Regulation of Nitric Oxide Production in Human Coronary Microvessels and the Contribution of Local Kinin Formation
Background The goal of this study was to define the regulation of nitric oxide release by coronary microvessels from the failing and nonfailing human heart and to determine the role of local kinin production in the elaboration of nitric oxide by human coronary microvascular endothelium.
Methods and Results Ten hearts from humans with end-stage heart failure and two hearts from patients without heart failure were harvested at the time of orthotopic cardiac transplantation. Microvessels were sieved and the production of nitrite was determined by the Griess reaction. Microvessels were incubated in the presence of agonists for nitric oxide production (acetylcholine and bradykinin), which caused dose-dependent increases in nitrite, a response that was blocked by NG-nitro-l-arginine methyl ester and receptor-specific antagonists (atropine and HOE 140, respectively). In addition, the production of nitrite by microvessels from the failing heart appeared to be less than that produced by microvessels from the nonfailing heart. Incubation with norepinephrine or the α2-adrenergic agonist BHT 920 also caused dose-dependent increases in nitrite production, which were blocked by the B2-receptor antagonist HOE 140. This implicated local kinin synthesis as an intermediate step in the production of nitric oxide in response to α2-adrenoceptor stimulation. The production of nitric oxide was also prevented by the addition of serine protease inhibitors, which blocked the action of local kallikrein, again suggesting a role for local kinin synthesis.
Conclusions Our results indicate that nitric oxide is produced by human coronary microvessels, that nitric oxide production may be reduced but certainly not increased in microvessels from the failing human heart, and that there is active local kinin generation in these blood vessels.
The coronary microcirculation is a dynamic vascular bed that responds through endothelium-dependent and -independent mechanisms to metabolic tissue demands, changes in blood flow, circulating humoral factors, and direct neuronal activation.1 2 The vascular endothelium produces vasodilators such as NO and prostanoids, the synthesis of which may be defective in disease states such as heart failure. For example, we3 recently showed that endothelium-mediated control of the canine coronary circulation is impaired in dogs with pacing-induced dilated cardiomyopathy and results in diminished vasodilator responses to coronary occlusion and ACH. Studies4 have shown abnormalities in endothelium-dependent dilation of the coronary microvasculature in patients with dilated cardiomyopathy who were administered ACH but a near-normal response to administration of the endothelium-independent vasodilator adenosine. Peripheral endothelial cell dysfunction, as exhibited by a blunted vasodilator response to ACH, of hindquarter resistance vessels has been reported as a consequence of an experimental form of heart failure in rats,5 and similarly, in forearm resistance vessels of humans with dilated cardiomyopathy.6
α2-Adrenoceptors, which are present on intact endothelial cells, have been shown to stimulate the production of EDRF or NO,7 8 9 10 perhaps to modulate the constriction caused by stimulation of these receptors. α-Adrenoceptors are present in the coronary microcirculation, especially on endothelial cells, and in cat and dog heart, it appears that α2-adrenoceptors are located preferentially in the small coronary arterioles, those <100 μm in diameter.11 Although α-adrenoceptors are present in the human coronary circulation,9 α-adrenergic stimulation of NO release has not been fully described in humans, nor has the mechanism through which NO release occurs been fully elucidated.
The purpose of the current study was to use direct measurement of the hydration product of NO, nitrite, in vitro to determine whether human coronary microvessels produce NO; to compare the regulation of NO release by the coronary microcirculation in the failing and nonfailing human heart; and to determine the role of local kinin production in the elaboration of NO by the coronary microvascular endothelium. A preliminary report was presented previously.12
Procurement of Human Hearts
Explanted hearts were harvested from 12 patients at the time of orthotopic cardiac transplantation and immediately placed in iced normal saline in the operating room. The apical portion of each heart was removed for study; this included the left ventricular free wall, the septum, and a portion of the right ventricular free wall. In addition, hemodynamic information was collected on all patients. Ten patients had severe, end-stage cardiac failure at the time of cardiac transplantation. Two patients had intact ventricular function and underwent transplantation for reasons other than ventricular failure.
