Acetylcholine-induced Coronary Vasodilation Research Articles

Increased superoxide production causes coronary endothelial dysfunction and depressed oxygen consumption in the failing heart

This study examined whether increased superoxide (O(2)(-).) production contributes to coronary endothelial dysfunction and decreased coronary blood flow (CBF) in congestive heart failure (CHF). To test this hypothesis, the effects of the low-molecular-weight SOD mimetic M40401 on CBF and myocardial oxygen consumption (MVo(2)) were examined in dogs during normal conditions and after CHF was produced by 4 wk of rapid ventricular pacing. The development of CHF was associated with decreases of left ventricular (LV) systolic pressure, maximum first derivative of LV pressure, MVo(2), and CBF at rest and during treadmill exercise as well as endothelial dysfunction with impaired vasodilation in response to intracoronary acetylcholine. M40401 increased CBF (18 +/- 5%, P < 0.01) and MVo(2) (14 +/- 6%, P < 0.01) in CHF dogs and almost totally reversed the impaired CBF response to acetylcholine. M40401 had no effect on acetylcholine-induced coronary vasodilation, CBF, or MVo(2) in normal dogs. Western blot analysis demonstrated that extracellular SOD (EC-SOD) was significantly decreased in CHF hearts, whereas mitochondrial Mn-containing SOD was increased. Cytosolic Cu/Zn-containing SOD was unchanged. Both increased O(2)(-). production and decreased vascular O(2)(-). scavenging ability by EC-SOD could have contributed to endothelial dysfunction in the failing hearts.

Effects of repeated brief episodes of ischemia and reperfusion in isolated perfused rat hearts.

The effects of ischemia and reperfusion on the coronary endothelium and myocardium as well as tolerance to ischemia/reperfusion injury were assessed using isolated retrogradely perfused rat hearts. Repeated brief episodes of myocardial ischemia followed by reperfusion is known to have a protective effect against subsequent myocardial infarction. However, no studies have been performed with perfusion in the absence of blood cells to determine the effect of repeated ischemia and reperfusion on the coronary endothelium and myocardium. Using the Langendorff perfusion technique, rat hearts were subjected to a 30-, 10-, 5-, or 2-min period of low-flow perfusion by reducing the coronary flow to 3 ml/min followed by reperfusion at 20 ml/min for the same period of time. Control perfusion was then performed at a constant flow rate of 20 ml/ min for 60 min. Acetylcholine-induced coronary vasodilation was significantly (P < 0.05) lower in hearts subjected to 30 min of ischemia and 30 min of reperfusion when compared with the control hearts. Myocardial creatinine kinase (CK) activity was significantly reduced (P < 0.01) in hearts subjected to ischemia and reperfusion for either 30, 10, or 5 min. To assess the effect of repeated episodes of ischemia and reperfusion, the following protocols were used: a control study with constant perfusion for 60 min (group A), 30 min of ischemia and 30 min of reperfusion (group B), three 10-min episodes of ischemia and reperfusion (group C), six 5-min episodes of ischemia and reperfusion (group D), and 15 2-min episodes of ischemia and reperfusion (group E). Acetylcholine-induced coronary vasodilation was significantly inhibited in group B (80% +/- 12%, P < 0.05) and group C (70% +/- 13%, P < 0.01), but did not change significantly in either group D (123% +/-19%) or group E (142% +/- 15%), compared with the control group (group A; 127% +/- 15%, mean +/-SEM). Nitroglycerin-induced coronary vasodilation was not altered by ischemia/reperfusion in any group. In contrast, myocardial CK activity was significantly lower in group B (3.6 +/- 0.6IU/mg protein, P < 0.01), group C (3.2 +/- 0.1 IU/mg protein, P < 0.01), and group D (3.3 +/- 0.21U/mg protein, P < 0.01) than in group A (47 +/- 6.7 IU/mg protein). The myocardial CK activity of group E was not significantly different from that of group A, but was significantly higher than in groups B, C, and D (P < 0.01). In isolated perfused rat hearts, both the coronary endothelium and myocardium are damaged by repeated episodes of ischemia and reperfusion. However, the coronary endothelium is more resistant to such damage than is the myocardium.

Does inhibition of coronary nitric oxide synthesis alter coronary vascular tone in normal dogs?

