Thomas F. Lüscher

Nitric Oxide Is Responsible for Flow-Dependent Dilatation of Human Peripheral Conduit Arteries In Vivo

BACKGROUND Experimental evidence suggests that flow-dependent dilatation of conduit arteries is mediated by nitric oxide (NO) and/or prostacyclin. The present study was designed to assess whether NO or prostacyclin also contributes to flow-dependent dilatation of conduit arteries in humans. METHODS AND RESULTS Radial artery internal diameter (ID) was measured continuously in 16 healthy volunteers (age, 24 +/- 1 years) with a transcutaneous A-mode echo-tracking system coupled to a Doppler device for the measurement of radial blood flow. In 8 subjects, a catheter was inserted into the brachial artery for measurement of arterial pressure and infusion of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA; 8 mumol/min for 7 minutes; infusion rate, 0.8 mL/min). Flow-dependent dilatation was evaluated before and after L-NMMA or aspirin as the response of the radial artery to an acute increase in flow (reactive hyperemia after a 3-minute cuff wrist occlusion). Under control conditions, release of the occlusion induced a marked increase in radial blood flow (from 24 +/- 3 to 73 +/- 11 mL/min; P < .01) followed by a delayed increase in radial diameter (flow-mediated dilatation; from 2.67 +/- 0.10 to 2.77 +/- 0.12 mm; P < .01) without any change in heart rate or arterial pressure. L-NMMA decreased basal forearm blood flow (from 24 +/- 3 to 13 +/- 3 mL/min; P < .05) without affecting basal radial artery diameter, heart rate, or arterial pressure, whereas aspirin (1 g PO) was without any hemodynamic effect. In the presence of L-NMMA, the peak flow response during hyperemia was not affected (76 +/- 12 mL/min), but the duration of the hyperemic response was markedly reduced, and the flow-dependent dilatation of the radial artery was abolished and converted to a vasoconstriction (from 2.62 +/- 0.11 to 2.55 +/- 0.11 mm; P < .01). In contrast, aspirin did not affect the hyperemic response nor the flow-dependent dilatation of the radial artery. CONCLUSIONS The present investigation demonstrates that NO, but not prostacyclin, is essential for flow-mediated dilatation of large human arteries. Hence, this response can be used as a test for the L-arginine/NO pathway in clinical studies.

Indirect evidence for release of endothelium-derived relaxing factor in human forearm circulation in vivo. Blunted response in essential hypertension.

In isolated blood vessels, acetylcholine releases endothelium-derived relaxing factor (EDRF). In vivo, the vasodilator action of acetylcholine may be mediated by EDRF, but prostacyclin or prejunctional inhibition of adrenergic neurotransmission may also be involved. Therefore, we investigated whether acetylcholine releases EDRF in humans in vivo and, if so, whether the response altered in essential hypertension. Acetylcholine was infused into the brachial artery, and forearm blood flow measured by venous occlusion plethysmography. In control subjects, acetylcholine (0.02-16 micrograms/min/100 ml tissue) increased flow from 2.4 +/- 5.0 to 20.6 +/- 5.2 ml/min/100 ml tissue (n = 14; p less than 0.05) and decreased forearm vascular resistance from 42.0 +/- 4.1 to 6.0 +/- 1.4 units (p less than 0.03), a response comparable to that of sodium nitroprusside (0.6 micrograms/min ml tissue). Acetylsalicylic acid (500 mg i.v.) given to block vascular prostacyclin production did not alter the response (n = 14). alpha-Adrenoceptor blockade by phentolamine (12 micrograms/min/100 ml tissue) did not prevent the increase in flow evoked by acetylcholine. In hypertensive patients, the decrease in forearm vascular resistance induced by acetylcholine but not evoked by sodium nitroprusside was reduced as compared with controls (14.5 +/- 3.1 and 6.1 +/- 1.6 units, respectively; n = 8; p less than 0.05). Thus, the vascular effects of acetylcholine in the human forearm circulation are independent of prostaglandins and adrenergic neurotransmission and therefore are most likely to be mediated by EDRF; the acetylcholine-induced release of EDRF is blunted in patients with essential hypertension.

Endothelin-1-induced vasoconstriction in humans. Reversal by calcium channel blockade but not by nitrovasodilators or endothelium-derived relaxing factor.

