Endothelium-dependent Coronary Vasodilation Research Articles

Cholesterol reduction improves myocardial perfusion abnormalities in patients with coronary artery disease and average cholesterol levels

OBJECTIVESWe sought to evaluate whether pravastatin treatment increases myocardial perfusion, as assessed by thallium-201 single-photon emission computed tomographic (SPECT) dipyridamole testing, in patients with coronary artery disease (CAD) and average cholesterol levels.BACKGROUNDPrevious studies in hypercholesterolemic patients have demonstrated that cholesterol reduction restores peripheral and coronary endothelium-dependent vasodilation and increases myocardial perfusion.METHODSThis was a randomized, placebo-controlled study with a cross-over design. Twenty patients with CAD were randomly assigned to receive 20 mg of pravastatin or placebo for 16 weeks and then were crossed over to the opposite medication for a further 16 weeks. Lipid and lipoprotein analysis and dipyridamole thallium-201 SPECT were performed at the end of each period. The SPECT images were visually analyzed in eight myocardial segments using a 4-point scoring system by two independent observers. A summed stress score and a summed rest score were obtained for each patient. Quantitative evaluation was performed by the Cedars-Sinai method. The magnitude of the defect was expressed as a percentage of global myocardial perfusion.RESULTSTotal and low density lipoprotein cholesterol levels during placebo were 214 ± 29 mg/dl and 148 ± 25 mg/dl, respectively. These levels with pravastatin were 170 ± 23 mg/dl and 103 ± 23 mg/dl, respectively. The summed stress score and summed rest score were lower with pravastatin than with placebo (7.2 ± 2.3 vs. 5.9 ± 2.3, p = 0.012 and 3.2 ± 1.6 vs. 2.4 ± 2.2, p = 0.043, respectively). Quantitative analysis showed a smaller perfusion defect with pravastatin (29.2%) as compared with placebo (33.8%) (p = 0.021) during dipyridamole stress. No differences were found at rest.CONCLUSIONSReducing cholesterol levels with pravastatin in patients with CAD improves myocardial perfusion during dipyridamole stress thallium-201 SPECT.

49-Year-Old Woman With Chest Pain

49-Year-Old Woman With Chest Pain

Reduced coronary NO production in conscious dogs after the development of alloxan-induced diabetes

The role of nitric oxide (NO) in the control of coronary blood flow (CBF) during the development of diabetes is unknown. To study this, mongrel dogs were chronically instrumented using sterile techniques for measurements of systemic hemodynamics and CBF. With heart rate controlled (150 beats/min), veratrine (1-10 micrograms/kg) caused dose-dependent increases in CBF; e.g., 5 mirograms/kg of veratrine increased CBF by 57 +/- 7% from 41 +/- 1.3 ml/min (P < 0.05). The dogs developed diabetes 4-5 wk after injection of alloxan (40-60 mg/kg iv, blood glucose levels were 384 +/- 18 mg/dl). After diabetes the same doses of veratrine caused smaller increases in CBF; i.e., 5 micrograms/kg of veratrine increased CBF by 32 +/- 2% (P < 0.05 compared with control) from 28 +/- 4 ml/min. ACh- and adenosine-induced coronary vasodilation were reduced after diabetes as well. In anesthetized dogs after diabetes, vagal stimulation caused smaller increases in CBF. ACh and bradykinin caused smaller increases in NO(-)(2) production in coronary microvessels from diabetic dogs. Furthermore, despite the fact that mRNA for endothelial cell NO synthase from the aorta was increased twofold with the use of Northern blotting, the protein for aortic endothelial constitutive NO synthase was reduced by 66% after diabetes, as determined by Western blotting. Our results indicate that the NO-dependent coronary vasodilation by the Bezold-Jarisch reflex is impaired in conscious dogs after diabetes. The mechanism responsible for the impaired endothelium-dependent coronary vasodilation is most likely the decreased release of NO from the endothelium.

