Search

Aims: To determine whether acute insulin-induced hypoglycaemia provokes a detectable alteration in peripheral plasma endothelin (ET) concentrations in humans with Type 1 diabetes., Methods: Serial plasma concentrations of ET were measured in 20 patients with Type 1 diabetes during controlled hypoglycaemia induced by intravenous infusion of soluble insulin., Results: A significant increase was observed in plasma ET concentrations, from 3.80 +/- 0.31 pg/ml at baseline to 6.72 +/- 1.47 pg/ml at 60 min after the onset of the hypoglycaemic reaction (P < 0.05)., Conclusions: Acute insulin-induced hypoglycaemia induces a rise in plasma endothelin concentrations in people with Type 1 diabetes. This finding is consistent with a putative role for ET in the mediation of hypoglycaemia-induced vasoconstriction, and the possible precipitation of macrovascular or microvascular events.

Evidence for interactions between the endothelin (ET) and renin-angiotensin systems is plentiful in vitro, but few studies have investigated these interactions in vivo. In the study reported here, we have investigated the influence of chronic angiotensin-II (A-II) infusion in vivo on expression of preproendothelin-1 (PPET-1) and endothelin-A- (ET(A)) and endothelin-B- (ET(B)) receptor mRNA in the heart. The role of the angiotensin type 1 (AT1)-receptor in mediating the actions of A-II was studied using losartan, the selective AT1-receptor antagonist. Male rats received an infusion of A-II (200 ng/kg/min) or vehicle for 14 days via mini-osmotic pumps; losartan (10 mg/kg/day) was administered in the drinking water. PPET-1 and ET(A)- and ET(B)-receptor mRNA were detected in heart sections using nonradioactive antisense in situ hybridization. Independent treatments with either A-II or losartan had no significant effect on PPET-1, ET(A)- or ET(B)-receptor expression. Combined treatment resulted in an increase in PPET-1 mRNA (p < 0.001) and ET(B)-receptor mRNA expression (p < 0.01), while ET(A)-receptor mRNA expression was decreased (p < 0.001). These results suggest that selective AT1-receptor blockade, in the presence of an elevated plasma A-II concentration, causes upregulation of ET-1 synthesis in the myocardium as well as modification of ET receptor expression. These effects may be mediated via angiotensin type 2 (AT2)-receptors.

Background: Endothelin-3 (ET-3) and its inactive precursor, big endothelin-3 (big ET-3), are both found in human plasma. We investigated whether big ET-3 is converted to ET-3 in the human forearm resistance vessels and dorsal hand veins in vivo., Methods: In a 4-phase study, 6 subjects received 90 minute intrabrachial artery infusions of big ET-3 (50 and 100 pmol x min(-1)) and ET-3 (5 and 10 pmol x min(-1)) in random order. Forearm blood flow was measured by venous occlusion plethysmography. In a second 3-phase study, 6 subjects received 90-minute dorsal hand vein infusions of saline solution, big ET-3 (50 pmol x min(-1)) and ET-3 (5 pmol x min(-1)) in random order. In a third 2-phase study, 6 subjects received 90-minute dorsal hand vein infusions of big ET-3 (100 pmol x min(-1)) and ET-3 (10 pmol x min(-1)). In the dorsal hand vein studies, vessel diameter was measured by the Aellig technique., Results: Intra-arterial ET-3 caused local forearm vasoconstriction of 20%+/-9% (P = .009) at 5 pmol x min(-1) and 20%+/-10% (P = .001) at 10 pmol x min(-1) after 90 minutes, with no difference between doses (P = .69). Intra-arterial big ET-3 also caused local forearm vasoconstriction of 22%+/-6% at 50 pmol x min(-1) (P = .004) and 18%+/-3% at 100 pmol x min(-1) (P<.0001) after 90 minutes, with no difference between doses (P = .44). There were no significant differences between the responses to intra-arterial big ET-3 and ET-3 at these doses. Local intravenous ET-3 caused a constriction of 9%+/-2% at 5 pmol x min(-1) (P = .04) and 22%+/-8% at 10 pmol x min(-1) (P = .002) after 90 minutes. Big ET-3 at 50 pmol x min(-1) and 100 pmol x min(-1) did not affect hand vein tone. All responses were slowly progressive., Conclusions: Based on vasoconstriction, measurable conversion of big ET-3 to ET-3 occurs in forearm resistance vessels but not in dorsal hand veins in vivo. An endothelin-converting enzyme, capable of converting exogenously administered big ET-3 to ET-3, appears to be present in upper limb resistance arteries but not in capacitance vessels in humans.

Aims: The aim of the study was to examine the effects of the ETB receptor selective agonists sarafotoxin S6c (SFTX6c) and BQ-3020 on the forearm resistance and capacitance vessels in healthy subjects in vivo., Methods: The local response to intra-arterial or intravenous infusion of SFTX6c (5 pmol min-1) or BQ-3020 (50 pmol min-1) was assessed, on separate occasions, in eight healthy men (aged 20-28 years). Data (mean +/- s.e.mean) were examined by ANOVA. Results are expressed as percentage change from baseline at 90 min., Results: SFTX6c and BQ-3020 reduced forearm blood flow, following local intra-arterial infusion (-25 +/- 7% and -27 +/- 7%, respectively; P < 0.001) and reduced hand vein diameter, following local intravenous infusion (-30 +/- 8% and -16 +/- 7%, respectively; P < 0.001)., Conclusions: We have shown that locally active infusions of the selective ETB receptor agonists SFTX6c and BQ-3020 cause arterial constriction and venoconstriction in healthy human blood vessels in vivo. These results indicate that ETB receptor stimulation may mediate vasoconstriction in humans.

Since its discovery in 1988, endothelin (ET) has been widely implicated in the pathophysiology of cardiovascular disease. ET antagonists have favourable effects in experimental models of these conditions and have proved useful in elucidating the role of the ET system. Orally acting ET antagonists appear very promising in clinical trials, particularly in patients with chronic heart failure and hypertension, but more information on the roles of the ET receptor subtypes in health and disease is required so that an informed choice can be made between the use of endothelin-A (ET-A) receptor-selective and nonselective receptor antagonists.