Preparation of Coronary Microvessels
Hearts were transported on ice to the laboratory, where microvessels were isolated by use of the method of Gerritsen and Printz13 as described by us recently.3 14 15 The hearts were received in the laboratory in less than 30 minutes in all but one instance; one heart was received 45 minutes after removal. Each heart was placed in iced PBS that contained 1% bovine serum albumin, pH 7.40. Left ventricular free-wall, midmyocardial tissue was dissected free of epicardium, endocardium, large coronary arteries, and fat. The remaining myocardium was cut with scissors into small pieces and suspended in ice-cold PBS. From this sample, microvessels were collected by use of sequential sieving and glass-bead separations as described previously.3 13 14 15 16 This yielded a preparation that was virtually free of contaminating myocytes and that consisted only of arterioles, venules, and capillaries <80 μm in diameter; approximately 850 mg of microvessels was collected per heart.
Microvessels were placed in a small packet made of 80-μm nylon mesh and placed in a constant-temperature (37°C) tissue bath that contained PBS. A gas mixture of 95% oxygen and 5% carbon dioxide was bubbled through the bath for 30 minutes. Twenty-milligram aliquots of tissue were placed in plastic tubes that contained 500 mL of PBS alone or 450 mL of PBS with the addition of 50 mL of drug to stimulate or inhibit NO production and were incubated in the bath for 20 minutes. All nitrite production is presented as pmol/mg wet weight per 20 minutes.
Measurement of Nitrite Release
Nitrite (NO2) is the hydration product of NO. With use of the Griess reaction, sulfanilic acid is diazotized by NO, and the resultant pink color is proportional to the amount of nitrite (hence, NO) present. Sulfanilamide (450 mL of 0.1% solution) and N-(1-naphthyl)ethylenediamine (50 mL of 0.2%) were added to each tube of microvessels after incubation. After a 10-minute period at room temperature, the supernatant was removed from each tube and the optical density measured at a wavelength of 540-μm absorbance (Uvikon 930 spectrophotometer, Kontron Instruments). Standard curves were generated each day by the addition of known amounts of nitrite to buffer.
NO/Nitrite Release From Human Coronary Microvessels
Microvessels from all hearts were incubated with an antagonist for 10 minutes before the addition of the maximal stimulatory dose of agonist used. Because of the limited amount of tissue obtained from each heart, complete dose-response curves to agonist with antagonist could not be generated, so that only the highest dose of each agonist was studied in the presence of an antagonist. We also specifically examined the potential role of local kinin formation in NO production. Microvessels were incubated with stimulators of local kinin production, and preincubation of microvessels with several inhibitors of enzymes that could generate kinins (including aprotinin, soybean trypsin inhibitor, and dichloroisocoumarin17 ) was performed to determine the potential role of local kinin formation in the production of NO.
Agonists used to stimulate NO production included ACH (10−8 to 10−5 mol/L) and BK (10−8 to 10−5 mol/L). The highest dose of BK was also studied after preincubation with the bradykinin-2 (B2) receptor blocker HOE 140 (10 μmol/L). Likewise, the highest dose of ACH was studied after preincubation with the muscarinic receptor–antagonist atropine methyl bromide (10 μmol/L) or HOE 140. To ensure that the measured nitrite truly reflected NO formation, preincubation with the NO synthase inhibitor L-NAME (100 μmol/L) was performed before the addition of the maximal dose of each agonist.
Contribution of Local Kinin Production in Human Coronary Microvessels
To test the response to α-adrenergic stimulation, microvessels were incubated with norepinephrine, the nonselective adrenergic agonist, in increasing doses (10−8 to 10−5 mol/L) alone, with L-NAME, or with the addition of the B2 receptor antagonist HOE 140 (10 μmol/L). Increasing doses of the selective α2-adrenergic receptor agonist BHT 920 (10−8 to 10−5 mol/L) were also used alone and with the addition of HOE 140 (10 μmol/L). To determine the role of local kinin synthesis as a mediator of the release of NO in response to α2-adrenergic receptor stimulation, serine protease inhibitors were used to block kallikrein-mediated local kinin formation. Microvessels were preincubated with soybean trypsin inhibitor (10 μmol/L), dichloroisocoumarin (10 μmol/L), or aprotinin (10 μmol/L); the maximal dose of norepinephrine (10−5 mol/L) or BHT 920 (10−5 mol/L) was then added. At the end of each incubation, the supernatant was separated from the microvessels and nitrite production was determined.
Potential for NO Production by Myocytes
As determined by phase-contrast microscopy, the eluted tissue obtained from human hearts during the last sieving step primarily contained myocytes. In two human heart preparations, these myocytes were collected, centrifuged, and incubated with the maximal stimulatory dose of BK or ACH, and nitrite was measured.