The effects of perhexiline on the rat coronary vasculature

The predominant site and mechanism(s) of perhexiline-induced coronary vasodilatation were investigated in the rat heart. Perhexiline was more potent in the Langendorff perfused heart than in the left anterior descending coronary artery (EC 50; 0.27 μM, confidence limits 0.19–0.39: 2.7 μM, 2.0–3.4, respectively). Selective endothelial inactivation with Triton X-100 in the perfused heart, reduced the response to perhexiline 1 μM (76+8% to 30+3% of control). 1 H-[1,2,4]Oxadiazolo[4,3- a]quinoxalin-1-one (ODQ) 3 μM, Nϖ-nitro- l-arginine 100 μM, or a combination of the latter with indomethacin 10 μM, had no significant effect on responses to perhexiline in the perfused heart. Unlike bradykinin-induced vasodilatation, responses to perhexiline were not inhibited by tetrabutylammonium 1 mM, or charybdotoxin 20 nM. SKF525A 5 μM inhibited both perhexiline and bradykinin responses, while apamin 1 μM and glibenclamide 3 μM inhibited neither. Perhexiline exerts partially endothelium-dependent coronary vasodilator effects in the rat, predominantly on small coronary arteries, which appear to be independent of nitric oxide (NO), prostacyclin and the endothelium-derived hyperpolarising factor (EDHF) released by bradykinin.

NITRIC OXIDE DOES NOT MEDIATE CORONARY VASODILATION DURING ACUTE NORMOVOLEMIC HEMODILUTION IN DOGS

S72 INTRODUCTION: Acute normovolemic hemodilution (ANH) is accompanied by increases in coronary blood flow (CBF) that are attributable to the combined effects of reduced blood viscosity and vasodilation [1]. The mechanism for this vasodilation is unknown. While previous findings have implied that endothelium-derived nitric oxide (EDNO) is involved in the systemic vasodilation accompanying ANH [2], the role of this mechanism in the coronary circulation remains to be evaluated. Accordingly, the present study was performed to test the hypothesis that inhibition of EDNO by the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) would compromise myocardial perfusion during ANH. METHODS: After approval from the University Animal Care Committee, studies were conducted in ten open-chest dogs anesthetized with isoflurane and fentanyl. Measurements of myocardial blood flow (MBF) were obtained in the perfusion territories of the left anterior descending (LAD) and circumflex (CX) coronary arteries using radioactive microspheres. Mean aortic pressure (MAP), mean left atrial pressure (MLAP), and heart rate (HR) were measured using standard methods. A Doppler flow transducer was used to measure CBF in the LAD. Two groups were studied. In Group 1 (NOS Inhibition; n=5), the LAD region received an intracoronary (i.c.) infusion of L-NAME (4.5 mg), and the CX region served as a control. Effectiveness of L-NAME was demonstrated by the its ability to inhibit the increases in CBF by the endothelium-dependent coronary vasodilator acetylcholine (20 [micro sign]g/min, i.c.), while having no effect on those by the endothelium-independent coronary vasodilator sodium nitroprusside (80 [micro sign]g/min, i.c.). In Group 2 (Endothelium Intact; n=5), neither the LAD nor the CX region was treated with L-NAME. This was done to validate the use of the CX bed as a control for the LAD bed in Group 1. Data were obtained under baseline conditions (hematocrit; 40 +/- 2 %) and following ANH with 5% dextran-40 to an hematocrit of 20 +/- 1 %. RESULTS: ANH caused similar increases in MBF in LAD and CX regions, whether or not the LAD region was treated with L-NAME (Figure 1). ANH had no effect on MAP, MLAP, or HR.Figure 1DISCUSSION: Pretreatment with L-NAME did not compromise myocardial perfusion during ANH. This finding implies that EDNO does not mediate coronary vasodilation under this condition.

The role of nitric oxide in coronary vascular effects of estrogen in postmenopausal women.