The endothelins are a family of endothelium-derived peptides that possess characteristically sustained vasoconstrictor properties. Endothelin-1 appears to be the predominant member of the family generated by vascular endothelial cells. In addition to its direct vascular effects, endothelin-1 has inotropic and mitogenic properties, influences homeostasis of salt and water, alters central and peripheral sympathetic activity and stimulates the renin-angiotensin-aldosterone system. Studies with endothelin receptor antagonists have indicated that endothelin-1 probably has complex opposing vascular effects mediated through vascular smooth muscle and endothelial ET(A) and ET(B)receptors. Endogenous generation of endothelin-1 appears to contribute to maintenance of basal vascular tone and blood pressure through activation of vascular smooth muscle ET(A)receptors. At the same time, endogenous endothelin-1 acts through endothelial ET(B) receptors to stimulate formation of nitric oxide tonically and to oppose vasoconstriction. In view of the multiple cardiovascular actions of endothelin-1, there has been much interest in its contribution to the pathophysiology of hypertension. Results of most studies suggest that generation of, or sensitivity to, endothelin-1 is no greater in hypertensive than it is in normotensive subjects. Nonetheless, the deleterious vascular effects of endogenous endothelin-1 may be accentuated by reduced generation of nitric oxide caused by hypertensive endothelial dysfunction. It also appears likely that endothelin participates in the adverse cardiac and vascular remodelling of hypertension, as well as in hypertensive renal damage. Irrespective of whether vascular endothelin activity is increased in hypertension, anti-endothelin agents do produce vasodilatation and lower blood pressure in hypertensive humans. There is more persuasive evidence for increased endothelin-1 activity in secondary forms of hypertension, including pre-eclampsia and renal hypertension. Endothelin-1 also appears to play an important role in pulmonary hypertension, both primary and secondary to diseases such as chronic heart failure. The hypotensive effects of endothelin converting enzyme inhibitors and endothelin receptor antagonists should be useful in the treatment of hypertension and related diseases. Development of such agents will increase knowledge of the physiological and pathological roles of the endothelins, and should generate drugs with novel benefits.

Endothelin-1, discovered in 1988, is a 21-amino-acid peptide and currently the most potent vasoconstrictor and pressor substance known. Generated by vascular endothelial cells in response to a variety of chemical and mechanical signals, endothelin-1 is known to potentiate the actions of other vasoconstrictor substances and act as a comitogen in addition to directly causing vasoconstriction. There is evidence that endothelin-1 may contribute to the pathophysiology of conditions associated with sustained vasoconstriction, such as hypertension and heart failure, vasospastic conditions, such as subarachnoid hemorrhage, and atherogenesis. Studies using endothelin receptor antagonists show that endothelin-1 plays an important role in the maintenance of vascular tone and blood pressure in healthy humans, predominantly via an effect on the vascular smooth muscle ETA receptors. The endothelin receptor antagonist bosentan also effectively lowers blood pressure in hypertensive subjects and produces sustained and favorable effects on systemic and pulmonary hemodynamics in patients with chronic heart failure. A good side-effect profile, together with a potential for inhibition of atherogenesis, makes the endothelin receptor antagonists a potentially interesting class of novel agents for the treatment of cardiovascular disease.

Endothelin-1 (ET-1) has been implicated in the pathogenesis of Raynaud's disease (RD). This study examined the effect of cooling on the response to ET-1 in human microvessels. Subcutaneous small arteries were dissected from gluteal fat biopsies taken from patients with RD (n = 20) and from age- and sex-matched control subjects (n = 17) and were cannulated in a small vessel arteriograph. Cumulative concentration-response curves to ET-1 (10(-12) to 3 x 10(-7) M) were obtained in vessels at 37 degrees C and 24 degrees C, with the endothelium either intact or removed (n = 6 per group). There were no significant differences in responses to ET-1 between RD patients and controls in either intact or denuded vessels, at either 37 degrees C or at 24 degrees C. There was, however, a significant endothelium-dependent interaction between the groups when the effect of temperature on the response to ET-1 was examined (p = 0.01; two-way ANOVA). Whereas cooling tended to reduce the sensitivity in RD, the opposite effect was observed in controls. Measurements of plasma ET-1 did not reveal any significant difference between patients with RD and healthy controls. These results suggest that ET-1 does not play a primary pathophysiologic role in RD. ET-1 might be responsible for mediating the prolonged vasospasm in RD, but secondary to another factor(s), such as impaired endothelium-dependent vasodilation.

Based on studies with a number of endothelin antagonists, endothelin-1 is now known to play a key physiological role in the maintenance both of basal systemic vascular resistance and of blood pressure in healthy people. Factors that regulate activity of the vascular endothelin system, and its physiological effects on the heart and kidney, now need to be determined.

Endothelin-1, a potent endothelium-derived 21-amino acid vasoconstrictor peptide was only discovered relatively recently. Antagonism of its actions may provide an important new approach to the modulation of cardiac and vascular function.

