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Cardiovascular Research 1998 39(3):657-664; doi:10.1016/S0008-6363(98)00151-5
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Copyright © 1998, European Society of Cardiology

Chronic blockade of endothelin ETA receptors improves flow dependent dilation in resistance arteries of hypertensive rats

Marc Iglarz, Khalid Matrougui, Bernard I. Lévy and Daniel Henrion*

Institut National de la Santé et de la Recherche Médicale (INSERM) U 141, IFR 6 (Circulation-Lariboisière), Université Paris VII, Hôpital Lariboisière, 41, Bd de la Chapelle, 75475 Paris, Cedex 10, France

* Corresponding author. Tel.: +33-1-4463-1864; Fax: +33-1-4281-3128; E-mail: daniel.henrion@inserm.lrb.ap-hop-paris.fr

Received 24 February 1998; accepted 11 May 1998


   Abstract
Top
 Abstract
1 Introduction
 2 Methods
 3 Results
 4 Discussion
 References
 
Objective: Flow (shear stress)-induced dilation (FD) is attenuated in hypertension. Flow triggers the release by endothelial cells of dilators, such as NO or cyclo-oxygenase (COX) derivatives and constrictor factors such as endothelin-1 (ET-1) which might be involved in several cardiovascular diseases. We hypothesized that ET-1 might play a functional role in FD and participate in the endothelial dysfunction in hypertension. Methods: We investigated the effect of a chronic treatment with the ETA receptor blocker LU135252 (50 mg/kg/day) for 2 weeks on the dilator response to flow in normotensive (Wistar–Kyoto; WKY) or hypertensive (SHR, n=7 or 8 per group) rats. Results: Systolic arterial pressure was not significantly affected by chronic ETA receptor blockade in both strains. In mesenteric resistance arteries (diameter: approximately 100 µm), isolated in vitro, FD was lower and myogenic tone higher in SHR than in WKY rats. Chronic ETA receptor blockade increased FD by 73% (7.5±1.5 to 13.0±2.7 µm dilation with a flow-rate of 150 µl/min) in SHR (no effect in WKY). The participation of NO to FD was increased in SHR and the participation of dilator COX product(s) (blocked by indomethacin 10 µmol/l) to FD was significantly increased in SHR and in WKY. In control rats FD was improved by acute ETA receptor blockade in WKY rats (18.5±2.0 to 23.2±1.8 µm dilation to flow-rate of 150 µl/min) and significantly more in SHR (6.0±1.8 to 15.1±1.6 µm). Acetylcholine-induced dilation was also improved by chronic ETA receptor blockade (no effect of an acute blockade). Myogenic and phenylephrine-induced tone were not affected by chronic or acute ETA receptor blockade. The improvement of endothelium-dependent dilation was not related to a change in blood pressure Conclusion: Chronic ETA receptor blockade increased flow-induced dilation in SHR possibly by suppressing flow-induced ETA stimulation and by improving the release of dilator products by the endothelium.

KEYWORDS Flow; Shear stress; Dilation; Myogenic tone; Resistance arteries; Indomethacin; L-NAME; Endothelin-1; Hypertension; WKY rats; SHR rats


   1 Introduction
Top
 Abstract
 1 Introduction
2 Methods
 3 Results
 4 Discussion
 References
 
Endothelin-1 (ET-1) is a potent vasoconstrictor and mitogen produced by endothelial and vascular smooth muscle cells. This hormone exerts its biological effects via activation of specific receptors. Type A (ETA) receptors are expressed by vascular smooth muscle cells and may cause vasoconstriction [1]. Type B (ETB) receptors expressed by vascular smooth muscle cells are also responsible for vasoconstriction and those expressed by endothelial cells can cause vasorelaxation [1–3]. In resistance arteries vascular tone is largely dependent on the level of pressure-induced (myogenic) tone [4, 5]and on flow (shear stress)-induced dilation or contraction [6, 7]. In resistance vessels the direction of the response to flow, dilation or contraction, depends on the level of initial vascular tone [8]. Both shear stress [9]and pressure [10, 11]are potent stimulator of ET-1 synthesis and release, even though a chronic increase in flow or a high flow-rate also generate NO and cGMP production which may inhibit ET-1 production [9].