To examine whether endothelial nitric oxide formation contributes to the vascular tone of resistance coronary vessels in vivo, we administered NG-nitro-L-arginine methyl ester (L-NAME) (10 and 100 micrograms/kg/min), a nitric oxide synthase inhibitor, as well as D-enantiomer into the left circumflex artery in normal dogs. Intracoronary L-NAME, which was associated with dose-related reductions in acetylcholine-induced coronary vasodilation, significantly reduced the baseline left circumflex blood flow by 6% and increased coronary vascular resistance of the left circumflex artery by 6%. D-enantiomer was ineffective in altering baseline coronary blood flow and vascular resistance of the left circumflex artery. These results indicate that continuous nitric oxide formation in the vasculature is important in the regulation of the coronary vascular tone of resistance vessels in vivo, and serves to maintain the vessels in a dilated state.

Endogenous and exogenous nitric oxide inhibits norepinephrine release from rat heart sympathetic nerves.

This study was designed to elucidate whether nitric oxide (NO) controls norepinephrine (NE) release from sympathetic nerves of the rat heart. Hearts were perfused in the Langendorff mode with Tyrode's solution. The right sympathetic nerve was stimulated with trains of 1 or 3 Hz and NE release was measured. The NO synthase (NOS) inhibitor NG-nitro-L-arginine (L-NNA) enhanced the evoked NE release in a concentration-dependent manner. This facilitation was independent of the increase in perfusion pressure and was stereospecifically reversed by L-arginine but not D-arginine. Another NOS inhibitor, NG-methyl-L-arginine, produced a similar increase in NE release. The NO-donor compound S-nitroso-N-acetyl-D,L-penicillamine, added in the presence of L-NNA, restored the suppression of NE release in a concentration-dependent fashion. A similar suppression was achieved with 3-morpholinosydnonimine. These results demonstrated that NE release is under the inhibitory control of endogenous NO. Western blots demonstrated the presence of neuronal NOS I and endothelial NOS III in the hearts. Perfusion of the hearts with a low concentration of the detergent CHAPS produced functional damage of the endothelium, as evidenced by an increase in perfusion pressure and a conversion of the acetylcholine-induced coronary vasodilation to a constriction. However, CHAPS treatment did not produce a facilitation of NE release (as did the NOS inhibitors), and L-NNA still increased NE release in CHAPS-treated hearts. Double-labeling immunofluorescence histochemistry showed NOS I immunoreactivity in stellate ganglion cells and in neurons of the heart, some of which also stained positive for tyrosine hydroxylase.(ABSTRACT TRUNCATED AT 250 WORDS)

744-3 Inhibition of Nitric Oxide Synthesis does not Increase Cardiac Contractile Response but Reduces Coronary Blood Flow Response to β-Adrenergic Stimulation in Normal Dogs

Although the induction of nitric oxide (NO) synthesis has been implicated as a cause of cytokine-induced depression of cardiac β-adrenergic responsiveness. whether the NO system constitutively present in the normal myocardium plays a role in its physiologic response to β-adrenergic stimulation in vivo remains controversial. Accordingly, we examined the effects of low and high doses of N W -nitro-L-arginine methyl ester (L-NAME)(10 and 100 μg/kg/min for 10 min), an NO synthase inhibitor, administered into left circumflex coronary artery (LCX) on responses of peak left ventricular (LV) dP/dt, regional wall thickening in LCX region and LCX blood flow to graded intracoronary (IC) doses of isoproterenol (ISO:0.002 to 0.016 μg/kg/min) in 7 anesthetized dogs. IC L-NAME was associated with dose-related reductions in IC acetylcholine-induced coronary vasodilation. Effects of L-NAME on ISO-induced changes are shown: baseline ISO:0.002 0.004 0.008 .0016 Peak LV dP/dt (mmHg/sec) (n = 7) control 2029 ± 136 2586 ± 192 2820 ± 200 3309 ± 255 4120 ± 419 * low L-NAME 2171 ± 149 2566 ± 176 2894 ± 206 3214 ± 223 3707 ± 250 * high L-NAME 2114 ± 166 2326 ± 193 2560 ± 152 3014 ± 140 3354 ± 171 * Wall thickening (%) (n = 2) control 22 ± 7 25 ± 6 29 ± 5 33 ± 7 35 ± 9 low L-NAME 25 ± 11 25 ± 15 28 ± 19 31 ± 18 36 ± 21 high L-NAME 28 ± 17 25 ± 15 25 ± 15 31 ± 19 34 ± 15 LCX blood flow (ml/min)(n = 7) control 33 ± 6 48 ± 7 52 ± 6 61 ± 8 70 ± 9 * low L-NAME 36 ± 7 41 ± 8 44 ± 9 47 ± 8 52 ± 9 * high L-NAME 33 ± 7 36 ± 8 38 ± 7 40 ± 7 48 ± 8 * mean ± SEM * p &lt; 0.05 Thus, inhibition of NO synthesis by L-NAME did not change baseline contractility nor did it increase its response to ISO. It also did not alter baseline blood flow, but reduced significantly its response to ISO. These data strongly suggest that the NO system in the normal myocardium does not modulate contractility, but NO formation in the vasculature contributes to the β-adrenergic coronary vasodilation.