At physiological concentrations, 17beta-estradiol selectively enhances endothelium-dependent coronary vasodilation by an unknown mechanism in postmenopausal women. To assess the contribution of nitric oxide (NO) to the vascular effects of estradiol, we measured coronary epicardial and microvascular responses to intracoronary acetylcholine (range, 3 to 300 microg/min for 2 minutes) before and after intracoronary estradiol 75 ng/min for 15 minutes in 20 estrogen-deficient women, 16 of whom had angiographic evidence of atherosclerosis or risk factors for atherosclerosis. This testing was repeated after inhibition of NO synthesis with intracoronary N(G)-monomethyl-L-arginine (L-NMMA) 64 micromol/min for 5 minutes. Estradiol increased acetylcholine-stimulated coronary flow from 54+/-48% (mean+/-SD) above baseline values before estradiol infusion to 100+/-63% above baseline values (P=.007) and decreased coronary resistance from 32+/-21% to 46+/-15% below baseline values (P=.007) at a coronary sinus estradiol concentration of 1725+/-705 pmol/L (470+/-192 pg/mL). Estradiol also tended to lessen the severity of acetylcholine-induced epicardial coronary artery vasoconstriction from 8+/-11% to 3+/-11% below baseline values (P=.123). However, during L-NMMA infusion, estradiol no longer potentiated the effects of acetylcholine on coronary flow dynamics; coronary flow increased 39+/-46% above baseline values and coronary resistance decreased 19+/-30% below baseline values (both P<.001 versus pre-L-NMMA responses). The epicardial diameter decreased 8+/-11% below baseline values (P=.06 versus pre-L-NMMA response). The effects of estradiol at physiological concentrations on endothelium-dependent coronary vasodilator responsiveness in postmenopausal women are mediated by enhanced bioavailability of NO, which may be responsible in part for the cardioprotective effects of estrogen.

Myocardial Contractile Effects of l-Arginine in the Human Allograft

Objectives. In the present study, we investigated, in transplant recipients, whether l-arginine (l-arg) potentiates the myocardial contractile effects of receptor-mediated coronary endothelial stimulation. Moreover, because inducible nitric oxide synthase (iNOS) is frequently expressed in transplanted myocardium, we also performed intracoronary infusion of l-arg in the absence of receptor-mediated coronary endothelial stimulation to investigate whether similar left ventricular (LV) contractile effects could be induced by providing more substrate for iNOS.Background. Nitric oxide (NO), released from coronary endothelium after receptor-mediated stimulation by substance P (SP), affects vascular smooth muscle tone and modulates LV contractile performance. l-arg augments receptor-mediated endothelium-dependent coronary vasodilation in transplant recipients by increasing substrate availability for endothelial NO production.Methods. Sixteen transplant recipients were studied at the time of annual coronary angiography. In eight transplant recipients, microtip LV pressures were recorded before and during intracoronary (IC) SP (20 pmol/min) and after the addition of IC l-arg (160 μmol/min) to IC SP. In eight transplant recipients, microtip LV pressures were recorded before and during IC l-arg (160 μmol/min) alone, and in six of these patients, endomyocardial biopsy samples were obtained to detect the expression of iNOS gene by reverse transcription–polymerase chain reaction.Results. Addition of IC l-arg to IC SP induced a fall (mean ± SEM) in LV peak systolic pressure (−16 ± 4 mm Hg), which was larger (p < 0.01) than that observed during IC SP (−7 ± 2 mm Hg). During IC l-arg alone, there was no change in LV peak systolic pressure despite the presence of iNOS mRNA in five of the six biopsy samples.Conclusions. In transplant recipients, l-arg potentiates the paracrine myocardial contractile effects of receptor-mediated coronary endothelial stimulation, probably by providing more substrate for endothelial NO production. Despite the myocardial expression of iNOS gene, l-arg alone fails to elicit similar contractile effects.(J Am Coll Cardiol 1997;29:1332–8)

Effects of L-arginine supplementation on endothelium-dependent coronary vasodilation in patients with angina pectoris and normal coronary arteriograms.