1. We examined the effects of systemic infusion, in healthy human volunteers, of the endothelin antagonist TAK-044 on the plasma concentrations of mature endothelin, big endothelin-1 and the C-terminal fragment of big endothelin-1, by selective solid-phase extraction and specific radioimmunoassays. 2. Unlabelled TAK-044 competed with specific [125I]-endothelin-1 binding to human left ventricle tissue in a biphasic manner giving KD values of 0.11 nM and 26.8 nM at the ETA and ETB receptor subtypes, respectively, indicating a 244 fold selectivity for the ETA receptor subtype. 3. A 15 min intravenous infusion of placebo or 30 mg TAK-044 (giving a serum concentration of 2 nM, calculated to block > 95% of ETA but < 5% ETB receptors) had no effect on the immunoreactive plasma concentrations of the three peptides. 4. At the higher dose of 750 mg TAK-044 (giving a serum concentration of 80 nM, calculated to block > 99% of ETA and > 75% ETB receptors), the immunoreactive plasma endothelin concentrations were increased 3.3 fold over basal levels (P < 0.01). The concentrations of big endothelin-1 or C-terminal fragment of big endothelin-1 were unchanged. 5. At both doses of TAK-044, there were significant decreases in diastolic blood pressure, and peripheral vascular resistance, with corresponding increases in cardiac index and stroke index. There were no changes in systolic or mean arterial blood pressures or heart rate. 6. Since only the concentrations of the mature peptide were increased, we conclude that the most likely sources of endothelin contributing to the observed rise were displacement of receptor-bound peptide and reduction in plasma clearance rather than peptide synthesis.

1. Endothelin-1 is a potent endothelium-derived vasoconstrictor peptide that is generated through cleavage of its precursor big endothelin-1 by 'endothelin converting enzyme' (ECE) in resistance vessels, including those of the forearm vascular bed. In some animal tissues, but not in resistance vessels of healthy human subjects, endothelin-1 appears to potentiate the actions of the sympathetic nervous system. We examined whether ECE activity is present in human hand veins and whether endothelin-1 or big endothelin-1 potentiate sympathetically mediated venoconstriction. 2. Six healthy subjects received dorsal hand vein infusion of local, non-systemic doses of endothelin-1 (5 pmol min-1), big endothelin-1 (50 pmol min-1) and, as a control, sodium chloride (0.9%; w/v) for 90 min. Vein diameter was measured using the Aellig displacement technique. Sympathetically mediated venoconstriction was elicited using the single deep breath reflex. 3. Endothelin-1 caused a progressive decrease in hand vein diameter, by 49% at 90 min (95% confidence intervals [CI]: -68 to -30%; P = 0.0001). Vein diameter did not change significantly after 90 min infusion of big endothelin-1 (+3%; CI: -11 to +17%; P = 0.0007 vs endothelin-1; P = 0.40 vs baseline) or sodium chloride (+2%; CI: -12 to +16%; P = 0.0002 vs endothelin-1; P = 0.60 vs baseline). Venoconstriction to deep breath was not potentiated by endothelin-1. 4. These results suggest that, in contrast to the situation in forearm resistance vessels, there is little or no local ECE activity in human hand veins and that endothelin does not potentiate sympathetic responses in these cutaneous capacitance vessels.

1. The vasoconstrictor peptide, endothelin-1 (ET-1) and a biologically inactive C-terminal fragment (CTF) are generated from an intermediate big ET-1 by a putative ET converting enzyme, sensitive to phosphoramidon. We have developed a procedure using selective solid-phase extraction and specific radioimmunoassays to measure the levels of immunoreactive (IR) big ET-1 and the products of conversion (ET-1 and CTF) in human plasma. These techniques have been used to determine the levels of the three peptides in venous plasma following local infusions of ET-1 and big ET-1, both alone and together with phosphoramidon. 2. Infusion of ET-1 into the brachial artery (5 pmol min-1) significantly increased (P < 0.05) IR ET levels from a basal level of 2.3 pM to 55.2 pM in plasma from the infused arm after 60 min of infusion. This corresponded with a marked decrease in forearm blood flow from a basal level of 2.6 ml dl-1 min-1 to 1.7 ml dl-1 min-1. The levels of IR big ET-1 and CTF were unchanged. Co-infusion of phosphoramidon (30 nmol min-1) with ET-1 had no significant effect on the plasma IR levels of ET, big ET-1, CTF, or blood flow. 3. Big ET-1 (50 pmol min-1) significantly increased (P < 0.05) venous concentrations of all three IR peptides after 60 min compared to basal (ET: from 2.2 to 7.7 pM, big ET-1; from 0 to 386.0 pM, CTF: from 0.2 to 37.0 pM). Forearm blood flow decreased significantly (P<0.05) from a basal level of 3.0 ml dl-1 min-1 to 1.6 ml dl-1 min-1.4. When phosphoramidon was co-infused with big ET-1, both the rise in IR ET and associated vasoconstriction were abolished. However, IR CTF was still detected, suggesting that either some conversion by phosphoramidon-insensitive enzyme(s) was occurring, and/or that CTF was being protected from further degradation by phosphoramidon.5. These data show that in the human forearm the activity of a phosphoramidon-sensitive ET converting enzyme is at least in part responsible for the vasoconstrictor properties of exogenous big ET-1. Furthermore, because measurable levels of newly synthesized ET-1 are likely to be rapidly reduced in the blood/plasma through receptor binding, assay of IR big ET-1 and CTF may be a more sensitive measure of ET-1 generation in disease.

Background: The role of endothelin ETB receptors in mediating vasoconstriction in humans is unclear. As yet, there have been no in vivo studies in resistance vessels, and in vitro data have been contradictory. We therefore investigated the function of ETB receptors in vivo in human forearm resistance and hand capacitance vessels using endothelin-1 as a nonselective agonist at ETA and ETB receptors and endothelin-3 and sarafotoxin S6c as selective agonists at the ETB receptor., Methods and Results: A series of single-blind studies were performed, each in six healthy men. Brachial artery infusion of endothelin-1 and endothelin-3 caused slow-onset dose-dependent forearm vasoconstriction. Although endothelin-3 caused significantly less forearm vasoconstriction than endothelin-1 at low doses, vasoconstriction was similar to the two isopeptides at the highest dose (60 pmol/min). Endothelin-3 caused transient forearm vasodilatation at this dose, whereas endothelin-1 showed only a nonsignificant trend toward causing early vasodilatation. Intra-arterial sarafotoxin S6c caused a progressive reduction in forearm blood flow, although less than that to endothelin-1 (P = .04). Dorsal hand vein infusion of sarafotoxin S6c caused local venoconstriction that was also less than that to endothelin-1 (P = .002)., Conclusions: Selective ETB receptor agonists cause constriction of forearm resistance and hand capacitance vessels in vivo in humans, suggesting that both ETA and ETB receptors mediate vasoconstriction. Hence, antagonists at both ETA and ETB receptors, or inhibitors of the generation of endothelin-1, may be necessary to completely prevent vasoconstriction to endogenously generated endothelin-1.

