Endothelial NO-dependent Relaxation Research Articles

The Beneficial Effect of Swimming Training Associated with Quercetin Administration on the Endothelial Nitric Oxide-Dependent Relaxation in the Aorta of Rats with Experimentally Induced Type 1 Diabetes Mellitus.

Type 1 diabetes mellitus is related to the vascular oxidative and nitrosative stress, the trigger for atherosclerosis and cardiovascular complications. The effects of moderate swimming training associated with quercetin oral administration were evaluated in aorta of rats with experimentally induced type 1 diabetes mellitus (T1DM), by analysing the nitric oxide-endothelial dependent relaxation (NO-EDR). T1DM rats received daily quercetin 30 mg/kg and followed the protocol of 5-weeks swimming exercise (30 min/day; 5 days/week). Aorta relaxation to acetylcholine (Ach) and sodium nitroprusside (SNP) were measured at the end of the experiment. Ach-induced endothelial dependent relaxation was significantly decreased in phenylephrine (PE) pre-contracted aorta of diabetic rats. Swimming exercise with quercetin administration preserved Ach-induced EDR but did not have any impact on SNP-induced endothelium-independent relaxation in the diabetic aorta. These findings suggest that quercetin administration associated with moderate swimming exercise could improve the endothelial NO-dependent relaxation in the aorta of rats with experimentally induced type 1 diabetes mellitus, showing that this therapeutical combination may improve and even prevent the vascular complications that occur in diabetic patients.

Exercise training improves cerebral endothelial function in juvenile and mature Ossabaw miniature‐pigs

BackgroundChronic hyperglycemia promotes excessive production of endothelial reactive oxygen species (ROS), which may impair nitric oxide (NO) signaling, a hallmark of vascular disfunction. How maturation and exercise training influence this process in the cerebrovasculature has yet to be explored. The purpose of this study was to examine the role of ROS on endothelial‐dependent cerebral dilation at different stages of maturation in sedentary and exercise trained hyperglycemic pigs. It was hypothesized that mature pigs would display reduced endothelial NO‐dependent dilation coinciding with augmented NAD(P)H Oxidase (NOX; involved in ROS production) activity and reduced superoxide dismutase (SOD; involved in ROS clearance) activity compared with juvenile pigs. Furthermore, compared with sedentary (SED) pigs, pigs that completed high‐intensity interval training (HIIT) would exhibit enhanced endothelial NO‐dependent dilation coupled with decreased NOX and increased SOD signaling.MethodsTwenty juvenile (n=10F/10M; 3±1 mo.) and 17 mature (n=9F/7M; 14±1 mo.) Ossabaw miniature‐pigs were divided into SED or HIIT groups. The HIIT consisted of 3 sessions per week (5X3 min intervals interspersed with 1 min recovery periods performed on a motorized treadmill) for 8 weeks. Animals were group housed with access to 1 kg of feed/pig/day and ad libitum access to a 10% sugar water solution. At the end of the intervention pigs were euthanized, and cerebral arteries were harvested for arterial myography experiments. To examine endothelial NO‐dependent relaxation, vasomotor responses to bradykinin (BK; 3e‐15M to 1e‐6M) were studied in the presence or absence of NO synthase inhibition (L‐NAME; 3e‐4 M). Further, to examine the role of ROS signaling, vasodilatory responses to BK were studied under the following conditions: 1) NOX inhibition (Apocynin; 1e‐4 M) and 2) SOD mimetic (TEMPOL; 1e‐4 M).ResultsAt the end of the trial all groups were considered hyperglycemic (mean plasma glucose> 8.5 mmol/L), and plasma glucose values were lower in mature (8.8±1.6 mmol//L) vs. juvenile pigs (10.3±2.2 mmol/L; main effect; P<0.05). Regarding cerebral vasomotor control, BK‐induced dilation was greater in HIIT vs. SED groups (main effect; P<0.01), owing to enhanced NO signaling (main effect; P=0.05). Treatment with a NOX inhibitor increased BK‐induced dilation in juvenile (P=0.05) and mature SED pigs (P=0.01). Treatment with a SOD mimetic did not influence dilation in juvenile SED pigs (P=0.87), and increases in BK‐induced dilation in mature SED pigs approached significance (P=0.07).ConclusionThese results show that despite being hyperglycemic, HIIT improved endothelial‐dependent cerebral dilation in juvenile and mature pigs likely owing to enhanced NO bioavailability, secondary to decreased NOX and increased SOD signaling. Maturity was associated with lower levels of hyperglycemia and did not impair cerebral endothelial‐dependent dilation.