The pathogenesis of impaired endothelium-dependent coronary vasodilation in angina pectoris and normal coronary arteriograms (microvascular angina pectoris) is not known. We examined whether supplementation with L-arginine, a precursor of endothelium-derived nitric oxide, improves endothelium-dependent coronary vasodilation in patients with microvascular angina. The effect of intracoronary infusion of L-arginine (50 mg/mm) on acetylcholine-induced coronary vasomotion was studied in eight patients with microvascular angina and eight control subjects. The responses of the large epicardial coronary artery diameter and coronary blood flow were measured with coronary arteriography and an intracoronary Doppler catheter, respectively. Acetylcholine increased coronary blood flow with modest vasoconstriction of the large coronary artery without altering arterial pressure and heart rate. The acetylcholine-induced increases in coronary blood flow were significantly less (P < .01) in patients than in control subjects. L-Arginine significantly augmented the coronary blood flow responses to acetylcholine in patients, but not in control subjects. L-Arginine did not alter responses of the large coronary artery in either group. Study results suggest that L-arginine improved endothelium-dependent vasodilation of coronary microcirculation in patients with microvascular angina pectoris.

Relationship Between Perfusion Pressure and Coronary Vasoreactivity in Saline Perfused Hearts Isolated from Normotensive and Spontaneously Hypertensive Rats: Role of Endothelium

Although reduced endothelium—dependent coronary vasodilation is a consistent finding in patients with hypertension, studies in perfused hearts isolated from spontaneously hypertensive rats (SHR) lead to controversial results. One possible explanation is that coronary vascular reactivity in SHR hearts were studied at different fixed perfusion pressure levels, which were outside the arterial blood pressure range found in these animals. Therefore, the objective of the present experiments was to systematically study the effect of an endothelium-dependent (serotonin) and endothelium—independent (sodium nitroprusside, SNP) vasodilator on coronary flow rate at a wide range of perfusion pressure levels (55 to 130 mmHg) in saline perfused hearts isolated from normotensive (NWR) and SH rats. The pressure-flow relationship showed coronary flow autoregulation at different pressure levels in the two groups (55 to 90 mmHg in NWR hearts and 110 to 130 mmHg in SHR hearts). At a maximally effective concentration (104M) co...

Acute supplementation with the nitric oxide precursor l-arginine does not improve cardiovascular performance in patients with hypercholesterolemia

Endothelial dysfunction based on lack of nitric oxide (NO) may contribute to several settings of cardiovascular disorder. Chronic oral supplementation with the NO precursor l-arginine counteracts the development of aortic atherosclerosis in cholesterol-fed rabbits, and i.v. infusion of l-arginine may acutely improve endothelium-dependent coronary epicardial vasodilation in patients with hypercholesterolemia (HC). To clarify whether excess NO precursor may also improve general cardiovascular performance in HC, we measured working capacity indices of myocardial ischemia, and basal and post-occlusive forearm and skin blood flow in nine patients with elevated plasma cholesterol (9.1 ± 0.2 μmol/l) following random double-blinded administration of l-arginine (16 g i.v.) or placebo. Infusion of l-arginine raised the plasma concentration of this amino acid from 85 ± 12 to 2460 ± 230 μmol/l but did not change the plasma level of the major NO metabolite nitrate. Maximal working capacity, indices of myocardial ischemia, and basal and post-occlusive blood flow in the skin or forearm did not differ between the treatments. The lack of positive effect of l-arginine compared to placebo indicates that excess NO precursor did not improve microvascular endothelial function in the patients, or alternatively, that the indices measured in the present study were not dependent on endothelial microvessel function. Thus, in patients with HC, deficiency of precursor for NO formation does not seem to impair either maximal exercise capacity myocardial perfusion during maximal exercise, or maximal vasodilator capacity in skeletal muscle or skin.