There are several pathological situations where ET-1 might be involved, including hypertension secondary to a DOCA-salt diet [12, 13], chronic angiotensin II infusion [14]or a chronic NO synthesis blockade [15]and spontaneous hypertension [16–18]. In spontaneously hypertensive rats (SHR) myogenic tone increases [19]and flow-induced dilation decreases [19, 20]. In SHR, the decreased flow-induced dilation might be related to an enhanced production of contractile factors upon flow stimulation, such as thromboxane A2 [19]or ET-1. We hypothesized that ET-1 might play a functional role in flow-induced dilation, especially in hypertension, and that flow-induced dilation might be improved by a chronic blockade of the ETA receptor. In addition, no study has yet investigated the effect of a chronic ETA receptor blockade on the vascular reactivity to pressure (myogenic tone) and flow (flow-induced dilation), although myogenic tone and flow-induced dilation are main determinants of vascular tone in resistance arteries. Indeed, chronic ETA receptor blockade might affect both flow-induced dilation and myogenic tone in resistance arteries as both pressure and flow have been shown to involve ET-1 production, as discussed above. In order to determine that chronic ET-1 treatment affected selectivity flow-induced dilation and/or pressure-induced (myogenic) tone, we also tested endothelium-dependent dilation to acetylcholine and contractions to phenylephrine. In addition, in resistance arteries from SHR, acetylcholine-induced dilation is attenuated and {alpha}-adrenoreceptor stimulation may lead to a higher contraction than in Wistar–Kyoto (WKY) [21].


   2 Methods
Top
 Abstract
 1 Introduction
 2 Methods
3 Results
 4 Discussion
 References
 
2.1 Animals
Ten week-old male WKY and SHR rats (Iffa-Credo, Lyon, France), weighing 250 g, were randomly allocated into two groups; a control group (fed with standard chow and tap water, n=8) and a LU135252 group which received the selective ETA receptor blocker LU135252 [22]in their food (50 mg/kg/day, n=7) for 2 weeks [14, 15]. After 2 weeks of treatment with LU135252, systolic blood pressure was monitored by the tail-cuff method (BP recorder 8006, Ugo Basile, Comerio, Italy). Rats were then anaesthetised [pentobarbital 50 mg/kg intraperitoneally (i.p.)] and a median laparotomy was performed. Several segments of mesenteric artery were then dissected out and immersed in ice cold physiological salt solution (PSS). The procedure followed in the care and euthanasia of the study animals was in accordance with the European Community standards on the care and use of laboratory animals (Ministère de l'Agriculture, France, authorization # 00577).

The efficiency of the treatment with LU135252 was verified in a preliminary group of experiments in which rats received LU135252 (50 mg/kg/day) for 2 weeks (n=4 per group). Rats were then anaesthetized (pentobarbital 50 mg/kg i.p.) and the right carotid artery cannulated (I.D. 0.6 mm) in order to measure blood pressure (pressure transducer: Gould P10EZ, Cleveland, OH, USA). The femoral artery was cannulated (I.D. 0.4 mm) in order to inject PSS containing ET-1 (10 nmol/l in PSS, 100 µl/min).

2.2 Isolated mesenteric resistance arteries
Segments of second order mesenteric artery (approximately 100 µm external diameter, n=7 in WKY and SHR treated with LU135252; n=8 in control WKY and SHR) were mounted in a myograph as previously described [23–26], according to the technique of Mulvany and Halpern [26]. Isometric tension was recorded and collected by a Biopac data acquisition system (Biopac MP 100, La Jolla, CA, USA) and continuously recorded (Apple computer, Cupertino, CA, USA). Arterial segments were bathed in PSS of the following composition (in mM): 135.0 NaCl, 15.0 NaHCO3, 4.6 KCl, 1.5 CaCl2, 1.2 MgSO4, 11.0 glucose, 5.0, N-2-hydroxy-ethylpiperazine-N-2-ethylsulfonic acid. The PSS was maintained at 37°C. The pCO2 was maintained at a value of 35 mmHg and the pO2 at a value of 160 mmHg [27]. Each vessel was placed under an optimal tension according to the technique of Mulvany and Halpern [26].