Deletion of Protein Tyrosine Phosphatase 1B (PTP1B) Enhances Endothelial Cyclooxygenase 2 Expression and Protects Mice from Type 1 Diabetes-Induced Endothelial Dysfunction.

Protein tyrosine phosphatase 1B (PTP1B) dephosphorylates receptors tyrosine kinase and acts as a molecular brake on insulin signaling pathway. Conditions of metabolic dysfunction increase PTP1B, when deletion of PTP1B protects against metabolic disorders by increasing insulin signaling. Although vascular insulin signaling contributes to the control of glucose disposal, little is known regarding the direct role of PTP1B in the control of endothelial function. We hypothesized that metabolic dysfunctions increase PTP1B expression in endothelial cells and that PTP1B deletion prevents endothelial dysfunction in situation of diminished insulin secretion. Type I diabetes (T1DM) was induced in wild-type (WT) and PTP1B-deficient mice (KO) with streptozotocin (STZ) injection. After 28 days of T1DM, KO mice exhibited a similar reduction in body weight and plasma insulin levels and a comparable increase in glycemia (WT: 384±20 vs. Ko: 432±29 mg/dL), cholesterol and triglycerides, as WT mice. T1DM increased PTP1B expression and impaired endothelial NO-dependent relaxation, in mouse aorta. PTP1B deletion did not affect baseline endothelial function, but preserved endothelium-dependent relaxation, in T1DM mice. NO synthase inhibition with L-NAME abolished endothelial relaxation in control and T1DM WT mice, whereas L-NAME and the cyclooxygenases inhibitor indomethacin were required to abolish endothelium relaxation in T1DM KO mice. PTP1B deletion increased COX-2 expression and PGI2 levels, in mouse aorta and plasma respectively, in T1DM mice. In parallel, simulation of diabetic conditions increased PTP1B expression and knockdown of PTP1B increased COX-2 but not COX-1 expression, in primary human aortic endothelial cells. Taken together these data indicate that deletion of PTP1B protected endothelial function by compensating the reduction in NO bioavailability by increasing COX-2-mediated release of the vasodilator prostanoid PGI2, in T1DM mice.

PTP1B Deletion Enhances Endothelial Cyclooxygenase 2 Expression and Protects Mice from Type 1 Diabetes‐Induced Endothelial Dysfunction

Protein tyrosine phosphatase 1B (PTP1B) dephosphorylates receptors tyrosine kinase and acts as a molecular brake on insulin signaling. Although vascular insulin signaling contributes to the control of glucose disposal, little is known regarding the role of PTP1B in the control of endothelial function. We hypothesized that metabolic dysfunctions increase PTP1B expression in endothelial cells and that PTP1B deletion prevents endothelial dysfunction in situation of diminished insulin secretion. Type I diabetes (T1DM) was induced in wild-type (WT) and PTP1B-deficient mice (KO) with streptozotocin (STZ). After 28 days of T1DM, body weight reduction, decrease in plasma insulin levels and increased in glycemia, cholesterol and triglycerides, were similar in WT and KO mice. T1DM increased PTP1B expression and impaired aortic endothelial NO-dependent relaxation. PTP1B deletion did not affect baseline endothelial NO-dependent relaxation, but preserved endothelium-dependent relaxation, in T1DM mice. NO synthase inhibition with L-NAME abolished endothelial relaxation in control and T1DM WT mice. L-NAME and the cyclooxygenases inhibitor indomethacin were required to abolish endothelium relaxation in T1DM KO mice. PTP1B deletion increased COX-2 expression and PGI2 levels, in mouse aorta and plasma respectively, in T1DM mice. In parallel, simulation of diabetic conditions increased PTP1B expression and knockdown of PTP1B increased COX-2 but not COX-1 expression, in primary human aortic endothelial cells. These data indicate that deletion of PTP1B protected endothelial function by compensating the reduction in NO bioavailability by increasing COX-2-mediated release of the vasodilator prostanoid PGI2, in T1DM mice.