Coronary vascular reactivity is abnormal in patients with Chagas' heart disease

Symptoms of myocardial ischemia, such as chest pain (sometimes with anginal features), acute myocardial infarction, and segmental wall motion abnormalities (including left ventricular apical aneurysm), frequently occur in patients with Chagas' heart disease. Because these clinical findings occur in the presence of normal coronary arteries, it is possible that an abnormality of the coronary vascular reactivity could be present in these patients. Therefore the current study was undertaken to determine whether endothelium-dependent coronary vasodilation is abnormal in Chagas' heart disease. Coronary endothelial function was assessed by infusing the endothelium-dependent vasodilator acetylcholine (10 −8 to 10 −6 mol/L) and the endothelium-independent vasodilator adenosine (10 −4 mol/L) into the left anterior descending coronary artery of nine patients (age 43 ± 4 years) with Chagas' heart disease. Coronary blood flow was measured with a Doppler flow velocity catheter and by quantitative coronary cineangiography. The left ventricular ejection fraction was 39% ± 5%; eight patients had a left ventricular apical aneurysm; and one had an area of anteroapical hypokinesis. An impairment of the endothelium-dependent coronary vasodilation was demonstrated by a reduction in coronary blood flow of 41.2% ± 12.8% produced by the infusion of acetylcholine at 10 −6 mol/L and by a blunted but preserved increase in coronary blood flow of 114.6% ± 65.0% with the infusion of adenosine at 10 −4 mol/L (p = 0.03). In conclusion, patients with Chagas' heart disease have an abnormality of the coronary endothelium-dependent vasodilation, and this abnormality may play a role in their chest pain syndrome and in the development of segmental wall motion abnormalities.

Early Impairment of Acetylcholine-Induced Endothelium-Dependent Coronary Vasodilation Is Not Predictive of Secondary Graft Atherosclerosis

To test the hypothesis that the magnitude of early constriction of coronary arteries to acetylcholine might be a useful predictor of secondary graft atherosclerosis. The responses of epicardial coronary arteries to stepwise intracoronary infusion of acetylcholine (10(-8)M to 10(-5)M) were compared in 7 control subjects and in 18 patients who had undergone transplants within 2 months after surgery. Vessel dimensions (percent basal diameter) were measured by quantitative angiography. Follow-up at 1 year showed angiographically normal coronary arteries in 12 patients (group 1) and coronary atherosclerosis in 6 patients (group 2). In control subjects, acetylcholine induced a dose-dependent dilation from 10(-8)M to 10(-6)M. No significant variation was observed at 10(-5)M. In patients with transplants early after surgery, diameters did not vary significantly from base at 10(-8)M in either group and constricted significantly at higher concentrations. Vasodilator responses to intracoronary isosorbide dinitrate were similar in both groups with transplants early after surgery, and at 1 year in group 1, but significantly lower than in control subjects. In patients who had undergone transplants, acetylcholine-induced endothelium-dependent coronary artery dilation is similarly impaired early after surgery (within 2 months) in patients with and without coronary atherosclerosis at 1-year follow-up. Thus, response to acetylcholine is not a predictor of secondary atherosclerosis in patients with heart transplants.

The Effect of Cholesterol-Lowering and Antioxidant Therapy on Endothelium-Dependent Coronary Vasomotion

