo-cyclic GMP (10 −4 M) and 1-methyl-3-isobutylxanthine (10 −5 M). When compared to IPA, IPV and trachealis muscle, the marked hyporesponsiveness of parenchyma to GTN and SNP suggests that the contractile cells which are primarily responsible for potassium-induced tone in bovine parenchymal strips differ markedly from smooth muscle cells in trachealis muscle and large, central IPA and IPV in their responsiveness to the vasodilators, GTN and SNP.

Contractile effects of 5-hydroxytryptamine in isolated intrapulmonary arteries and veins.

The contractile effects of serotonin (5-hydroxytryptamine, 5-HT) were compared in helical strips of intrapulmonary artery (IPA) and vein (IPV) isolated from lungs of dog, rabbit, cow, and man. 5-HT (10(-8) - 10(-4) M) increased force generation by IPA and IPV from the four species in a concentration-related manner. Sensitivities to 5-HT of IPA and IPV from the four species were similar, with median effective concentrations ranging from 3 X 10(-8) to 3 X 10(-7) M. However, maximal and contractile responses to 5-HT relative to those elicited by potassium were greater in canine and human IPA and bovine IPA and IPV than in rabbit IPA and IPV and canine and human IPV. Results obtained using specific pharmacology blocking agents suggest that 5-HT-induced contraction of canine and rabbit intrapulmonary vessels does not involve alpha-adrenergic mechanisms except, possibly, in rabbit IPV. The contractile effects of 5-HT observed in the present study suggest that 5-HT released from platelets could contribute to increased vascular resistance during pulmonary thromboembolism by actively constricting arterial and venous segments of the pulmonary vascular bed in a variety of species, including man.