Objective: To elucidate the sequential mechanisms underlying cyclosporin-induced hypertension and nephrotoxicity., Design: A study of healthy males over the first 9 days of drug ingestion to permit the detection of serial changes in renal function and blood pressure in a situation free from the confounding variables of concomitant disease or drugs., Methods: Double-blind, placebo-controlled, randomized crossover study with cyclosporin (5 mg/kg twice a day) or placebo. Blood pressure and urinary sodium excretion were measured each day, and glomerular filtration rate (GFR) and effective renal plasma flow (ERPF) were measured on days 1, 4, 7 and 9. Cholesterol, lipoprotein (a) and endothelin were measured on days 1 and 9., Results: GFR decreased by 9% with cyclosporin and was significantly lower than with placebo on day 4 of therapy. ERPF fell by 24%. The fall in GFR correlated significantly with suppressed plasma renin activity (P < 0.0001). Cyclosporin-induced hypertension occurred in the absence of any change in urinary sodium output or in plasma endothelin. Cyclosporin did not affect lipoprotein (a) levels during 9 days of cyclosporin therapy., Conclusions: Cyclosporin-induced hypertension and renal vasoconstriction are well established after 9 days of cyclosporin 5 mg/kg twice a day. We found no evidence to implicate either circulating endothelin or renal sodium retention in the onset of cyclosporin-induced hypertension. Cyclosporin-induced renal vasoconstriction appeared to occur when the protective mechanism of plasma renin activity suppression became exhausted.

Endothelin-1 is a potent endothelium-derived vasoconstrictor and pressor peptide with uniquely sustained activity. We have examined the contribution of endogenously-generated endothelin-1 to the maintenance of basal vascular tone in healthy subjects. In these studies, on separate occasions, a combined inhibitor of endothelin converting enzyme (ECE) and neutral endopeptidase (NEP), phosphoramidon, a selective inhibitor of NEP, thiorphan, and a selective ETA receptor antagonist, BQ-123, were given via the brachial artery. Big endothelin-1, the precursor to endothelin-1, caused a slow onset dose-dependent forearm vasoconstriction, the magnitude of which was consistent with about 10% conversion to mature endothelin-1 in the forearm. Vasoconstriction to big endothelin-1 was abolished by co-infusion of phosphoramidon, whereas vasoconstriction to endothelin-1 was unaffected. Phosphoramidon caused progressive vasodilatation when infused alone, with blood flow increasing by 37% at 90 min (P = 0.02), whereas thiorphan caused vasoconstriction, consistent with NEP inhibition exerting its major effect on degradation of constrictor peptides, such as angiotensin and endothelin-1. Vasoconstriction to endothelin-1 was completely abolished by coinfusion of BQ-123, and BQ-123 alone produced progressive forearm vasodilatation, with blood flow increasing by 64% after 60 min (P = 0.001). These results demonstrate that endogenous production of endothelin-1 acts to sustain vascular tone in humans and indicate that ECE inhibitors and endothelium receptor antagonists may have therapeutic potential as vasodilators.

The role of endothelin (ET)-B (ETB) receptors in mediating vasoconstriction in humans is unclear. As yet, in vitro data have been contradictory, and there have been no in vivo studies in resistance vessels. We investigated the function of ETB receptors in vivo in human forearm resistance vessels using ET-1 as a nonselective agonist at ETA and ETB receptors and ET-3 and sarafotoxin S6c as ETB receptor agonists. Brachial artery infusion of ET-1 and ET-3 caused slow-onset, dose-dependent forearm vasoconstriction. Although ET-3 caused significantly less forearm vasoconstriction than ET-1 at low doses, vasoconstriction to the two isopeptides was similar at the highest dose (60 pmol/min). ET-3 caused initial transient forearm vasodilatation at this dose, whereas ET-1 showed only a nonsignificant trend toward causing early vasodilatation. Intra-arterial sarafotoxin S6c caused a progressive reduction in forearm blood flow, although less than that to ET-1. Therefore, ETB receptor agonists contract human resistance vessels in vivo. The effects of ET-3 and sarafotoxin S6c, compared with ET-1, suggest that both ETA and ETB receptors mediate vasoconstriction. Antagonists at both ETA and ETB receptors, or inhibitors of the generation of ET-1, may be necessary to completely prevent vasoconstriction to endogenously generated ET-1.

We developed a procedure for individual measurement of big endothelin-1 (big ET-1) and the two products of big ET-1 conversion (ET-1 and C-terminal fragment (CTF) of big ET-1 (big ET-1(22-38)), using selective solid-phase extraction and specific radioimmunoassays. These techniques were used to measure the levels of these peptides in extracts of human plasma after brachial artery infusions of big ET-1. Infusion of big ET-1 (50 pmol/min, n = 6) caused a decrease in forearm blood flow from 3.03 to 1.55 ml/dl/min after 60 min (p < 0.05). In agreement, the levels of plasma immunoreactive (IR) big ET-1, ET, and CTF were significantly increased from basal levels in the infused arm (from undetectable to 386 pM, 2.2-7.5 pM, and 0.17-37 pM, respectively; p < 0.05). In the presence of the metalloprotease inhibitor phosphoramidon (30 nmol/min), the increase in IR-ET and the associated vasoconstriction were abolished. However, IR-CTF was still detected, suggesting that either some conversion by phosphoramidon-insensitive endothelin-converting enzyme (ECE) was occurring and/or that CTF was being preserved from further proteolysis by phosphoramidon. These data confirm that exogenous big ET-1 is locally converted to ET-1 and CTF in the human forearm, at least in part by a phosphoramidon-sensitive ECE. Furthermore, because measurable levels of newly synthesized ET-1 are probably rapidly reduced as a result of receptor binding, the assay of IR-CTF may be a more sensitive measure of the overexpression of ET-1 in disease.

