AJP - Heart Calcium Transients and Cell-Sarcomere
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Heart Circ Physiol 292: H1722-H1727, 2007. First published December 1, 2006; doi:10.1152/ajpheart.00612.2006
0363-6135/07 $8.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
292/4/H1722    most recent
00612.2006v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (2)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Siddiqui, A. H.
Right arrow Articles by Hussain, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Siddiqui, A. H.
Right arrow Articles by Hussain, T.

Enhanced AT1 receptor-mediated vasocontractile response to ANG II in endothelium-denuded aorta of obese Zucker rats

Athar H. Siddiqui and Tahir Hussain

Department of Pharmacological and Pharmaceutical Sciences, University of Houston, Houston, Texas

Submitted 9 June 2006 ; accepted in final form 22 November 2006


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
In the present study, we tested the hypothesis that ANG II causes a greater vasoconstriction in obese Zucker rats, a model of type 2 diabetes, with mild hypertension. Measurement of isometric tension in isolated aortic rings with intact endothelium revealed a modest but not significantly greater ANG II-induced contraction in obese than lean rats. Removal of endothelium or inhibition of nitric oxide (NO) synthase by NG-nitro-L-arginine methyl ester (L-NAME) enhanced 1) ANG II-induced contraction in both lean and obese rats, being significantly greater in obese rats (Emax g/g tissue, denuded: lean 572 ± 40 vs. obese 664 ± 16; L-NAME: lean 535 ± 14 vs. obese 818 ± 23) and 2) ANG II sensitivity in obese compared with lean rats, as revealed by the pD2 values. Endothelin-1 and KCl elicited similar contractions in the aortic rings of lean and obese rats. ACh, a NO-dependent relaxing hormone, produced greater relaxation in the aortic rings of obese than lean rats, whereas sodium nitroprusside, an NO donor, elicited similar relaxations in both rat strains. The expression of the ANG type 1 (AT1) receptor protein and mRNA in the endothelium-intact aorta was significantly greater in obese than lean rats, whereas the endothelium-denuded rings expressed modest but not significantly greater levels of AT1 receptors in obese than lean rats. The endothelial NO synthase protein and mRNA expression levels were higher in the aorta of obese than lean animals. We conclude that, although ANG II produces greater vasoconstriction in obese rat aortic rings, enhanced endothelial AT1 receptor-mediated NO production appears to counteract the increased ANG II-induced vasoconstriction, suggesting that arterial AT1 receptor may not be a contributing factor to hypertension in this model of obesity.

angiotensin II; endothelium; thoracic aorta; NG-nitro-L-arginine methyl ester; angiotensin II type 1 receptor messenger ribonucleic acid

ANG II, THE primary active product of the renin-angiotensin system (RAS), plays an important role in the regulation of cardiovascular hemodynamics via two distinct subtypes of receptors, ANG type 1 (AT1) and type 2 (AT2) receptors. The AT1 and the AT2 receptors belong to the superfamily of G protein-coupled receptors (12). ANG II, via the activation of AT1 receptors, causes vasoconstriction, aldosterone secretion, sympathetic activation, and renal sodium reabsorption and thereby regulates the blood pressure and sodium and water homeostasis (9, 22, 26). There is evidence that suggests the role of AT2 receptors to elicit vasodilatation, sodium excretion, and blood pressure reduction and thereby counteract the effects of AT1 receptors (25, 35).

Obesity is an important risk factor for hypertension (24), and higher activity of RAS has been shown to be associated with obesity-related hypertension in humans and animal models (1, 10, 17, 18). The obese Zucker rat is a model of insulin resistance with mild hypertension. Treatment with AT1 receptor antagonist losartan causes greater reduction in the blood pressure in obese than control lean animals (1). Because the plasma renin activity and the kidney contents of renin are similar or lower in obese than in lean rats (1, 4, 19), the greater blood pressure reduction by AT1 receptor antagonist suggests an enhanced AT1 receptor sensitivity to ANG II in obese rats. There have been conflicting reports as to the vascular sensitivity of AT1 receptors to ANG II in the obese Zucker rats. Although Harker et al. (19) have shown similar responses to ANG II in the aortic rings of lean and obese Zucker rats, recent studies conducted by Nishimatsu et al. (27) have shown an increased vascular response to ANG II in the aortic rings of obese Zucker rats. An in vivo study (37) reported that ANG II infusion causes greater pressor response in obese than in lean Zucker rats. Collectively, these studies led us to hypothesize that there is an increased AT1 receptor expression/function in the blood vessels of obese Zucker rats that may contribute to hypertension in these animals.