Modulation by aging of the coronary vascular response to endothelin-1 in the rat isolated perfused heart

Changes with age in the coronary vascular response to endothelin-1 were investigated in perfused hearts isolated from 2-, 6- and 24-month-old (mo) male Fisher-344 rats. Endothelin-1 injected as a single bolus (0.3, 3 and 30 nmol) into the coronary artery supply caused dose-dependent vasoconstriction in all three age groups. While there was no age-related change in the vasoconstriction induced by the lower doses (0.3 and 3 nmol), the higher dose (30 nmol) elicited a more pronounced vasoconstriction in 6- and 24-mo rats than that in 2-mo rats. NG-nitro-L-arginine (L-NNA), an inhibitor of nitric oxide formation, markedly enhanced the vasoconstriction induced by 30 nmol endothelin-1 in 2- and 6-mo rats but only slightly and non-significantly enhanced that vasoconstriction in 24-mo rats. Haemoglobin, which inhibits activation of guanylate cyclase by nitric oxide, enhanced the endothelin-1-induced vasoconstriction in 2-mo rats, but not in 6- and 24-mo rats. The acetylcholine-induced coronary vasodilation was more pronounced in 2- and 6-mo rats than in 24-mo rats and was attenuated by L-NNA in 2- and 6-mo rats. The coronary vasodilation induced by nitroprusside (0.1 mmol), a pharmacological precursor of nitric oxide, did not change with age. Endothelin-1 (30 nmol) markedly increased the release of 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha) in all three age groups. The prostaglandin synthesis inhibitor indomethacin enhanced the endothelin-1-induced vasoconstriction in 2- and 6-mo rats to a similar extent.(ABSTRACT TRUNCATED AT 250 WORDS)

Hemoglobin inhibits endothelium-dependent relaxation to acetylcholine in human coronary arteries in vivo.

The endothelium can regulate vascular tone by releasing both endothelium-derived relaxing factor (EDRF or nitric oxide) and contracting factors. To date, there has only been circumstantial evidence to indicate EDRF activity in vivo in human coronary arteries. Using human hemoglobin as a specific inhibitor, the hypothesis that acetylcholine-induced coronary vasodilation is due to EDRF release was tested. We studied the response of normal coronary arteries to acetylcholine (an endothelium-dependent vasodilator) and isosorbide dinitrate (an endothelium-independent vasodilator) in seven patients. The specificity of any vasodilator response was assessed by the infusion of reduced free human hemoglobin. Hemoglobin 10(-5) M infusion alone had no effect on coronary artery diameter. Drugs were infused into the coronary artery, and the diameter changes were assessed by quantitative angiography. Acetylcholine 10(-7) M increased left anterior descending coronary artery diameter from control: 2.30 +/- 0.12 mm to 2.79 +/- 0.20 mm (mean +/- SEM, n = 7, p < 0.01). Hemoglobin both in a concentration of 10(-6) M and 10(-5) M reversed this vasodilator effect, causing constriction to 2.11 +/- 0.18 mm (p < 0.001 compared with acetylcholine 10(-7) M) and 2.29 +/- 0.14 mm (p < 0.05 compared with acetylcholine 10(-7) M). Isosorbide dinitrate in the presence of hemoglobin caused dilatation of the coronary artery in all cases to 3.04 +/- 0.24 mm (p < 0.001 compared with acetylcholine 10(-7) M and hemoglobin 10(-6) M). Using a specific inhibitor of nitric oxide, reduced free hemoglobin, we have demonstrated that basal EDRF release does not appear to play an important role in the maintenance of human epicardial coronary artery diameter in vivo but is responsible for the acetylcholine-induced dilatation.