The physiologic significance of endothelin-1 (ET-1) generation in human resistance vessels is unknown. We therefore investigated whether endothelin-converting enzyme (ECE) activity could be demonstrated in human vessels, and the effects of inhibition of the generation or actions of ET-1 on vascular tone in healthy men. Brachial artery infusion of local doses of big ET-1 caused a slow-onset, dose-dependent forearm vasoconstriction that was abolished by co-infusion of the ECE inhibitor phosphoramidon. Phosphoramidon did not affect responses to ET-1. Phosphoramidon caused slow-onset vasodilatation when infused alone, with blood flow increasing by 37% (p = 0.03). Vasoconstriction to ET-1 was completely abolished by co-infusion of the ETA receptor antagonist BQ-123 (p = 0.006), with forearm blood flow tending to increase. Infusion of BQ-123 alone resulted in progressive vasodilatation, with blood flow increasing by 64% (p = 0.007). These results suggest that endogenous generation of ET-1 contributes to the maintenance of vascular tone in states of normal and elevated blood pressure. ECE inhibitors and ETA receptor antagonists may have potential as vasodilators in the treatment of diseases associated with vasoconstriction, such as hypertension and chronic heart failure.

The pathogenesis of the prolonged cold-induced vasospasm of Raynaud's disease (RD) is unknown. Cooling has been shown to decrease the sensitivity of cutaneous rabbit ear arteries to endothelin-1 (ET-1), perhaps by increasing the availability of nitric oxide (NO). If the endothelium in RD lacks this NO-mediated inhibitory function during cooling, increased production and enhanced constriction to ET-1 could cause vasospasm. This study examined the effect of cooling on the contractile response to ET-1 in human microvessels. Small arteries were dissected from biopsy specimens of human fat and were cannulated in a small vessel arteriograph. Cumulative concentration-response curves to ET-1 (10(-12) to 3 x 10(-7) M) were obtained in vessels at 37 degrees C (n = 8) and 24 degrees C (n = 7). Cooling to 24 degrees C resulted in an eightfold decrease in sensitivity to ET-1 (EC50 6.6 +/- 2.5 x 10(-10) M at 37 degrees C vs. 5.5 +/- 2.5 x 10(-9) M at 24 degrees C; p < 0.05, unpaired t test). The Emax was not significantly different at 37 degrees C and 24 degrees C (114 +/- 9 at 37 degrees C vs. 138 +/- 14 at 24 degrees C; p = 0.18). These results suggest that cooling decreases the sensitivity of human microvessels to ET-1. Further investigation is needed to establish the mechanism of this effect, including studies in vessels obtained from patients with RD.

Background: Among patients with independent evidence of coronary disease and recent onset unstable angina or non-Q wave myocardial infarction the incidence of subsequent cardiovascular events is high. Markers predictive of adverse cardiac outcome in unstable angina and non-Q wave myocardial infarction need to be defined more accurately. Endothelin-1 is a potent endothelium derived vasoconstrictor peptide that may play a part in the pathophysiology of acute myocardial ischaemia. AIM AND STUDY DESIGN: In a study that specifically identified high risk patients a group of 16 consecutive patients with either unstable angina at rest or non-Q wave myocardial infarction were prospectively investigated to establish whether these conditions are associated with high plasma immunoreactive endothelin and whether endothelin concentration at presentation is related to cardiovascular events within the next 12 weeks. Controls consisted of a group of 40 healthy subjects., Results: Patients had significantly higher mean (SD) plasma endothelin at presentation than did healthy controls (7.4 (1.1) v 5.0 (1.2) pg/ml, P < 0.0001). At nine weeks plasma endothelin was still significantly higher in those patients who had subsequent cardiovascular events, (n = 9, acute myocardial infarction or refractory angina with electrocardiographic changes and revascularisation procedures, 8.5 (2.6) pg/ml, P < 0.005 v controls) whereas its concentration returned to normal in those patients who had a favourable outcome (n = 7, 5.9 (0.7) pg/ml). Compared with those patients who had an uneventful course, patients with subsequent events had significantly higher plasma endothelin, both at presentation and at nine weeks (P < 0.05 on both occasions)., Implications: Endothelin may contribute to the pathophysiology of acute coronary syndromes and may relate to subsequent cardiovascular outcome.

Endothelin-1 is a potent endothelium-derived vasoconstrictor peptide. Although circulating concentrations are not increased in essential hypertension, enhanced sensitivity to endothelin-1 has been observed in animal models of hypertension. We investigated dorsal hand vein responses to local infusion of endothelin-1 and norepinephrine in 12 patients with essential hypertension who had never received treatment and in 12 age and sex matched normotensive control subjects. The maximal venoconstriction and the geometric mean of the dose of norepinephrine that caused 50% of maximal venoconstriction were similar in hypertensive (mean +/- SE; 80 +/- 4%; 31 +/- 8 pmol/min) and normotensive subjects (87 +/- 5%, 22 +/- 9 pmol/min). In contrast, mean venoconstriction to endothelin-1 was significantly greater in hypertensive (49 +/- 5%) than in normotensive subjects (27 +/- 2%; P = 0.004). Sympathetically mediated venoconstriction elicited by deep breath was substantially potentiated by endothelin-1 in hypertensive (67 +/- 7% at 90 min) but not normotensive subjects (11 +/- 3% at 90 min; P = 0.001). Venoconstriction to endothelin-1 correlated positively with mean arterial pressure in the hypertensive subjects (r = 0.82; p = 0.001) but negatively in the normotensive subjects (r = -0.58; p = 0.047). Endothelin-1 may contribute to the reduction of venous compliance occurring in the early stages of essential hypertension and to the altered systemic hemodynamics in this condition.