Differences from control for each dose of agonist were determined by use of a Student’s t test and ANOVA. Differences between nitrite production of microvessels from normal and failing canine hearts were determined by unpaired t test, and a Bonferroni correction was used when multiple comparisons were performed. Standard curves for nitrite production as determined by spectrophotometry were constructed each day; best fit was determined by use of a linear least-squares analysis. Only standard curves with r>.90 were used. We calculated the amount of nitrite by substituting the measured absorbance of each sample into the equation of the line. All of these techniques have been used by us previously.3 14 15 18
Pertinent clinical and hemodynamic data are listed in the Table⇓. Of the 12 patients from whom hearts were harvested, 10 were male and 2 were female. The age range was 6.5 months to 63 years, with a median age of 12 years. The most common clinical diagnosis was idiopathic dilated cardiomyopathy (6 of 12 patients). Ten hearts were designated as failing (patients F-1 to F-10 in the Table⇓); these were obtained from patients whose main indication for transplantation was low cardiac output and failure of the left ventricle. Two hearts were nonfailing (patients NF-1 and NF-2 in the Table⇓); 1 of these was obtained from a patient who was transplanted for cyanotic congenital heart disease not amenable to conventional surgery and the other from a patient who underwent retransplantation because of accelerated coronary vasculopathy of his first cardiac allograft. All patients were considered NYHA class 4. Seven of the 10 patients with heart failure required intravenous inotropic support before transplantation.
On pathological examination, all hearts exhibited some degree of myocardial hypertrophy. This was quantified as the percent above estimated normal heart weight for age and sex, which was more useful than absolute weight in grams because the population from which the explanted hearts were obtained included both pediatric and adult patients of both sexes. The mean percent above normal heart weight was 190±20% (mean±SE).
NO/Nitrite Release From Human Coronary Microvessels
Increasing doses of BK or ACH caused dose-related increases in nitrite production in all human hearts. The response to BK by microvessels isolated from all failing and nonfailing human hearts is shown in Fig 1⇓. The maximal dose of BK (10−5 mol/L) yielded a mean percent change from control of 157±28% (n=10) in microvessels from failing hearts. The magnitude of the response to ACH was similar.
The maximal dose of BK was also studied after preincubation with HOE 140 (10 μmol/L; n=12), which abolished any significant increase in nitrite production. Likewise, the maximal dose of ACH was studied after preincubation with atropine or HOE 140. Atropine blocked the increases in nitrite production in response to ACH (n=7; P<.05), whereas nitrite production was unaffected by HOE 140 (data not shown). Preincubation of microvessels with the NO synthase inhibitor L-NAME before the addition of the maximal dose of each agonist significantly diminished the response to either BK or ACH, as shown in Fig 2⇓.
Contribution of Local Kinin Production in Human Coronary Microvessels
Nonspecific α-adrenergic stimulation with norepinephrine or α2-adrenergic receptor selective stimulation with BHT 920 caused dose-dependent increases in nitrite release from human coronary microvessels. Results from studies in microvessels from failing and nonfailing human hearts are shown in Fig 1⇑.
The addition of the B2-receptor blocker HOE 140 to the maximal dose of each agonist eliminated the increases in nitrite production (n=12). Incubation with L-NAME before the addition of agonist had a similar effect. Data are shown in Fig 3⇓.
Preincubation of microvessels isolated from all human hearts with the serine protease inhibitors soybean trypsin inhibitor (10 μmol/L), dichloroisocoumarin (10 μmol/L), or aprotinin (10 μmol/L) was performed to block the action of kallikrein. The maximal dose of norepinephrine (10−5 mol/L) or BHT 920 (10−5 mol/L) was then added. No significant difference was observed in nitrite production by microvessels compared with basal levels. These data are shown in Fig 4⇓. Preincubation with the serine protease inhibitors had no effect on the stimulatory action of the maximal dose of ACH or BK.
Potential for NO Production by Myocytes
Human myocytes, collected from the final elution step in microvessel preparations, were centrifuged and incubated with the maximal stimulatory dose of BK or ACH. None of the tissue that was tested generated measurable amounts of nitrite in the basal state or in the presence of these agonists.