Cumulative concentration–response curves were obtained to phenylephrine (10 nmol/l–0.1 mmol/l). Data was expressed as milli-Newtons (mN) force per mm vessel [26]. Acetylcholine concentration–response curves (1 nmol/l–0.1 µmol/l) were obtained after preconstriction of the artery with phenylephrine (10 to 100 nmol/l, approximately 70% of the maximal contractile response). Data was expressed as % dilation of phenylephrine-induced preconstriction (n=7 in WKY and SHR treated with LU135252; n=8 in control WKY and SHR).

Concentration–response curves to phenylephrine and acetylcholine were repeated after incubation of the arteries for 20 min with the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME, 100 µM) and then with the nonselective cyclo-oxygenase inhibitor indomethacin (10 µM) in addition to L-NAME. In a separate series of experiments no drug was added (time control group, n=5 in WKY and n=6 in SHR).

2.3 Perfused and pressurized mesenteric arteries
Other segments of mesenteric resistance artery (approximately 100 µm external diameter), from the same rats than in the protocol described above, were isolated, cannulated at both ends and mounted in a video monitored perfusion system, as previously described [27, 28]using an arteriograph for resistance arteries [29]. The artery was bathed in a 5 ml organ bath containing PSS. The artery was superfused at a rate of 4 ml/min. Perfusion of the artery was set at rates ranging from 0 to 150 µl/min (under a pressure of 75 mmHg). The pressure in the proximal and in the distal ends of the artery segment was monitored and the output pump was controlled by a pressure servo control system based on the average pressure between the pressures measured in the proximal and distal ends of the artery [27, 28]. Arterial diameter was recorded using a video monitoring system (Living System Instrumentation, Burlington, VT, USA). Pressure and flow-rate could be changed independently [27, 28]. Equilibrium diameter changes were also measured in each segment, under no flow, when intraluminal pressure was set at 25, 50, 75, 100, 125 and 150 mmHg. Stepwise increases in pressure and flow were subsequently repeated after addition of either L-NAME (100 µM) or indomethacin (10 µM) to the perfusate and superfusate. At the end of each experiment arteries were perfused and superfused with a Ca2+-free PSS containing ethylene glycol-bis(-aminoethyl ether)-N,N,N',N'-tetraacetic acid (2 mM) and sodium nitroprusside (10 µM) and the pressure steps (25 to 150 mmHg) were repeated in order to determine the passive diameter of the vessel, i.e., in the absence of smooth muscle tone [19, 27, 28]. Diameter values measured in normal PSS were considered as diameter under active tone or "active diameter" [19, 27, 28]. Pressure and diameter measurements were collected and recorded as described above. Myogenic tone was expressed as percentage change from passive diameter using the equation:


Formula

Flow-induced relaxation was expressed as increases in diameter induced by flow (µm) (n=7 in WKY and SHR treated with LU135252; n=8 in control WKY and SHR).

The integrity of the vessel wall was tested in each vessel before the experimental protocol. The integrity of the smooth muscle was assessed by testing the constrictor effect of KCl (80 mM) and phenylephrine (0.1 µM). The integrity of the endothelium was assessed by testing the vasodilator effect of acetylcholine (1 µM) after preconstriction of the mesenteric arteries with phenylephrine (0.1 µM), under an intraluminal pressure of 50 mmHg. Only arteries fully dilated by acetylcholine (1 µM) were used for the experimental protocol.

2.4 Statistical analysis
Results are expressed as mean±standard error of the mean (S.E.M.), for concentration–response curves to acetylcholine and phenylephrine EC50 (IC50 for relaxation) and Emax were calculated individually for each concentration response curve using the equation [30]:


Formula

where E is the contraction, Emax is the maximum contraction, C is the concentration, EC is the EC50 (IC50 for relaxation) (concentration of agonist required to induce half the maximum response) and m is Hill's coefficient.

The effect of L-NAME and indomethacin was estimated by calculating the percentage of decrease in acetylcholine-induced dilation due to each drug.

Comparisons between groups were made using a one factor analysis of variance (ANOVA) followed by a Dunnett's t-test when significant or by two factor ANOVA for repeated measurements to compare the whole concentration–response curves in the different groups. A probability level of P<0.05 was considered significant.