Endothelin-1 is an endothelium-derived vasoconstrictor peptide, possibly involved in the pathophysiology of cardiovascular disease. We examined the contribution of endogenously generated endothelin-1 to maintenance of peripheral vascular tone in healthy subjects by local intraarterial administration of an inhibitor of endothelin converting enzyme, phosphoramidon, and of a selective endothelin receptor A antagonist, BQ-123. Brachial artery infusion of local doses of proendothelin-1, the precursor to endothelin-1, caused a slow-onset dose-dependent forearm vasoconstriction which was abolished by co-infusion of phosphoramidon. Phosphoramidon did not affect responses to endothelin-1. Phosphoramidon caused slow-onset vasodilatation when infused alone, with blood flow increasing by 37% at 90 min (p = 0.03). Vasoconstriction to endothelin-1 was abolished by co-infusion of BQ-123 (p = 0.006), with forearm blood flow tending to increase. Infusion of BQ-123 alone caused progressive vasodilatation, with blood flow increasing by 64% after 60 min (p = 0.007). These results show that endogenous production of endothelin-1 contributes to the maintenance of vascular tone. Endothelin converting enzyme inhibitors and receptor antagonists may have therapeutic potential as vasodilators.

Cardiac arrest is associated with major metabolic disturbances, including severe hypoxia and large increases in circulating catecholamines, both of which are known to stimulate generation of the potent endothelium-derived vasoconstrictor peptide endothelin-1. We have, therefore, examined plasma immunoreactive endothelin concentrations following cardiac arrest. Blood was sampled at 10-min intervals from a central venous catheter inserted at onset of resuscitation in 38 patients (13 female; mean age, 67 years) presenting with cardiac arrest to the Accident and Emergency Department at the Royal Infirmary of Edinburgh. Plasma immunoreactive endothelin concentrations (mean +/- S.D.) in patients following cardiac arrest (5.4 +/- 2.3 pg/ml) were no different from those in healthy subjects (5.1 +/- 1.2 pg/ml). There was no significant difference between endothelin concentrations at presentation in survivors and non-survivors of cardiac arrest. However, non-survivors had a significant fall in endothelin concentrations with time from onset of resuscitation from 5.4 +/- 2.2 pg/ml to 3.5 +/- 1.8 pg/ml (P = 0.002), while survivors had a non-significant increase in concentrations. On multiple regression analysis there was a significant association between higher plasma endothelin concentration and survival (r = 0.37; P = 0.009). The failure of plasma endothelin to increase after cardiac arrest is unexpected. Although the fall in plasma endothelin with time in non-survivors may reflect the adverse physiological milieu that occurs during cardiac arrest, it is also possible that low endothelin concentrations contribute to the poor prognosis in this condition.

Recent studies in vitro have suggested that there may be an interaction between endothelin-1 and ATP-sensitive K+ channels in vascular smooth muscle. Here we have investigated whether agents acting on membrane Ca2+ and K+ channels modulate endothelin-1-induced venoconstriction in vivo in human subjects. In a series of studies, six healthy subjects received, on separate occasions, local infusions into dorsal hand veins of endothelin-1 coinfused with 1) the ATP-sensitive K+ channel opener, cromakalim; 2) the dihydropyridine Ca2+ antagonist, nicardipine; 3) a control vasodilator, hydralazine; and 4) saline placebo. Endothelin-1 caused local venoconstriction with a maximum reduction in vein size of 66 +/- 4% at 60 min (P = 0.0001 vs. basal). Cromakalim prevented endothelin-1-induced venoconstriction (9 +/- 10% maximum constriction; P = 0.68 vs. basal). By contrast, nicardipine, in a dose sufficient to block depolarization-induced constriction caused by K+ infusion, had only a partial effect on endothelin-1-induced venoconstriction (35 +/- 8% maximum constriction; P = 0.001 vs. basal; P = 0.02 vs. endothelin-1), whereas a 10-fold higher dose of nicardipine had no additional effect and hydralazine had no effect. In further studies, cromakalim, but not nicardipine, reversed endothelin-1-induced venoconstriction. Cromakalim did not prevent constriction induced by norepinephrine. Although calcium entry through dihydropyridine-sensitive Ca2+ channels may account in part for the vasoconstrictor action of endothelin-1 in humans, the abolition of endothelin-1 responses by a K+ channel opener suggests additional mechanisms of action for endothelin-1.

1. We have investigated whether local vascular production of nitric oxide or prostacyclin regulates venoconstriction induced by the endothelium-derived peptide, endothelin-1, in vivo in man. 2. Six healthy subjects received local dorsal hand vein infusion of endothelin-1 for 60 min alone or, on two separate occasions, co-infused with the donator of nitric oxide, glyceryl trinitrate, or the vasodilator prostaglandin, prostacyclin. In further studies, endothelin-1 was co-infused with an inhibitor of nitric oxide production, NG-monomethyl-L-arginine, or after oral administration of the irreversible inhibitor of prostaglandin production, acetylsalicylic acid (aspirin). 3. At a low dose (5 pmol/min), endothelin-1 alone caused slowly developing and long-lasting venoconstriction (maximal constriction: 66 +/- 4%). Although glyceryl trinitrate partially prevented endothelin-1-induced venoconstriction (maximum: 33 +/- 5%), inhibition of nitric oxide production did not affect endothelin-1-induced venoconstriction (maximum: 55 +/- 4%). 4. Prostacyclin was more effective at blocking the venoconstriction in response to endothelin-1 than glyceryl trinitrate (maximum: 12 +/- 3%), and there was substantial potentiation of endothelin-1-induced venoconstriction after pretreatment with aspirin (maximum: 90 +/- 3%). 5. Despite the capacity of nitric oxide to attenuate responses to endothelin-1, NG-monomethyl-L-arginine did not potentiate endothelin-1-induced venoconstriction, suggesting little or no stimulated production of nitric oxide in human veins. However, the potentiation of responses to endothelin-1 by aspirin indicates that endothelial production of prostacyclin attenuates responses to endothelin-1 in human veins in vivo.