Characterization of the components of urban particulate matter mediating impairment of nitric oxide-dependent relaxation in intrapulmonary arteries

We have previously shown that exposure to urban particulate matter (UPM) impairs endothelial nitric oxide (NO) bioactivity in intrapulmonary arteries. As UPM is composed of heterogeneous constituents, the aim of this study was to clarify the class of pollutants responsible for such effect. Extracts (aqueous, acidic or organic) were prepared from SRM1648, an UPM sample collected in St. Louis (MO, USA). The metal composition of extracts as well as endotoxin content was determined. The effects of each extract, metal mixture and endotoxin were evaluated on endothelium-dependent relaxation to acetylcholine (reflecting endothelial NO production) in rat isolated intrapulmonary arteries. Aqueous or organic SRM1648 pretreatment altered acetylcholine-induced relaxation, similar to that induced by native SRM1648. Organic extract induced similar attenuation of acetylcholine relaxation than organic-treated SRM1648, whereas aqueous extract had no effect. Acidic pretreatment, which impoverished metal and endotoxin content of SRM1648, prevented the impairment of acetylcholine-induced relaxation. However, neither the acidic extract enriched in metals, nor a metal mixture representative of SRM1648 content, modified acetylcholine relaxation, while endotoxin impaired it. Polymyxin B, which chelates endotoxin, prevented SRM1648-induced decrease in relaxation to acetylcholine. It is concluded that SRM1648-induced impairment of endothelial NO-dependent relaxation in intrapulmonary arteries unlikely involved a soluble factor released by vascular cells during UPM exposure, but rather an organic extractible and acidic-sensitive constituents of UPM. Endotoxin, but not metals, may be responsible for UPM-induced impairment of endothelial NO-dependent relaxation.

Intense exercise training is not effective to restore the endothelial NO-dependent relaxation in STZ-diabetic rat aorta

BackgroundThe aim of this study was to examine the effects of intense physical training on vascular function in streptozotocin-diabetic rats. We focused on the endothelium-dependent relaxation (EDR) induced by acetylcholine (ACh) and stable ADP adenosine-5′- O – (2-thiodiphosphate) (ADPβS).MethodsControl or diabetic male Wistar rats (n=44) were randomly assigned to sedentary or trained groups. The training program consisted in a regular period of running on a treadmill during 8 weeks (10° incline and up to 25 m/min, 60 min/day). The reactivity of isolated thoracic aorta rings of healthy, diabetic and/or trained has been tested.ResultsACh and ADPβS-induced EDR were observed in phenylephrine (PE) pre-contracted vessels. As compared to sedentary control group, diabetic rats showed an increase in PE-induced contraction and a decrease in ACh and ADPβS-induced EDR (p<0.05). Moreover, there were no increase in ACh and ADPβS-induced EDR in diabetic rats. N-Nitro-L-Arginine Methyl Ester inhibited the nitric oxide synthase in diabetic and control rats, thereby resulting in a strong inhibition of the EDR induced by ACh and ADPβS (10-6 M).ConclusionDiabetes induced an endothelium dysfunction. Nevertheless, our intense physical training was not effective to restore the aorta endothelial function.

Anemonin, from Clematis crassifolia, potent and selective inducible nitric oxide synthase inhibitor

The aim of this study was to examine the anti-inflammatory effects of aerial part of Clematis crassifolia Benth. (Ranunculaceae) based on an iNOS inhibition in lipopolysaccharide (LPS) activated macrophages. Bioassay-guided fractionation and purification led to the isolation of ibotanolide B ( 1), calceolarioside B ( 2), trans-caffeic acid ( 3), anemonin ( 4) and 3′,4′,5,7-tetrahydroxy-6-C-glucopyranosylflavone ( 5). Their structures were elucidated on the basis of spectroscopic analysis. All these compounds inhibited NO production, detected as nitrite, in activated macrophages except 5. Among them, anemonin ( 4) was the most potent. Analyses of reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting revealed that it decreased the expression of iNOS mRNA and protein in activated RAW 264.7 cells. In isolated rat thoracic aortic rings, anemonin prevented the vascular hyporeactivity to phenylephrine induced by LPS whereas it did not affect acetylcholine-induced endothelial NO-dependent relaxation, an index of endothelial NOS (eNOS) activity. These results indicated that the potential anti-inflammatory effect of anemonin, the naturally occurring selective iNOS inhibitor, may provide a rationale for the medical use of Clematis crassifolia.