There were several significant findings of this study. First, BK, ACH, norepinephrine, and BHT 920 caused dose-dependent increases in nitrite release from coronary microvessels isolated from failing and nonfailing human hearts. When these two groups of hearts were compared, we found that nitrite release by failing human hearts appeared to be less than the nitrite release measured in nonfailing human hearts. However, it was not possible to procure a sufficient number of nonfailing human hearts for statistical comparisons. Our studies with human microvessels indicated that nitrite release in response to all agents that stimulated nitrite production was markedly reduced by L-NAME, an inhibitor of NO synthase, which confirmed that nitrite production was an indication of NO metabolism. As expected, HOE 140 specifically inhibited nitrite release in response to BK; similarly, atropine selectively blocked the effects of ACH. Surprisingly and perhaps most interestingly, nitrite release in response to α2-adrenergic receptor stimulation with norepinephrine or BHT 920 was inhibited by the B2-receptor antagonist HOE 140. To test the involvement of local kinin production in this process, the serine protease inhibitors soybean trypsin inhibitor, aprotinin, or dichloroisocoumarin were used to block the action of local kallikrein. All three agents caused significant decreases in nitrite release during α2-receptor stimulation.
The patient population from which the explanted hearts were obtained included 10 patients with symptomatic severe congestive heart failure and hemodynamics consistent with ventricular dysfunction and 2 patients with no symptoms of heart failure and hemodynamics that reflected preserved ventricular function. The population included both adults and children. It is evident that the first 10 patients were “failing,” as evidenced by elevated pulmonary capillary wedge pressures, low mixed venous oxygen saturations, low cardiac indexes, and high resting heart rates. This observation was supported by measures of ventricular function, which were obtained routinely in adults as left ventricular ejection fraction by multigated acquisition scan and in children as percent fractional shortening of the left ventricle from transthoracic M-mode echocardiogram. In the 10 patients with failing hearts, these values were well below normal. Of the 2 patients with nonfailing hearts, 1 (designated as NF-1 in the Table⇑) had congenital heart disease and persistent cyanosis but low pulmonary capillary wedge pressure and qualitatively normal ventricular function by transthoracic M-mode echocardiogram. The other patient (designated as NF-2 in the Table⇑) underwent retransplantation for posttransplant coronary vasculopathy; this patient had normal hemodynamics and qualitatively normal ventricular function by echocardiogram. Although we could not designate these patients as “failing,” we also could not call these patients truly “normal,” given their anatomic or pathological indications for cardiac transplantation.
The patients with symptomatic heart failure were all treated with an oral regimen that included digoxin and diuretics. Six of the 10 patients were treated with ACE inhibitors. Seven of the 10 patients were treated chronically in the hospital with intravenous dobutamine until the time of transplantation. There is some question as to the effect of these medications on endogenous NO production. Dobutamine primarily stimulates β1-adrenergic receptors, and therapy before transplantation and isolation of coronary microvessels appears to have little or no effect on NO/nitrite production in vitro, ie, this did not cause increased nitrite production in the basal or stimulated state in microvessels from failing hearts compared with normal. Of note, direct application of specific β-adrenergic receptor agonists has been shown to have no effect on stimulation of EDRF production or release in isolated canine coronary artery rings.19 The vasodilator properties of ACE inhibitors and their modulation of the vascular response to BK have been well described.20 21 22 Although it is possible that ACE-inhibitor therapy may alter nitrite production in isolated coronary microvessels, we have not found this to be true.
Our data support work that previously demonstrated that endothelium-dependent dilation is abnormal in congestive heart failure and that this impairment extends to the coronary microcirculation. In studies in humans, Drexler et al6 demonstrated the consequences of impaired NO production by showing that patients with congestive heart failure exhibit a blunted increase in forearm blood flow to endothelium-dependent dilating agents, such as ACH, compared with healthy patients. Those data also suggest that the basal release of NO is preserved or may be enhanced in forearm resistance vessels in patients with heart failure, which implies a compensatory role in the modulation of tissue perfusion in response to humoral agents after the development of heart failure. The probability that intrinsic microvascular disease exists in the coronary circulation has been explored by Treasure et al,4 who demonstrated a differential response in small-vessel resistance to endothelium-dependent (ACH) and -independent (adenosine) dilators in normal patients and those with cardiomyopathy. Although coronary blood flow was increased in both groups with adenosine, ACH caused significant increases in coronary blood flow in the control group only; blood flow was not altered in patients with cardiomyopathy.