Patients with coronary artery disease and abnormalities of serum lipids often have endothelial vasodilator dysfunction, which may contribute to ischemic cardiac events. Whether cholesterol-lowering or antioxidant therapy can restore endothelium-dependent coronary vasodilation is unknown. We randomly assigned 49 patients (mean serum cholesterol level, 209 +/- 33 mg per deciliter [5.40 +/- 0.85 mmol per liter]) to receive one of three treatments: an American Heart Association Step 1 diet (the diet group, 11 patients); lovastatin and cholestyramine (the low-density lipoprotein [LDL]-lowering group, 21 patients); or lovastatin and probucol (the LDL-lowering-antioxidant group, 17 patients). Endothelium-dependent coronary-artery vasomotion in response to an intracoronary infusion of acetylcholine (10(-8) to 10(-6) M) was assessed at base line and after one year of therapy. Vasoconstrictor responses to these doses of acetylcholine are considered to be abnormal. Treatment resulted in significant reductions in LDL cholesterol levels of 41 +/- 22 percent in the LDL-lowering-antioxidant group and 38 +/- 20 percent in the LDL-lowering group (P < 0.001 vs. the diet group). The maximal changes in coronary-artery diameter with acetylcholine at base line and at follow-up were -19 and -2 percent, respectively, in the LDL-lowering-antioxidant group, -15 and -6 percent in the LDL-lowering group, and -14 and -19 percent in the diet group (P < 0.01 for the LDL-lowering-antioxidant group vs. the diet group; P = 0.08 for the LDL-lowering group vs. the diet group). (The negative numbers indicate vasoconstriction). Thus, the greatest improvement in the vasoconstrictor response was seen in the LDL-lowering-antioxidant group. The improvement in endothelium-dependent vasomotion with cholesterol-lowering and antioxidant therapy may have important implications for the activity of myocardial ischemia and may explain in part the reduced incidence of adverse coronary events that is known to result from cholesterol-lowering therapy.

Impaired endothelium-dependent vasodilation of large epicardial and resistance coronary arteries in patients with essential hypertension. Different responses to acetylcholine and substance P.

Hypertensive patients have impaired endothelium-dependent coronary vasodilation evoked with acetylcholine. The aim of this study was to examine whether the impaired endothelium-dependent dilation of coronary arteries is related to a specific abnormality of the muscarinic receptor that mediates the effects of acetylcholine. Responses of the large epicardial and resistance coronary arteries were assessed in seven hypertensive patients (mean arterial pressure, 106 +/- 14 mm Hg) and seven control subjects (83 +/- 6 mm Hg, P < .01) during cardiac catheterization. To assess coronary endothelial function, we infused acetylcholine and substance P (endothelium-dependent agents that act on different receptors) and papaverine and nitrate (direct vascular smooth muscle dilators) into the left anterior descending coronary artery and determined coronary artery diameter by arteriography and coronary blood flow with an intracoronary Doppler catheter technique. In control subjects, 3 micrograms/min acetylcholine increased (P < .05) and 30 micrograms/min acetylcholine decreased (P < .05) arterial diameter, and in hypertensive patients, 1, 3, 10, and 30 micrograms/min acetylcholine decreased arterial diameter in a dose-dependent manner. Substance P at 3, 10, and 30 ng/min caused comparable increases in diameter in both groups. Increases in coronary blood flow with both acetylcholine and substance P were significantly (P < .01) blunted in hypertensive patients compared with control subjects. No significant differences were noted between the groups in the responses of large epicardial coronary artery diameter and coronary blood flow to papaverine and nitrate.(ABSTRACT TRUNCATED AT 250 WORDS)

Organ Protection

Patients with arterial hypertension often have angina pectoris and a positive exercise tolerance test, even when their epicardial coronary arteries are of normal anatomic appearance. In these patients, coronary vasodilator capacity is frequently impaired by 30-50%, predominantly owing to microvascular alterations. These include medical thickening, decreased capillary density, perivascular fibrosis, and an impaired endothelium-dependent coronary vasodilation. An antihypertensive treatment that can be considered cardioreparative restores myocardial structure through both regression of myocardial fibrosis and regression of the structural coronary microvascular alterations. We believe that this is the first clinical study to show that coronary flow reserve was considerably improved after long-term medical blood pressure normalization.