-adrenergic relaxation in pulmonary arteries: preservation of the endothelial nitric oxide-dependent 2 component in pulmonary hypertension

beta-adrenoceptor (beta-AR)-mediated relaxation was characterized in pulmonary arteries from normoxic and hypoxic (as model of pulmonary hypertension) mice. The endothelial NO synthase (eNOS) pathway was especially investigated because of its potential vasculoprotective effects. Pulmonary arteries from control or hypoxic (0.5 atm for 21 days) wild-type or eNOS-/- mice were used for pharmacological characterization of beta-AR-mediated relaxation in myograph, and for immunohistochemistry using anti-beta-AR antibodies. In pulmonary arteries from normoxic mice, isoproterenol (beta-AR agonist) and procaterol (selective beta2-AR agonist) elicited relaxation, while cyanopindolol and CL316243 (beta3-AR agonists) were ineffective. The effect of isoproterenol was antagonized by CGP20712A and ICI118551 (beta1- or beta2-AR antagonists, respectively) and also partially inhibited by N omega-nitro-L-arginine methylester (L-NAME, a NOS inhibitor), endothelium denudation, or eNOS gene deletion. Relaxation to procaterol was abolished by L-NAME or endothelium removal. In eNOS-/- mice, procaterol-induced relaxation was decreased but was insensitive to L-NAME, this residual effect involving other endothelium-dependent relaxant factors as compensatory mechanisms. Immunostaining for beta2-AR was observed in the endothelial layer, but not the medial layer of pulmonary arteries. Pulmonary arteries from hypoxic mice exhibited decreased endothelial NO-dependent relaxation to acetylcholine. However, in these arteries, relaxation to procaterol was either unaffected (extralobar segments) or even increased (intralobar segments) and was still abolished by L-NAME or endothelium removal. beta1- and beta2-AR, but not beta3-AR, mediate relaxation of mice pulmonary arteries. The beta2-AR component is dependent on eNOS activity and is preserved following chronic hypoxia. These data highlight the role of the beta2-AR as a pharmacological target to induce/restore endothelial NO-dependent protective effects in pulmonary circulation.

Selective inducible nitric oxide synthase suppression by new bracteanolides from Murdannia bracteata

Murdannia bracteata has been used as a Taiwanese folk medicine for its anti-inflammatory properties. However, neither its active ingredients nor its anti-inflammatory actions are well defined. Nitric oxide (NO), overproduced by activated macrophages via inducible NO synthase (iNOS), is suggested to be a significant pathogenic factor in various inflammatory tissue injuries. In order to elucidate the anti-inflammatory actions of M. bracteata, the present study was designed to isolate its active constituents and examine its effects on iNOS in lipopolysaccharide (LPS)-activated macrophages. Two new hydroxybutenolides, bracteanolide A ( 1) and B ( 2), together with (+)-( R)- p-hydroxyphenyllactic acid ( 3) and isovitexin ( 4), were isolated and identified from M. bracteata by the NO production assay . All of the compounds inhibited NO production except 3. Their rank order of potency was 1 > 2 > 4. Among these, 1 significantly inhibited NO production, which is associated with its suppression on iNOS induction in a concentration-dependent manner, with an IC 50 of 33.27 ± 0.86 μM. Nevertheless, isometric tension recordings in isolated endothelium-intact rat aorta revealed that 1–4 did not affect acetylcholine-induced endothelial NO-dependent relaxation, an index of endothelial NOS (eNOS) activity. The selective inhibition on iNOS provides a possible explanation for the anti-inflammatory use of M. bracteata.

Distinct Activity of Peptide Mimetic Intracellular Ligands (Pepducins) for Proteinase‐Activated Receptor‐1 in Multiple Cells/Tissues