Data from our laboratory3 showed attenuated dilation of the large coronary artery and increases in coronary blood flow in conscious dogs with pacing-induced cardiomyopathy after brief occlusion of the left circumflex coronary artery, intravenous administration of ACH, or administration of arachidonic acid. The diminished response to these vasodilators suggests a defect in endothelial function of both the large coronary arteries and the coronary microvasculature of failing hearts. Recently, in our laboratory, Zhao et al18 showed that reflex cholinergic coronary vasodilation is almost abolished during pacing-induced heart failure in the dog because of the disappearance of NO. Both isolated large coronary arteries and microvessels of failing hearts produced significantly less nitrite in vitro in response to all agonists in our previous studies,3 16 and this effect was blocked by the addition of L-NAME. These data support the contention that endothelium-mediated control of the coronary circulation may be depressed in the failing heart.
It has also been postulated that NO modulates α-adrenergic vasoconstriction in a number of blood vessels. Jones et al7 showed that α-adrenergic receptors are present throughout the canine coronary microcirculation, with α1-receptors predominating in small arteries and α2-receptors in arterioles. When norepinephrine, β-adrenergic receptor blockade, and selective inhibition of either α1-receptors with prazosin or α2-receptors with rauwolscine were used, constriction caused by α1- or α2-adrenergic receptor activation was potentiated during inhibition of NO synthase.
In comparing the response to norepinephrine of carotid, mesenteric, renal, and femoral large arteries of the pig, greyhound, and mongrel dog, Angus et al8 have shown that there is a nonuniformity of distribution of α2-adrenoceptors on endothelium and smooth muscle. They also demonstrated that α2-receptors on endothelium mediate the release of EDRF, as evidenced by an enhanced concentration-contraction curve to norepinephrine in vessels denuded of endothelium. More specifically, norepinephrine relaxed precontracted arteries in the presence of β-adrenergic receptor blockade, and this was inhibited by α2-adrenoceptor antagonists.8
Studies in humans conducted by Indolfi et al9 showed that in vivo administration of the selective α2-receptor agonist BHT 933 to patients with normal coronary arteries induced a reduction in coronary artery diameter and coronary blood flow velocity. α2-Adrenergic blockade did not change coronary blood flow in normal subjects; however, in patients with atherosclerosis, regional coronary blood flow decreased after receptor blockade. This supports our data as well in that α2-receptors participate in the modulation of vascular resistance in the human heart.
It has not been shown before in blood vessels from humans, however, that the stimulatory effects of α2-adrenergic receptor activation on NO release occur via activation of local kinin synthesis. In the present study, nonselective α-stimulation with norepinephrine and the selective α2-adrenergic stimulator BHT 920 caused dose-dependent increases in nitrite release from coronary microvessels isolated from both failing and nonfailing human hearts. The addition of the B2-receptor antagonist HOE 140 prevented the increase in nitrite, and given this evidence, we postulated a novel potential mechanism of NO release in response to α2-adrenoceptor activation, ie, local kinin production was the intermediate step responsible for the ultimate release of EDRF. To further investigate this hypothesis, inhibition of kallikrein activity was achieved by use of three different serine protease inhibitors. As demonstrated by others, aprotinin can block local kinin production23 24 25 26 and in fact is used currently to improve hemostasis during open-heart surgery that requires cardiopulmonary bypass. Aprotinin inhibits the contact activation of tissue kallikrein and thereby blocks the intrinsic coagulation cascade and prevents the total body inflammatory response that is seen with cardiopulmonary bypass.27
Kallikrein or a kallikrein-like enzyme appears to be present ubiquitously in human vascular tissue and in circulating plasma.26 28 Kinin precursors, such as high-molecular-weight kininogen, also circulate in plasma and are taken up by high-affinity binding sites present on the surface of endothelial cells.29 BK is then formed and binds to B2-receptors on the endothelial cell surface to release NO and prostacyclin. In the current study, we have shown that activation of the kallikrein-kinin system can be initiated by α2-adrenoceptor stimulation, and the end product of this is nitrite (NO2). The importance of this is clear: local kinin production, and hence NO, may modulate coronary vasoconstriction after α-adrenergic receptor stimulation in the human coronary circulation.