The vascular effects of endothelin-1 (ET) in humans were investigated by brachial artery infusions of ET into 25 healthy volunteers. Forearm blood flow increased from a mean +/- SD value of 2.3 +/- 1.5 to 2.5 +/- 1.5 ml/min/100 ml forearm tissue (n = 25, p less than 0.05) in response to low dose (0.5 ng/min/100 ml forearm tissue) ET infusion and decreased to 1.78 +/- 1.3 and 1.1 +/- 0.9 ml/min/100 ml forearm tissue (p less than 0.001) during higher dosages (25 and 50 ng/min/100 ml forearm tissue). Sodium nitroprusside (0.6 micrograms/min/100 ml forearm tissue, n = 6), acetylcholine (16 micrograms/min/100 ml forearm tissue, n = 7), nifedipine (6 micrograms/min/100 ml forearm tissue, n = 6), and verapamil (80 micrograms/min/100 ml forearm tissue, n = 6) were infused alone and in combination with ET to evaluate the interactions between ET-induced vasoconstriction and stimulation of vascular muscle cyclic GMP levels by sodium nitroprusside, release of endothelium-derived relaxing factor by acetylcholine, and blockade of voltage-operated calcium channels by nifedipine and verapamil. Neither the vasodilator nor the vasoconstrictor response to ET was influenced by sodium nitroprusside or acetylcholine. In contrast, both calcium antagonists converted ET-induced vasoconstriction (e.g., delta forearm vascular resistance to ET 50 ng/min/100 ml forearm tissue, 151 +/- 100% and 164 +/- 92% in verapamil and nifedipine groups, respectively) to vasodilation (-35 +/- 12% and -21 +/- 16%, p less than 0.05). Our results demonstrate both ET-induced vasodilation (at low dosages) and vasoconstriction (at high dosages) in resistance vessels of normal humans. Blockade of voltage-operated calcium channels prevented ET-induced vasoconstriction and unmasked the vasodilator effect of high ET dosages. In human resistance vessels, blockade of voltage-operated Ca2+ channels but not cyclic GMP-dependent vasodilation may be an effective tool to inhibit ET-induced vasoconstriction.

Antihypertensive therapy augments endothelium-dependent relaxations in coronary arteries of spontaneously hypertensive rats.

BACKGROUND Coronary artery disease is an important complication of hypertension. Therefore, the effects of antihypertensive therapy on the endothelial nitric oxide (NO)/L-arginine pathway and vascular smooth muscle were studied in left anterior descending coronary arteries of Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). Angiotensin II (AT1) receptor antagonists CGP 48369 and valsartan, angiotensin-converting enzyme inhibitor benazepril HCl, and calcium antagonist nifedipine were used as antihypertensive agents. METHODS AND RESULTS Rings were examined in myograph systems for isometric tension recording. In untreated WKY and SHR rings, acetylcholine (10(-9) to 10(-5) mol/L) but not bradykinin, substance P (both 10(-6) mol/L), or thrombin (1 U/mL) induced comparable endothelium-dependent relaxations. These relaxations were markedly decreased by NG-monomethyl-L-arginine (10(-4) mol/L) and fully prevented by N omega-nitro-L-arginine methyl ester (10(-4) mol/L) or methylene blue (10(-5) mol/L). In vitro treatment of WKY and SHR rings with benazeprilat, CGP 48369, or valsartan (3 x 10(-7) mol/L) did not affect responses to acetylcholine. In SHR, chronic therapy for 8 weeks with benazepril HCl, CGP 48369, valsartan, or nifedipine (each 10 mg.kg-1.d-1 PO) similarly reduced blood pressure and increased endothelium-dependent relaxations to acetylcholine (log shift at IC50, ie, half-maximal inhibition of a preceding contraction, 10-, 8-, 13-, and 13-fold, P < .05 versus control), whereas relaxations to the NO donor 3-morpholino sydnonimine (SIN-1, 10(-9) to 10(-5) mol/L) remained unaffected. In WKY, chronic therapy with nifedipine (10 mg.kg-1.d-1 PO) affected neither blood pressure nor relaxations to acetylcholine or SIN-1. CONCLUSIONS In rat coronary arteries, NO is synthesized via the endothelial L-arginine pathway and released after stimulation with acetylcholine. In SHR, chronic antihypertensive therapy with either angiotensin receptor antagonists, an angiotensin-converting enzyme inhibitor, or a calcium antagonist specifically increased the normal endothelium-dependent relaxations to acetylcholine, probably because of their blood pressure-lowering effects, whereas the responsiveness of vascular smooth muscle to NO remained unaffected.