Proteinase-activated receptor-1 (PAR1), a G protein-coupled receptor (GPCR) for thrombin, can be activated not only by PAR1-activating peptides (PAR1APs) based on the N-terminal cryptic tethered ligand sequence but also by an N-palmitoylated (Pal) peptide, Pal-RCLSSSAVANRSKKSRALF-amide (P1pal-19), based on the intracellular loop 3 of PAR1, designated pepducin, in human platelets or PAR1-transfected cells. The present article evaluated the actions of P1pal-19 and also the shorter peptide, Pal-RCLSSSAVANRS-amide (P1pal-12), known as a possible PAR1 antagonist, in multiple cells/tissues that naturally express PAR1. P1pal-19 as well as a PAR1AP, TFLLR-amide, evoked cytosolic Ca(2+) mobilization in cultured human lung epithelial cells (A549) and rat gastric mucosal epithelial cells (RGM1). P1pal-19 and TFLLR-amide, but not a PAR2-activating peptide, SLIGRL-amide, caused delayed prostaglandin E(2) formation in RGM1 cells. P1pal-19, like TFLLR-amide, produced endothelial NO-dependent relaxation in rat aorta and epithelial prostanoid-dependent relaxation in mouse bronchus. The P1pal-19-induced relaxation remained constant even after desensitization of PAR1 with TFLLR-amide in either tissue. P1pal-19 failed to mimic the contractile effects of TFLLR-amide in the endothelium-denuded preparations of rat aorta or superior mesenteric artery and the rat gastric longitudinal smooth muscle strips. P1pal-12 partially inhibited the vasorelaxation caused by TFLLR-amide and P1pal-19, but not SLIGRL-amide, in the rat aorta. Our data thus indicate that P1pal-19 is capable of mimicking the effects of PAR1APs in the endothelial and epithelial, but not smooth muscle, cells/tissues, and suggest that P1pal-12 may act as a PAR1 antagonist in the vascular endothelium.

Attenuated endothelium-mediated relaxation by acteoside in rat aorta: Role of endothelial [Ca 2+] i and nitric oxide/cyclic GMP pathway

Acteoside and other phenylethanoid glycoside are contained in many plants that are widely used in traditional Chinese herbal medicine. Acteoside possesses multiple biological actions. Its effect on the vascular system is, however, incompletely understood. This study was aimed to investigate the role of endothelial [Ca 2+] i, nitric oxide (NO), and cyclic GMP in acteoside-induced inhibition of endothelial NO-mediated relaxation in rat aorta. Acteoside reduced endothelial NO-dependent relaxation induced by acetylcholine (Ach) or A23187. Acteoside inhibited Ach-stimulated increase in tissue content of cyclic GMP in endothelium-intact rings. L-NNA abolished the stimulatory effect of Ach. Treatment with acteoside significantly suppressed bradykinin-induced increase in [Ca 2+] i of cultured rat aortic endothelial cells. Acute exposure to acteoside (30 μM) did not affect the expression of eNOS mRNA in endothelium-intact rings. In summary, acteoside impairs endothelial NO-mediated aortic relaxation partially through inhibition of agonist-induced endothelial Ca 2+ mobilization and Ca 2+-dependent NO production and subsequent suppression of cyclic GMP formation. This novel pharmacological action if occurring in small vessels in vivo, may contribute to the reported anti-inflammatory effect of acteoside against NO-mediated vascular permeability-related acute edema.

Decreased expression of myocardial eNOS and caveolin in dogs with hypertrophic cardiomyopathy.

Because nitric oxide (NO) regulates cardiac and vessel contraction, we compared the expression and activity of the endothelial NO synthase (eNOS) and caveolin, which tonically inhibits eNOS in normal and hypertrophic cardiomyopathic hearts. NOS activity (L-[(3)H]citrulline formation), eNOS immunostaining, and caveolin abundance were measured in heart tissue of 23 mongrel dogs before and at 3 and 7 wk of perinephritic hypertension (PHT). Hemodynamic parameters in vivo and endothelial NO-dependent relaxation of macro- and coronary microvessels in vitro were assessed in the same animals. eNOS immunostaining and total calcium-dependent NOS activity decreased at 7 wk in all four heart cavities (in left ventricle, from 17.0 +/- 1.3 to 0.2 +/- 0.2 fmol. min(-1). mg protein(-1), P < 0.001). Caveolin-1 and -3 also decreased in PHT dog hearts. Accordingly, basal vascular tone was preserved, but maximal endothelial NO-dependent relaxation was impaired in all vessels from 7-wk PHT dogs. The latter had preserved systolic function but impaired diastolic relaxation [relaxation time constant (T(1)), 25.1 +/- 0.9 vs. 22.0 +/- 1 ms in controls; P < 0.05]. Peripheral infusion of the NOS inhibitor N(G)-nitro-L-arginine methyl ester increased mean aortic pressure in both groups and reduced diastolic (T(1), 31.9 +/- 1.4 ms) and systolic function in PHT dogs (DP40, 47.5 +/- 2.5 vs. 59.4 +/- 3.8 s(-1) in control animals). In conclusion, both eNOS and caveolin proteins are decreased in the hypertrophic hearts of PHT dogs. This is associated with altered maximal (but not basal) vascular relaxation and impaired diastolic function. Further degradation of cardiac function after NOS inhibition suggests a critical role of residual NOS activity, probably supported by the concurrent downregulation of caveolin.