Fish oil improves endothelium-dependent coronary vasodilation in heart transplant recipients

Objectives. The purpose of this study was to determine whether dietary fish oil supplementation enhances endothelium-mediated vasodilator responses in human heart transplant recipients, a group known to have coronary artery disease and endothelial dysfunction.Background. Omega-3 fatty acid supplementation has been shown to enhance endothelium-dependent coronary vasodilation in animal models of atherosclerosis,Methods. Endothelium-dependent vasodilator responses to intracoronary acetylcholine infusion and endothelium-independent responses to nitroglycerin were evaluated in heart transplant recipients who received a high dose of dietary supplementation with omega-3 fatty acids for 3 weeks (5 g of eicosapentaenoic acid plus docosahexacnoic acid/day, n = 7) and compared with responses in a group of matched heart transplant recipients who did not receive fish oil (control patients, n = 7). Acetylcholine was selectively infused into the midportion of the left anterior descending or left circumflex coronary artery, with the noninfused left coronary artery serving as a control vessel. Serial coronary angiograms were performed after infusion with increasing doses of acetylcholine (infused concentrations 10−6to 10−3mol/liter) and after intracoronary nitroglycerin administration.Results. The patients treated with fish oil showed a normal vasodilator response to acetylcholine with 14 ± 2.5% and 15 ± 7% vasodilation (vs. baseline diameter) at infused acetylcholine concentrations of 10−5and 10−4mol/liter, respectively. In contrast, the control patients demonstrated vasoconstrictor responses (−1 ± 1% and −9 ± 4%) to acetylcholine at these same doses (p < 0.05 and < 0.005, respectively, for treated vs. control patients). There were no differences in the response to nitroglycerin between the control and treated patients.Conclusions. Dietary supplementation with fish oil significantly alters endothelium-dependent coronary vasodilation in heart transplant recipients without alteration of the responses to endothelium-independent vasodilation. Whether this enhancement of endothelial function can beneficially alter the natural history of heart transplant atherosclerosis warrants further study.

Mechanism of Thrombin-Induced Endothelium-Dependent Coronary Vasodilation in Dogs: Role of its Proteolytic Enzymatic Activity

We have recently reported that exogenous thrombin produced a dose- and endothelium-dependent coronary vasodilation in both intact open-chested dogs and in isolated dog coronary artery preparations. To determine whether the observed vasodilatory effect may be related to thrombin proteolytic enzymatic activity, effects of other proteases, such as trypsin, chymotrypsin, and pepsin, on the mechanical responses of isolated dog coronary arteries were studied. Among the four proteases evaluated, only thrombin (0.01-0.1 U/ml) and trypsin (0.03-0.67 U/ml) consistently produced a potent dose- and endothelium-dependent relaxation, that was reproducible with repeated testings. Addition of chymotrypsin (0.01-1.0 U/ml) produced only a minimal effect and was not reproducible, while addition of pepsin, as much as 10 U/ml, did not produce any effect. The specific soybean trypsin inhibitor and aprotinin, but not heparin and hirudin, competitively shifted the trypsin dose-response to the right, whereas heparin, hirudin, and antithrombin III proved to be more effective than trypsin inhibitors in inhibiting the thrombin-induced vasodilation. In all cases, the thrombin- and trypsin-induced vasodilation were equally sensitive to inhibition by the specific synthetic thrombin inhibitor, PPACK (D-phenylalanyl-L-propyl-L-arginine chloromethyl ketone, 1-30 nM). PPACK, however, had no effect on the other endothelium-dependent coronary vasodilators, such as acetylcholine and adenosine triphosphate, in our isolated dog coronary artery preparations. Biochemical determinations of the amidolytic activity of thrombin, using Tosylglycyl-L-prolyl-L-arginine-p-nitroanilide as a chromogen, also indicated a similar PPACK and heparin-antithrombin III dose-dependent inhibition of the thrombin enzymatic activity.(ABSTRACT TRUNCATED AT 250 WORDS)