Endothelium-Derived Nitric Oxide as a Mediator of Acetylcholine-Induced Coronary Vasodilation in Dogs

To investigate the role of endothelium-derived nitric oxide (NO) in regulation of the tonus of the coronary artery system in vivo, the effect of NG-mono-methyl-L-arginine (L-NMMA), a specific inhibitor of the synthesis of NO from L-arginine, on acetylcholine (ACh)-induced coronary vasodilation was examined in open-chest, anesthetized dogs. At baseline, the infusion of ACh and adenosine into the left circumflex coronary artery (LCX) increased the LCX blood flow. Continuous infusion of L-NMMA (2 mumol/min, for 20 min) into LCX significantly attenuated ACh-induced increase in the LCX flow: percentage increases in the LCX flow by ACh (10 and 100 ng/kg) before L-NMMA were 42.0 +/- 8.7 and 137.3 +/- 12.5%, respectively, whereas those after L-NMMA were 14.0 +/- 4.9 (p less than 0.02) and 88.9 +/- 10.9% (p less than 0.005), respectively. L-NMMA showed no effect on adenosine-induced increase in the LCX flow. After the L-NMMA infusion was terminated, the effect of ACh partially returned to the baseline level. In other dogs, L-arginine (3 mg/min, for 20 min) was administered just after the L-NMMA infusion, which completely reversed the inhibitory effect of L-NMMA on ACh-induced vasodilation. Endothelium-derived NO is a mediator of ACh-induced vasodilation of the coronary resistance vessels in dogs.

Role of purines in acetylcholine-induced coronary vasodilation

Acetylcholine, when infused directly into the coronary arteries, increases coronary flow and stimulation of the vagus causes parasympathetic coronary vasodilation. Recent evidence suggests that this acetylcholine-induced relaxation of coronary resistance vessels is not the result of a direct effect on vascular smooth muscle cells, but rather involves endothelial cell mediated release of some unidentified substance(s) [ 5]. Possible candidates which may participate in this coupling are adenine nucleotides and adenosine. These purine compounds which act on the surface of coronary myocytes via specific receptors are powerful dilators of coronary vessels [ 3, 8, 11]. We report here that stimulation of cardiac muscarinic receptors by acetylcholine causes a marked increase in the release of ATP, AMP, and adenosine into the coronary system of isolated guinea-pig hearts independent of myocardial oxygen consumption. The amounts of adenosine liberated appear quantitatively sufficient to be causally involved in the coronary dilatory action of acetylcholine.

Transmural distribution of blood flow during activation of coronary muscarinic receptors.

The role of coronary muscarinic receptors in the distribution of transmural blood flow across the left ventricular wall of the working heart was studied in anesthetized open-chest dogs. Tissue blood flow in subepicardium, midmyocardium, and subendocardium was determined with radioactive microspheres before and during activation of muscarinic vasodilator receptors by intracoronary infusions of acetylcholine. Myocardial and coronary vascular beta-receptors were blocked by sotalol (2.0 mg/kg iv). Equivalent submaximal coronary vasodilator doses of acetylcholine and adenosine were compared for effects on transmural blood flow. Intracoronary infusions of acetylcholine (5.0 and 10.7 micrograms/min) produced a dose-related increase in the subendocardial-subepicardial blood flow ratio (endo/epi) from 1.07 to 1.32 and 1.57, respectively. A progressively larger decrease in coronary vascular resistance occurred in the subendocardium than midmyocardium or subepicardium following acetylcholine administration. In contrast, intracoronary administration of adenosine (54.4 micrograms/min) produced no change in endo/epi. Atropine effectively blocked acetylcholine-induced coronary vasodilation but not vasodilation produced by adenosine. Neither agent affected heart rate, left ventricular pressure, coronary perfusion pressure, or myocardial contractility. These results suggest that activation of muscarinic coronary vasodilator receptors redistributes blood flow preferentially to the subendocardium independent of cardiac mechanical influences.