Endothelium plays an important role in the regulation of blood flow and in maintaining the vascular tone via the release of factors such as endothelin-1 and nitric oxide (NO). Endothelium is also known to modulate the ANG II-induced vasoconstriction responses (5, 6, 14). Because endothelial cell dysfunction has been reported under pathophysiological conditions such as diabetes and hypertension (29, 31), it is not known how endothelium affects ANG II-induced contractile response in obese Zucker rats. Therefore, in the present study, we studied the role of endothelium and NO on ANG II-induced vasocontractile responses in the isolated aortic rings of lean and obese Zucker rats. We also determined the protein and mRNA expression levels of the AT1 receptor and endothelial nitric oxide synthase (eNOS) in the aorta of lean and obese Zucker rats.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Experimental animals. Male lean and obese Zucker rats (10–12 wk old) were obtained from Charles River Laboratories (Wilmington, MA). They were housed in the University of Houston animal care facility, fed with the standard rat chow diet, and had free access to water. The Institutional Animal Care and Use Committee approved the experimental protocol.

Aortic ring preparation. The thoracic aorta, excised from the rats under pentobarbital sodium anesthesia (50 mg/kg ip), was placed into ice-cold modified Krebs-Henseleit buffer, pH 7.4 (21) with continuous oxygenation (95% O2-5% CO2). The aortas were cleaned of all adherent connective tissue and cut into rings (3–4 mm long).

Organ bath experiments. The aortic rings were vertically mounted in 15-ml organ baths filled with Krebs-Henseleit buffer (pH 7.4, 37°C), continuously bubbling with 95% O2-5% CO2. The changes in isometric tension of the rings were measured with a digital force isometric transducer (Harvard Apparatus, South Natick, MA) connected to data acquisition system (AD Instruments, Colorado Springs, CO). The rings were equilibrated for 1 h under an initial tension of 1 g, with the buffer being changed every 15 min. After equilibration, the rings were challenged with 50 mM KCl until reproducible contractions were achieved. Various agonists with indicated concentrations were added to the baths in a concentration cumulative manner to obtain concentration-response curves. Because ANG II may cause tachyphylaxis, we therefore obtained one ANG II concentration-response curve per aortic rings. Additionally, the AT1 receptor antagonist losartan (10–6 M) was added to the baths 20 min before the agonist, and the ANG II-induced concentration-response curves were obtained in parallel. The contractions were induced at a basal tension of 1 g and calculated as grams tension per gram tissue. The vasorelaxation responses with sodium nitroprusside (SNP, 10–9 to 10–7 M) and ACh (10–9 to 10–4 M) were elicited in vascular rings precontracted with phenylephrine (PE, 20 nM). PE at 20 nM produced submaximal but similar contractions (g tension/g tissue) in the aortic rings of lean (996 ± 92) and obese (1,197 ± 97) Zucker rats. The vascular rings that produced <50% relaxation at 100 µM in response to ACh were excluded from the studies.

To understand the role of endothelium and NO, the response to ANG II was obtained either in the endothelium-denuded aortic rings or in the presence of an inhibitor of NO synthase (NOS), NG-nitro-L-arginine methyl ester (L-NAME, 10–4 M), which was added to the baths 30 min before ANG II. The endothelium was removed by gentle rubbing of the intimal surface with the tip of a pair of small forceps. The absence and presence of endothelium was assessed by the inability and ability, respectively, of ACh (10–9 to 10–4 M) to relax the PE precontracted rings.

Western blotting and RT-PCR. The aorta was minced using a scissor and homogenized in lysis buffer (50 mmol/l beta-glycerophosphate, 100 mmol/l NaVO3, 2 mmol/l MgCl2, 1 mmol/l EGTA, 0.5% Triton X-100, 1 mmol/l dl-dithiothreitol, and 1 mmol/l phenylmethylsulfonyl fluoride) containing a cocktail of protease inhibitors that had a broad inhibitory specificity, including serine, cysteine, metalloproteases, and calpains activity. Sample protein was measured using a kit (Pierce, Rockford, IL), and was dissolved in Laemmli buffer. Equal amounts of aortic proteins (20 µg for AT1 receptor and 40 µg for eNOS), of lean and obese rats were resolved on 10 and 7.5% SDS-PAGE for AT1 receptor and eNOS, respectively, and transblotted on Immobilon P membrane (blot). The blots were incubated with primary antibodies (AT1 receptors and eNOS), followed by horseradish peroxidase-conjugated anti-rabbit or anti-mouse IgG, as secondary antibody. The signal was detected by an enhanced chemiluminescence system and recorded on X-ray films. After the bands were recorded, the blots were stripped and reprobed for glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as loading control. Densitometric analysis of the bands was performed and compared as a ratio of AT1 to GAPDH and eNOS to GAPDH between lean and obese rats.