Hypotensive mechanism of [Leu13]motilin in dogs in vivo and in vitro

The effects of [Leu13]motilin were examined in vivo after its intravenous administration into anesthetized dogs and in vitro with isolated preparations of canine mesenteric artery. [Leu13]Motilin (0.1-10 nmol·kg-1, i.v.) induced both strong and clustered phasic contractions in the gastric antrum and duodenum. At doses of over 1 nmol·kg-1, [Leu13]motilin also produced transient decreases in arterial blood pressure, left ventricular pressure, maximum rate of rise of left ventricular pressure, and total peripheral resistance, and an increase in aortic blood flow and heart rate. A selective motilin antagonist, GM-109 (Phe-cyclo[Lys-Tyr(3-tBu)-betaAla]betatrifluoroacetate), completely abolished the gastric antrum and duodenal motor responses induced by [Leu13]motilin. In contrast, hypotension induced by [Leu13]motilin (1 nmol·kg-1) was unchanged in the presence of GM-109. In isolated mesenteric artery preparations precontracted with U-46619 (10-7 M), [Leu13]motilin (10-8-10-5 M) induced an endothelium-dependent relaxation, and this was inhibited by a pretreatment with Nomega-nitro-L-arginine, a competitive inhibitor of NO synthase (10-4 M). A high dose (10-4 M) of GM-109 slightly decreased [Leu13]motilin-induced relaxation, and shifted the concentration-response curve of [Leu13]motilin to the right. However, the pA2 value (4.09) of GM-109 for [Leu13]motilin in the present study was conspicuously lower than that previously demonstrated in the rabbit duodenum (7.37). These results suggest that [Leu13]motilin induces hypotension via the endothelial NO-dependent relaxation mechanism and not through the receptor type that causes upper gastrointestinal contractions.Key words: motilin, gastrointestinal motility, hypotension, hemodynamics, anesthetized dog, mesenteric artery, endothelium, nitric oxide.

Hypotensive mechanism of [Leu13]motilin in dogs in vivo and in vitro.

The effects of [Leu13]motilin were examined in vivo after its intravenous administration into anesthetized dogs and in vitro with isolated preparations of canine mesenteric artery. [Leu13]Motilin (0.1-10 nmol x kg(-1), i.v.) induced both strong and clustered phasic contractions in the gastric antrum and duodenum. At doses of over 1 nmol x kg(-1), [Leu13]motilin also produced transient decreases in arterial blood pressure, left ventricular pressure, maximum rate of rise of left ventricular pressure, and total peripheral resistance, and an increase in aortic blood flow and heart rate. A selective motilin antagonist, GM-109 (Phe-cyclo[Lys-Tyr(3-tBu)-betaAla] trifluoroacetate), completely abolished the gastric antrum and duodenal motor responses induced by [Leu13]motilin. In contrast, hypotension induced by [Leu13]motilin (1 nmol x kg(-1)) was unchanged in the presence of GM-109. In isolated mesenteric artery preparations precontracted with U-46619 (10(-7) M), [Leu13]motilin (10(-8)-10(-5) M) induced an endothelium-dependent relaxation, and this was inhibited by a pretreatment with N(omega)-nitro-L-arginine, a competitive inhibitor of NO synthase (10(-4) M). A high dose (10(-4) M) of GM-109 slightly decreased [Leu13]motilin-induced relaxation, and shifted the concentration-response curve of [Leu13]motilin to the right. However, the pA2 value (4.09) of GM-109 for [Leu13]motilin in the present study was conspicuously lower than that previously demonstrated in the rabbit duodenum (7.37). These results suggest that [Leu13]motilin induces hypotension via the endothelial NO-dependent relaxation mechanism and not through the receptor type that causes upper gastrointestinal contractions.