2.5 Drugs
L-NAME and indomethacin were purchased from Sigma (St. Louis, MO, USA). LU135252 was kindly provided by Dr. M. Kirchengast (Knoll AG, Ludwigshafen, Germany). The other reagents were obtained from Prolabo (Paris, France).


   3 Results
Top
 Abstract
 1 Introduction
 2 Methods
 3 Results
4 Discussion
 References
 
3.1 Blood pressure
Systolic blood pressure, measured by the tail-cuff method, was significantly higher in SHR (215±7 mmHg, P<0.05, n=15) than in WKY rats (172±7 mmHg, n=15). A 2 week treatment with the chronic ETA receptor blocker LU135252 (50 mg/kg/day) had no significant effect on the systolic arterial pressure in SHR (226±8 mmHg, n=7) as well as in WKY rats (175±11 mmHg, n=7).

In a preliminary series of experiments the efficiency of the chronic ETA receptor blockade with LU135252 was tested by measuring the contractile effect of ET-1 in vivo. The increase in blood pressure due to the infusion of ET-1 (42±3 mmHg in the control group, n=4) was significantly reduced (11±2 mmHg, n=4, P<0.001) in the group receiving LU135252 for 2 weeks.

3.2 Phenylephrine-induced contraction
Phenylephrine induced a concentration-dependent contraction in mesenteric resistance arterial segments (maximal contraction: 3.8±0.3 mN/mm and EC50: 1.1±0.2 µmol/l, n=8). Phenylephrine induced contraction was equivalent in both strains and was not significantly affected by the chronic ETA receptor blockade (data not shown).

3.3 Acetylcholine-induced dilation
Acetylcholine (1 nmol/l to 10 µmol/l) induced a concentration-dependent dilation of mesenteric artery ring segments precontracted with phenylephrine, in both strains (Fig. 1). Precontraction with phenylephrine was equivalent in all the groups described. Acetylcholine-induced dilation was lower in SHR than in WKY (Fig. 1; Emax=100±2% relaxation in WKY and 86±3% relaxation in SHR, n=8, P<0.05; no significant difference between IC50 values). In SHR submitted to a chronic ETA receptors blockade acetylcholine-induced dilation was significantly higher than in SHR (Fig. 1; Emax 86±3% dilation in control SHR vs. 97±2% in SHR+LU135252, n=8 per group, P<0.05; no significant change in IC50). There was no significant difference in acetylcholine-induced dilation between control WKY and WKY+LU135252 (Fig. 1). The consecutive addition of L-NAME (100 µmol/l) and indomethacin (10 µmol/l) significantly attenuated acetylcholine-induced dilation in mesenteric arterial segments (Fig. 2), allowing us to differentiate the dilator pathway involving NO, suppressed by L-NAME and the pathway involving cyclo-oxygenase products, suppressed by indomethacin. The maximal dilation to acetylcholine was not significantly modified by L-NAME in both WKY and SHR, with or without LU135252 (Fig. 2). Nevertheless, the sensitivity to acetylcholine was significantly decreased by L-NAME as evidenced by a significant increase in IC50 (10±2 vs. 35±6 nmol/l, P<0.01) in WKY and in SHR (9±3 vs. 30±5 nmol/l). The effect of L-NAME was not significantly different in the control groups (WKY or SHR) than in the groups treated with the chronic ETA receptor blocker (Fig. 2). Indomethacin further attenuated acetylcholine-induced dilation in WKY but not in SHR (Fig. 2). Indomethacin did not affect the maximal dilation to acetylcholine but significantly increased the IC50 (35±6 to 105±16 nmol/l, P<0.01). In WKY rats submitted to a chronic ETA receptor blockade indomethacin had no significant effect on the dilation induced by acetylcholine (Fig. 2). In SHR submitted to a chronic ETA receptor blockade indomethacin did not significantly affect acetylcholine-induced dilation (Fig. 2).