Endothelin-1 (ET-1) is known to be a potent and long-lasting constrictor of arteries and veins. We investigated whether local endothelial production of nitric oxide (NO) or prostacyclin modulates venoconstriction induced by the endothelium-derived peptide ET-1 in vivo in humans. Six healthy volunteers each received local dorsal hand vein infusion of ET-1 (5 pmol/min) for 60 min in five separate studies: once given alone; on three occasions co-infused with either the NO donor glyceryl trinitrate (GTN), the vasodilator prostaglandin, prostacyclin, or the inhibitor of nitric oxide synthase (NOS) NG-monomethyl-arginine (L-NMMA); and once given 30 min after oral administration of the irreversible inhibitor of cyclooxygenase, acetylsalicylic acid (aspirin). ET-1 alone caused slowly developing and long-lasting venoconstriction (maximal constriction 66 +/- 4%). Although GTN partially prevented ET-1-induced constriction (maximum 33 +/- 5%, p = 0.004 versus ET-1), inhibition of NOS did not affect ET-1-induced venoconstriction (maximum 55 +/- 4%). Prostacyclin was more effective at blocking the venoconstriction to ET-1 than GTN (maximum 12 +/- 3%, p = 0.0001) and there was substantial potentiation of ET-1-induced venoconstriction after pretreatment with aspirin (maximum 90 +/- 3%, p = 0.001). Despite the capacity of NO to attenuate responses to ET-1, L-NMMA did not potentiate ET-1-induced venoconstriction, suggesting little or no stimulated production of NO by ET-1 in human hand veins. However, substantial potentiation of ET-1-induced venoconstriction by aspirin indicates that endothelial production of prostacyclin modulates responses to ET-1 in human veins in vivo.

Endothelin-1 (ET-1) is a potent vasoconstrictor peptide produced by endothelial cells. We investigated whether ET-1, like other potent endothelium-derived vasoactive agents, interacts directly with human platelets in vitro. Platelet-rich plasma was obtained from healthy male volunteers and incubated with ET-1 (1 microM) or vehicle (sodium chloride 154 mM) for 10 min at 37 degrees C. Platelet aggregation was measured by the Born method, using light transmittance through the plasma sample as an index of activation. Although a significant increase in light transmittance was observed when plasma was incubated with ET-1 compared with vehicle, (3.8 +/- 0.4% versus 2.7 +/- 0.2%; n = 24; p = 0.038), this effect was small and is unlikely to be of biologic significance. To investigate the possibility that ET-1-stimulated platelet nitric oxide (NO) synthesis might be masking a direct aggregatory effect of ET-1, in a second study in six subjects NG-monomethyl-L-arginine (L-NMMA, 10 and 100 microM), an inhibitor of NO synthase, was preincubated with the plasma before the addition of ET-1 (1 nM and 1 microM). No significant difference was observed whether samples were incubated with L-NMMA alone or with L-NMMA and ET-1. The results of this study suggest that ET-1 does not have a major direct effect as a platelet aggregating agent.

Endothelin-1 (ET-1) causes long-lasting vasoconstriction associated with a prolonged elevation of intracellular free Ca2+. Because this may be mediated through an effect on membrane ion channels, we investigated the effects of the dihydropyridine calcium channel antagonist nifedipine; two structurally distinct K+ channel openers, pinacidil and levcromakalim; and the inactive stereoisomer of levcromakalim (D-cromakalim), as well as ET-1 and ET-3, on binding of 125I-labeled endothelin-1 to rat cardiac membranes. Specific binding of 125I-ET-1 was inhibited in a concentration-dependent manner by unlabeled ET-1 (IC50 = 1.56 +/- 0.78 nM; slope = -0.49 +/- 0.10) and ET-3 (IC50 = 314 +/- 54 nM; slope = -0.34 +/- 0.11). Nifedipine, in concentrations < or = 10(-5) M, did not affect 125I-ET-1 binding. However, levcromakalim significantly inhibited 125I-ET-1 binding (maximum binding 49 4/- 9%; p = 0.04), whereas the inactive isomer, D-cromakalim, had no effect. Pinacidil also inhibited 125I-ET-1 binding, although to a lesser extent than levcromakalim (maximum binding 63 +/- 7%). These findings provide evidence for a stereospecific interaction between K(+)-channel openers and ET-1 binding in rat cardiac membranes. Because the slope of the logistic fit was substantially less than unity, and the effects of pinacidil and levcromakalim were incomplete, there may be two or more receptors for ET-1 in rat heart, only one of which is sensitive to K(+)-channel openers.

Binding of [125I]-labelled endothelin-1 (ET-1) to rat cardiac membranes and the effects of endothelin-1 (ET-1), endothelin-3 (ET-3), the calcium channel antagonist nifedipine, and both enantiomers of the potassium channel opener cromakalim (BRL 34915) on binding have been examined. Specific binding of [125I]-ET-1 was inhibited in a concentration dependent manner by both unlabelled ET-1 (10(-12)-10(-7) M) and ET-3 (10(-12)-10(-6) M). Nifedipine (10(-11)-10(-5) M) did not affect [125I]-ET-1 binding. However, BRL 38227 (10(-11)-10(-5) M), the biologically active isomer of cromakalim, significantly inhibited [125I]-ET-1 binding. The inactive isomer, BRL 38226 (10(-11)-10(-5) M) had no effect. These findings provide the first evidence for a stereospecific interaction between BRL 38227 and an ET-1 binding site in rat cardiac membranes.

Endothelin is a local peptide hormone produced by vascular endothelial cells and is the most potent vasoconstrictor yet identified. There is increasing evidence that endothelin plays a role in the regulation of vascular tone and blood pressure and in the pathophysiology of cardiovascular, renal and respiratory disease.