Aging differentially affects direct and indirect actions of endothelin-1 in perfused mesenteric arteries of the rat

1 The effects of age on the vascular action of endothelin‐1 were studied in mesenteric resistance arteries of 4,9 and 27 month old Fischer 344 rats. 2 Third order branches (about 200 μm in diameter) of mesenteric resistance arteries were dissected free and mounted on glass cannulae in organ chambers. Changes in intraluminal diameter of the perfused and pressurized vessels were continuously measured with a video dimension analyzer. 3 Endothelin‐1 (10−14−3 × 10−8 m.) caused contractions that were augmented after removal of the endothelium. The inhibitory effects of the endothelium were greater in young than in old rats. 4 The sensitivity of vascular smooth muscle to endothelin‐1 decreased with age, while the maximal response was maintained. In contrast, the contractions to noradrenaline were unaffected by aging. 5 Threshold concentrations of endothelin‐1 potentiated the contractions evoked by low and moderate concentrations of noradrenaline (10−7−10−6m) in old, but not in young, rats. 6 Endothelium‐dependent relaxations to acetylcholine inhibited maximal contractions to endothelin‐1 and this effect decreased with age. In contrast, the relaxations to the nitric oxide donor, 3‐morpholinosydnonimine (SIN‐1; the active metabolite of molsidomine), did not differ in the three age groups. 7 Aging specifically decreases the direct contractile effects of endothelin‐1 and the inhibitory effects of the endothelium against these contractions, while the indirect (potentiating) effects of the peptide become more pronounced.

Renovascular hypertension impairs formation of endothelium-derived relaxing factors and sensitivity to endothelin-1 in resistance arteries

1 Endothelium‐dependent vascular regulation was investigated in mesenteric resistance arteries of Goldblatt two kidney‐one clip (2K1C) renovascular hypertensive rats. 2 Third order branches of mesenteric arteries were dissected free and mounted on glass cannulae in an organ chamber. Changes in vascular diameter were measured in pressurized and perfused arteries with a video dimension analyzer. 3 Acetylcholine evoked endothelium‐dependent relaxations that were much more pronounced with intraluminal than with extraluminal application. In 2K1C rats the relaxation induced by intraluminal, but not extraluminal acetylcholine was decreased compared to normotensive Wistar Kyoto rats (WKY). Increased duration of hypertension further decreased the response to intraluminal but not extraluminal acetylcholine. 4 Endothelin‐1 and noradrenaline caused contractions which were augmented by removal of the endothelium. This augmentation was reduced in 2K1C rats compared to WKY; the difference was small with noradrenaline but more pronounced with endothelin‐1. 5 In arteries without endothelium the sensitivity, but not the maximal contraction to endothelin‐1 was lower in 2K1C rats, while the response to noradrenaline was not different in 2K1C rats and WKY. The sensitivity to the peptide was not further affected by increasing the duration of hypertension. 6 Thus, renovascular hypertension leads to an impaired intraluminal, but not extraluminal activation of the release of endothelium‐derived relaxing factor and a decreased inhibitory effect of the endothelium against endothelin‐1‐ and noradrenaline‐induced contractions in mesenteric resistance arteries. Furthermore, the sensitivity, but not the maximal response of vascular smooth muscle to endothelin‐1 was reduced.

Effects of calcium channel blockers on the response to endothelin-1, bradykinin and sodium nitroprusside in porcine ciliary arteries*

Calcium channel blockers are increasingly used in ophthalmology, for instance in patients with visual field defects caused by vasospasm. Endothelin is a new vasoactive peptide which also has been implicated in hypoperfusion of the ophthalmic circulation. This study investigated the effects of the calcium channel blockers on the response to endothelin-1, bradykinin and sodium nitroprusside in isolated porcine ciliary arteries (diameter 200-250 microns). Isolated porcine ciliary arteries were suspended in myograph systems filled with modified Krebs-Ringer solution (37 degrees C; 95% O2/5% CO2) for isometric tension recording. Endothelin-1 (10(-12) -10(-7) M) induced potent concentration-dependent contractions of porcine ciliary arteries (PD50 = 8.3 +/- 0.1; n = 7). Lacidipine (10(-5) -10(-7) M) and nifedipine (10(-5) M) significantly reduced the contractions and decreased the sensitivity to endothelin-1 as compared to control (P < 0.03). On the other hand, endothelium-dependent relaxations to bradykinin (10(-10) -10(-6) M) and endothelium-independent relaxations to sodium nitroprusside (10(-10) -10(-4) M) remained unaffected by the calcium channel blocker. These findings demonstrate that in porcine ciliary arteries, the calcium channel blockers selectively inhibit endothelin-1-induced contractions, while leaving endothelium-dependent and endothelium-independent relaxations unaffected. This property of calcium channel blockers may contribute to the clinical efficacy of this class of drugs in patients with ocular vasospasms.