Figure 1
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  		Fig. 1 Concentration-dependent dilation to acetylcholine in mesenteric resistance artery segments isolated from SHR (right panel) and WKY rats (left panel) treated for 2 weeks with the ETA receptor blocker LU135252 (50 mg/kg/day). Data is given as mean±S.E.M., n=7 or 8 per group. *P<0.05, two factor ANOVA for consecutive measurements, LU135252 vs. control.

 

Figure 2
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  		Fig. 2 Effect of the consecutive addition of L-NAME (100 µmol/l) and indomethacin (10 µmol/l) on the concentration-dependent dilation to acetylcholine in mesenteric resistance artery segments isolated from SHR (lower panel) and WKY rats (upper panel). Rats were treated for 2 weeks with the ETA receptor blocker LU135252 (50 mg/kg/day, right panel, vs. control in the left panel). Data is given as mean±S.E.M., n=7 or 8 per group. Two factor ANOVA for consecutive measurements: *P<0.05, L-NAME vs. control; §P<0.05, L-NAME+indomethacin vs. L-NAME.

 
In a separate series of experiments indomethacin (10 µmol/l) was added to the PSS without a pretreatment with L-NAME. In such conditions the attenuation of acetylcholine-induced dilation in WKY rats was similar to that given above and in Fig. 2 (IC50 from 15±4 to 42±7 nmol/l, n=5, P<0.05). In the SHR and in the rats (WKY and SHR) treated with the ETA receptor blocker indomethacin had no significant effect.

Acute ETA receptor blockade (LU135252 1 µmol/l, 60 min in organ bath) did not significantly affect acetylcholine-induced dilation or phenylephrine-induced contraction (data not shown).

3.4 Flow-induced dilation and myogenic tone
In another series of experiments, isolated mesenteric resistance arteries were submitted to different pressures and flow-rates. Stepwise increases in intraluminal pressure induced the development of myogenic tone. Myogenic tone was higher in SHR than in WKY (Fig. 3). Myogenic tone was not significantly affected by the chronic ETA receptor blockade, in either strains (Fig. 3). Passive arterial diameter, in the absence of tone ranged from 105±4 µm to 153±15 µm in WKY and from 95±7 µm to 146±8 µm in SHR, for pressure steps ranging from 25 to 150 mmHg (no significant effect of the chronic ETA receptor blockade). Acute ETA receptor blockade with LU135252 (1 µmol/l, 60 min) had no significant effect on myogenic tone in either WKY rats or SHR (not shown).


Figure 3
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  		Fig. 3 Myogenic tone determined in mesenteric resistance artery segments isolated from normontensive (WKY) or hypertensive (SHR) rats treated for 2 weeks with the ETA receptor blockade LU135252 (50 mg/kg/day). Data is expressed as % change in diameter from passive diameter, n=7 or 8 per group. *P<0.01; two factor ANOVA for consecutive measurements, SHR compared to WKY. Two factor ANOVA for consecutive measurements: no significant difference vs. LU135252.

 
Stepwise increases in flow (0 to 150 µl/min, corresponding to shear stress values from 0 to 65 dynes/cm2), at a pressure of 75 mmHg produced a significant dilation in WKY rats (Fig. 4). Arterial diameter (under 75 mmHg, before induction of flow) was 115±5 µm in WKY (n=8), 113±6 µm in WKY+LU135252 (n=7); 97±5 µm in SHR (n=8) and 98±6 µm in SHR+LU135252 (n=7). In SHR, flow-induced dilation was significantly lower as compared to WKY (Fig. 4). Chronic ETA receptor blockade increased significantly flow-induced dilation in SHR (7.5±1.5 to 13.0±2.7 µm dilation with a flow-rate of 150 µl/min) and had no significant effect in WKY rats (Fig. 4).


Figure 4
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  		Fig. 4 Flow-induced dilation in mesenteric resistance artery segments isolated from normontensive (WKY) or hypertensive (SHR) rats treated for 2 weeks with the ETA receptor blockade LU135252 (50 mg/kg/day). Data is expressed as mean±S.E.M., n=7 or 8 per group. *P<0.001; two factor ANOVA for consecutive measurements, SHR compared to WKY. #P<0.01; two factor ANOVA for consecutive measurements, LU135252 compared to control (CONT).