1. The hypothesis that endothelin (ET) influences sympathetically mediated vasoconstriction was investigated in 13 healthy, male subjects. 2. ET (1 pmol min-1) was infused for 60 min into the left brachial arteries of seven healthy male subjects. Resting forearm blood flow, and sympathetic vasoconstriction produced by lower body negative pressure (LBNP; 15 mm Hg), was measured in both arms by strain gauge plethysmography. In a further six subjects, noradrenaline (NA) was infused intra-arterially at doses of 150-600 pmol min-1, with and without co-infusion of ET (1 pmol min-1), with blood flow measured in both forearms. 3. ET produced a small but significant reduction of blood flow in the infused forearm from 3.9 +/- 0.6 ml 100 ml-1 min-1 during infusion of saline, to 3.3 +/- 0.7 ml 100 ml-1 min-1 during infusion of ET at 60 min (P less than 0.05). Blood flow in the non-infused forearm was not altered by ET infusion. 4. NA produced a significant and dose-dependent reduction of blood flow in the infused forearm from 3.13 +/- 0.5 ml 100 ml-1 min-1 during saline infusion, to 1.49 +/- 0.2 ml 100 ml-1 min-1 with NA at 600 pmol min-1 (P less than 0.001). During co-infusion of ET, blood flow was reduced similarly in the infused arm from 3.15 +/- 0.7 ml 100 ml-1 min-1 during saline infusion to 1.55 +/- 0.2 ml 100 ml-1 min-1 with NA at 600 pmol min-1. Blood flow in the non-infused arm was not altered by ET and NA infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Endothelin-1 (ET-1) is a potent vasoconstrictor and pressor agent with a prolonged action in animal preparations. We have investigated the vascular pharmacology of ET-1 in healthy human volunteers. In arterial studies, blood flow was measured in both forearms and the left brachial artery cannulated for infusion of drugs and vehicle. Local blood flow was reduced in a dose-dependent manner by ET-1. At 5 pmol/min, the response was maximal by 60 min, and flow returned to baseline only after a further 120 min. ET-1 and angiotensin II, both at 5 pmol/min, reduced flow by 40% at 60 min. These responses were reversed to a similar degree by the dihydropyridine calcium antagonist nicardipine (0.3-10 micrograms/min). At 1 pmol/min, ET-1 did not affect sympathetic vasoconstriction to arterial norepinephrine or lower body negative pressure. In venous studies, the internal diameter of a dorsal hand vein was measured during local infusion of drugs and vehicle. ET-1 (5 pmol/min) produced local venoconstriction, with a maximal reduction in vein size of 83 +/- 12% at 60 min. Reversal of responses was slow, and unaffected by nicardipine (1.5 micrograms/min). We have shown that ET-1 is a potent vasconstrictor producing prolonged effects in resistance and capacitance vessels in humans. Low-dose ET-1 does not appear to affect sympathetically mediated vasoconstriction. Studies with nicardipine do not provide support for the hypothesis that ET-1 is an endogenous ligand of the dihydropyridine-sensitive calcium channel.

Inactivation of endothelin B receptors (ETB), either through selective pharmacological antagonism or genetic mutation, increases the circulating concentration of endothelin-1 (ET-1), suggesting ETB plays an important role in clearance of this peptide. However, the cellular site of ETB-mediated clearance has not yet been determined. We have used a novel mouse model of endothelial cell-specific knockout (KO) of ETB (EC ETB-/-) to evaluate the relative contribution of EC-ETB to the clearance of ET-1. Phenotypic evidence of EC-specific ETB KO was confirmed by immunocytochemistry and autoradiography. Binding of the radiolabelled selective ETB ligand BQ3020 was significantly and selectively decreased in EC-rich tissues of EC ETB-/- mice, including the lung, liver, and kidney. By contrast, ETA binding was unaltered. RT-PCR confirmed equal expression of ET-1 in tissue from EC ETB-/- mice and controls, despite increased concentration of plasma ET-1 in EC ETB-/-. Clearance of an intravenous bolus of [125I]ET-1 was impaired in EC ETB-/- mice. Pretreatment with the selective ETB antagonist A192621 impaired [125I]ET-1 clearance in control animals to a similar extent, but did not further impair clearance in EC ETB-/- mice. These studies suggest that EC-ETB are largely responsible for the clearance of ET-1 from the circulation. L’inactivation des récepteurs de type B de l’endothéline (ETB), par un antagonisme pharmacologique sélectif ou une mutation génétique, augmente la concentration d’endothéline-1 circulante, ce qui laisse supposer que l’ETB joue un rôle important dans la clairance de ce peptide. Toutefois, le site cellulaire de la clairance véhiculée par l’ETB n’est pas connu. Nous avons utilisé un nouveau modèle d’inactivation (KO) de l’ETB spécifique des cellules endothéliales (ETB−/− CE) chez la souris dans le but d’évaluer la contribution relative de l’ETB-CE à la clairance de l’ET-1. Nous avons confirmé, par immunocytochimie et autoradiographie, le caractère phénotypique de l’inactivation (KO) de l’ETB-CE. La fixation d’un ligand radiomarqué sélectif de l’ETB, BQ3020, a diminué significativement et sélectivement dans les tissus riches en CE des souris ETB−/− CE, y compris le poumon, le foie et le rein. En revanche, la fixation sur l’ETA est demeurée inchangée. La RT-PCR a confirmé une expression identique d’ET-1 dans les tissus des souris CE ETB−/− et témoins, malgré une augmentation de la concentration d’ET-1 plasmatique chez les souris ETB−/− CE. La clairance d’un bolus intraveineux d’[125I]ET-1 a été altérée chez les souris CE ETB−/−. Un prétraitement avec l’antagoniste sélectif de l’ETB, A192621, a perturbé la clairance de l’[125I]ET-1 à un degré similaire chez les animaux témoins, mais n’a pas altéré davantage celle des souris ETB−/− CE. Ces études donnent à penser que l’ETB-CE joue un rôle de premier plan dans la clairance de l’ET-1 de la circulation. [ABSTRACT FROM AUTHOR]