Many authors20 21 22 28 have speculated that the blood pressure–lowering effects of ACE-inhibitor therapy are not just a manifestation of angiotensin II inhibition but are also due to diminished BK degradation. One study30 showed that plasma kinin levels of hypertensive patients who received ACE-inhibitor therapy were not significantly altered; however, enhanced local synthesis or storage of kinins within the vascular endothelium could be responsible for the hypotensive effects of the drug. It is also possible that in pathological states such as heart failure, endogenous BK may be present in subthreshold amounts and its vasodilator capacity may only be unmasked by ACE-inhibitor therapy. In addition, these compounds can potentiate other in vitro endothelium-dependent relaxations, such as the vascular response to ACH and shear stress.22
The systemic benefit of EDRF production rests not only in its action as a vasodilator but also in its ability to diminish total tissue oxygen consumption and limit tissue metabolism, as we have recently shown31 ; myocardial tissue oxygen consumption may be regulated in part by NO synthesized by the coronary microvessel endothelium. It is interesting and perhaps only coincidental that many patients with severe heart failure are treated with ACE inhibitors. In fact, 6 of 10 patients in our study received ACE inhibition. Perhaps at least part of the beneficial effect is to preserve NO production by extending the half-life of locally formed kinins.
Recent studies indicated that myocytes may produce NO. Pinsky et al32 demonstrated increased inducible NO synthase expression in rat myocytes in response to cytokines such as interleukin-1 and tumor necrosis factor-α. Cytokines may be elaborated in response to immunologic challenge and inflammatory states, such as cardiac allograft rejection and cardiomyopathy. The end result of enhanced myocardial inducible NO synthase activity may be autotoxicity, reduced contractility (myocyte shortening) that contributes to myocyte necrosis, and eventual ventricular dysfunction. It is unclear, however, whether myocyte inducible NO synthase is active in basal conditions and whether the enhanced synthesis observed in the rat model is also seen in healthy humans and in disease states. In the present studies in human coronary microvessels, the preparations were virtually free of contaminating myocytes.
There are a number of potential limitations of the present study that should be discussed. First, our vessel preparation was a mixture of small coronary blood vessels, and reduced nitrite production may have been present in some or all of the segments, including arterioles, capillaries, and venules. Because some of our recent studies33 indicated reduced NO synthase gene expression in aorta from the dog with overt congestive heart failure, it is most likely that this reduced NO production extends throughout the vascular tree. Another potential criticism is that human microvessel nitrite production does not reflect the magnitude of the actual defect in heart failure, since the patients were medicated to preserve cardiac function and since even the microvessels from the nonfailing hearts were from “sick hearts.” Be that as it may, microvessels from the hearts of dogs with pacing-induced heart failure that were not medicated for the treatment of heart failure may be indicative of the true magnitude of the blood vessel defect, ie, they have even lower nitrite production.3 If vessels from the nonfailing human heart do not reflect normal human coronary microvessels and hypothetically also exhibit altered NO production, then the logical explanation from all of our data, human and canine, is that NO production from normal human coronary microvessels should be even higher than we measured in the nonfailing group. Finally, because of medication of these patients, the drugs may have altered NO production even after the microvessels were isolated. The microvessels were extensively washed during preparation, which took 4 to 5 hours, and this should have removed water-soluble drugs. However, if drugs such as the ACE inhibitors altered endothelial gene expression, then an altered NO production may have persisted. Even so, nitrite production from human coronary microvessels appeared to be low.
In summary, viable coronary microvessels were isolated successfully from failing and nonfailing human hearts. These microvessels produced NO in response to known agonists for NO production (ACH, BK, norepinephrine, and BHT 920), and the stimulatory action of these substances was blocked by L-NAME. The nonselective α-adrenergic receptor agonist norepinephrine and the selective α2-adrenergic receptor agonist BHT 920 also caused NO production; their action was blocked by the B2-receptor antagonist HOE 140, which implicates local kinin production as an intermediate step. Inhibitors of kallikrein activity (the serine protease inhibitors aprotinin, soybean trypsin inhibitor, and dichloroisocoumarin) all prevented NO production in response to norepinephrine and BHT 920.
Selected Abbreviations and Acronyms
|EDRF||=||endothelium-derived relaxing factor|
|L-NAME||=||NG-nitro-l-arginine methyl ester|
This study was supported by grant PO-1-HL43012 from the National Heart, Lung, and Blood Institute.
Presented in abstract form at FASEB 1994, Anaheim, Calif, and at the 67th Scientific Sessions of the American Heart Association, Dallas, Tex, November 14-17, 1994, and published in abstract form (Circulation. 1994;90:I-295).
- Received October 5, 1995.
- Revision received December 19, 1995.
- Accepted December 21, 1995.
- Copyright © 1996 by American Heart Association
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