 
In mesenteric resistance arteries from WKY rats, flow-induced dilation was significantly increased by acute ETA receptor blockade (18.5±2.0 to 23.2±1.8 µm dilation to flow, flow=150 µl/min, P<0.01). Similarly, in arteries from SHR acute ETA receptor blockade significantly increased flow-induced dilation (6.0±1.8 to 15.1±1.6 µm, flow=150 µl/min, P<0.001). The increase in flow-induced dilation due to acute ETA receptor blockade was significantly higher in SHR than in WKY (P<0.001).

The NO synthesis blocker L-NAME (100 µmol/l, Fig. 5) significantly attenuated flow-induced dilation in WKY (90±6 to 30±5% attenuation of flow-dilation, flow from 9 to 150 µl/min, P<0.001). L-NAME (100 µmol/l) had no significant effect on flow-induced dilation in SHR (Fig. 5).


Figure 5
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  		Fig. 5 Effect of L-NAME (100 µmol/l) and indomethacin (INDO, 10 µmol/l) on flow-induced dilation in mesenteric artery segments isolated from WKY rats treated for 2 weeks with the ETA receptor blocker LU135252 (50 mg/kg/day in the lower panel, vs. control rats: upper panel). Data is given as mean±S.E.M., n=7 or 8 per group. Two factor ANOVA for consecutive measurements: *P<0.001, L-NAME compared to control (CONT); #P<0.001, L-NAME+INDO compared to control (CONT).

 
The cyclo-oxygenases inhibitor indomethacin (10 µmol/l, Figs. 4 and 5Go) significantly attenuated flow-induced dilation in WKY (85±5 to 30±5% attenuation, 9 to 150 µl/min, P<0.001) and in SHR (50±17 to 34±8%, 9 to 150 µl/min, P<0.01: SHR vs. WKY). The effect of indomethacin on flow-induced dilation was significantly less important in SHR than in WKY (e.g. 50±17% vs. 85±5% attenuation of flow-induced dilation, P<0.005).

In WKY rats the attenuation of flow-induced dilation by L-NAME was not significantly modified by the chronic ETA receptor blockade (Fig. 5). In SHR the decrease in flow-induced dilation due to L-NAME was significant in SHR submitted to chronic ETA receptor blockade whereas L-NAME had no significant effect on flow-induced dilation in control SHR (Fig. 6). The attenuation of flow-induced dilation by indomethacin was significantly higher in WKY and SHR submitted to chronic ETA receptor blockade than in their respective controls (Figs. 5 and 6Go). For example, indomethacin suppressed 31±5% of flow-induced dilation in control SHR and 52±7% of flow-induced dilation in SHR+LU135252 (P<0.005). In control WKY indomethacin suppressed 46±5% of flow-induced dilation vs. 61±6% of flow-induced dilation in WKY+LU135252 (P<0.01).


Figure 6
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  		Fig. 6 Effect of effect L-NAME (100 µmol/l) and indomethacin (INDO, 10 µmol/l) on flow-induced dilation in mesenteric artery segments isolated from SHR treated for 2 weeks with the ETA receptor blocker LU135252 (50 mg/kg/day, lower panel, vs. control: upper panel). Data is given as mean±S.E.M., n=7 or 8 per group. Two factor ANOVA for consecutive measurements: *P<0.001, L-NAME compared to control (CONT); #P<0.001, L-NAME+INDO compared to control (CONT).

 
In a separate series of experiments (n=4 per group) indomethacin (10 µmol/l) was added first to the PSS without a pretreatment with L-NAME. In such conditions the attenuation of flow-induced dilation was similar to that given above (data not shown).


   4 Discussion
Top
 Abstract
 1 Introduction
 2 Methods
 3 Results
 4 Discussion
References
 
In the present study we showed that chronic ETA receptor blockade improved mainly flow-induced dilation in mesenteric resistance arteries from hypertensive rats.

In agreement with previous works [21], vascular reactivity to acetylcholine was decreased in SHR versus WKY. Similarly, myogenic tone was found to be higher in SHR (present study and Refs. [8, 19]) whereas flow-induced dilation was lower (present study and Refs. [19, 20, 31]) and resistant to NO synthase blockade, as previously demonstrated [19, 20]. The sensitivity of flow-induced dilation to cyclo-oxygenase inhibition was also decreased in SHR (present study and Ref. [19]).

The main effect of the chronic ETA receptor blockade was an increase in flow-induced dilation and in a lesser proportion in acetylcholine-induced dilation, in mesenteric resistance arteries in SHR, not in WKY rats. A possible explanation is that ET-1, produced upon flow stimulation [9]counteracted the dilation. This would apply mainly in SHR in which acute and chronic ETA receptor blockade improved flow-induced dilation much more than in WKY rats (not significant in WKY). Indeed, ET-1 produced upon flow stimulation may act as an endothelium-derived contracting factor ("EDCF"). This effect might be pronounced in SHR and could be inhibited or reversed by the chronic treatment with an ETA receptor blocker, at least in SHR, in which flow-induced dilation was effectively increased after chronic ETA receptor blockade. This may not be the only explanation as acute ETA receptor blockade also improved flow-induced dilation, although in a lesser proportion, in WKY. Moreover, acetylcholine-induced dilation was not affected by acute ETA receptor blockade whereas it was higher after chronic ETA receptor blockade. Additional explanations could involve an effect of the long-term ETA receptor blockade on endothelium-dependent dilators, as discussed below.

After chronic ETA receptor blockade the participation of cyclo-oxygenase products in flow-induced dilation was increased, in both strains. Similarly, the participation of NO in the flow-induced response was also increased after chronic ETA receptor blockade (significant only in SHR). One possible explanation could be that ET-1 would contract vessels, at least in part, through vasoconstrictor prostanoid(s). Chronic ETA receptor blockade could prevent this vasoconstrictor prostanoid(s)-dependent activity and thus favor the effect of vasodilator prostanoid(s) upon flow stimulation. This possibility is supported by the observation that ET-1 may act through the release of vasodilator prostanoids [32–37]and that flow stimulates the release of both vasodilator and vasoconstrictor prostanoids [19]. Interestingly, mesenteric arteries from SHR produce more vasoconstrictor prostanoids and less vasodilator prostanoids than SHR [19], thus the mechanism described above would be more pronounced in SHR. This could explain, at least in part, that flow-induced dilation was improved only in SHR. Another possible explanation might be that the absence of flow-induced activation of ETA receptors or the overactivation by ET-1 of ETB receptors would activate NO and cyclo-oxygenase derivatives production. This cannot be directly concluded from our study as no ETB receptor blocker was used, but previous studies have shown that the effect of ETB receptor stimulation could be evidenced mainly after ETA blockade [32]. Moreover, ET-1-induced dilation has been shown to involve NO [33–36]and/or cyclo-oxygenase derivatives production [33, 37]. Thus, together with the loss of activation of the ETA receptor, an increased effect of ETB receptor activation might occur during the chronic blockade of the ETA receptors.

Flow and acetylcholine both induced a endothelium-dependent dilation in rat mesenteric resistance arteries. NO-synthase and cyclo-oxygenases blockade could only block part of this dilation, suggesting, as previously shown in resistance arteries [38–40]including the mesenteric arteries [41], that other dilating factors are involved. Among these factors, endothelium-derived hyperpolarizing factors ("EDHF") are the most important, although other not yet identified factors are also involved [39].

Finally, and interestingly, no change in blood pressure was observed after chronic ETA receptor blockade in WKY as well as in SHR, despite a significant increase in flow-induced dilation in SHR. On the other hand, we have recently reported results showing a significant decrease in flow-induced dilation in a mice model with a normal blood pressure [28]. Thus flow-induced dilation may not necessarily be a major determinant of blood pressure. Nevertheless such an issue requires further investigation.

In conclusion, chronic ETA receptor blockade increased mainly flow-induced dilation in hypertensive rats by inhibiting the "EDCF" role of ET-1 and/or by activating NO and cyclo-oxygenase derivatives production.

Time for primary review 24 days.


   Acknowledgements
 
Marc Iglarz was a fellow of the "Fondation pour la Recherche Médicale" (Paris, France).


   References
Top
 Abstract
 1 Introduction
 2 Methods
 3 Results
 4 Discussion
 References
 

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