For RT-PCR experiment, RNA was isolated from the aorta by the RNeasy mini kit protocol (QIAGEN, Valencia, CA). RNA (0.5 µg) was used in the Advantage RT-for-PCR kit (Clonetech, Mountain View, CA) for the cDNA synthesis. The resulting cDNA was used to amplify AT1 receptor, eNOS, and GAPDH cDNA. GAPDH was used as an internal control in each reaction. The primers used for AT1 receptor, eNOS and GAPDH, are shown in Table 1. The PCR products obtained were run on 1.5% agarose gels and subsequently stained in ethidium bromide. The images obtained were analyzed by FluorChem 8800 (Alpha Innotech Imaging System, San Leandro, CA) for band densitometry.


View this table:
[in this window]
[in a new window]

  		Table 1. Primer sequences used in RT-PCR procedure

 
Statistical analyses. Data are presented as means ± SE. Concentration-response curves were analyzed by nonlinear regression to obtain Emax and pD2 using GraphPad Prism 4 (GraphPad Software, San Diego, CA). Statistical significance was determined by Student's t-test and/or ANOVA followed by the Newman-Keuls post hoc test for multiple comparisons. A P < 0.05 was considered to be significant.

Chemicals and drugs. ANG II, endothelin-1, PE, KCl, SNP, ACh chloride, L-NAME, and all other chemicals used were purchased from Sigma (St. Louis, MO). Losartan was a generous gift from Merck Sharp & Dohme. AT1 receptor antibody (catalog no. sc-1173) was obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Polyclonal antibody for eNOS (catalog no. ab5589) was obtained from Abcam (Cambridge, MA). The antibody for GAPDH (catalog no. RDI-TRK5G4-6C5) was obtained from Research Diagnostics (Concord, MA). The anti-rabbit and anti-mouse secondary antibodies (catalog nos. 20320 and 40320) and Enhanced NuGlo Substrate Kit (for enhanced chemiluminescence, catalog no. 80215) were obtained from Alpha Diagnostics (San Antonio, TX).


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
ANG II-induced contraction in aortic rings. ANG II (10–9 to 10–7 M) induced contraction in the endothelium-intact aortic rings of lean and obese Zucker rats (Fig. 1). The maximal contraction was observed at 10 nM of ANG II in both lean and obese rats; thereafter, the contractile response decreased as ANG II concentrations increased. Although the Emax (g tension/g tissue) was modestly greater in obese (349 ± 43) than lean (258 ± 27) rats, it was not statistically significant (Fig. 1). Similarly, the pD2 values of ANG II in aortic rings were modestly lower in obese (8.5 ± 0.01) than lean (8.6 ± 0.002) Zucker rats.


Figure 1
View larger version (12K):
[in this window]
[in a new window]

  		Fig. 1. Concentration-response curve of ANG II (10–9 to 10–7 M) in the endothelium-intact and -denuded aortic rings of lean and obese Zucker rats. Data are expressed as means ± SE (n = 18 rats). *P < 0.05, significantly different compared with lean rats (1-way ANOVA followed by Newman-Keuls post hoc test for multiple comparisons).

 
Effect of endothelium on ANG II-induced contraction in aortic rings. The removal of endothelium resulted in a significant increase in the contractile response to ANG II in both the lean and obese Zucker rats compared with the endothelium-intact rings (Fig. 1). However, unlike in the endothelium-intact aortic rings, ANG II-induced contraction (Emax, g tension/g tissue) in endothelium-denuded aortic rings was greater in obese (664 ± 16) compared with lean (572 ± 40) rats. Although endothelium removal increased the pD2 value of ANG II in obese rat aortic rings, there was a significant decrease in the pD2 in lean rats. This led to a significant (P < 0.05) difference in the pD2 values for ANG II between lean (8.0 ± 0.02) and obese (8.8 ± 0.09) rats, suggesting enhanced sensitivity to ANG II in obese compared with lean rats.

Effect of L-NAME on ANG II-induced contraction in aortic rings. To investigate the role of NO on the ANG II-induced contraction in the endothelium-intact aortic rings of lean and obese rats, a concentration-response curve to ANG II was elicited in the presence of L-NAME (10–4 M), an NOS inhibitor. The ANG II-induced contraction was enhanced in both lean and obese rats (Fig. 2) compared with the control response of ANG II in endothelium-intact rings. However, ANG II-induced contraction in the presence of L-NAME was significantly greater in obese than in lean rats (Emax: lean, 535 ± 14 vs. obese, 818 ± 23 g tension/g tissue). Similar to the effect of endothelium removal, the presence of L-NAME decreased pD2 of ANG II in lean rats but did not significantly affect obese rats. This effect of L-NAME on ANG II pD2 led to a significant difference (P < 0.05) between lean (8.0 ± 0.09) and obese (8.7 ± 0.30) rats, suggesting enhanced sensitivity to ANG II in obese compared with lean rats, similar to the effect of endothelium removal on ANG II sensitivity in these animals.


Figure 2
View larger version (11K):
[in this window]
[in a new window]

  		Fig. 2. Concentration-response curve of ANG II (10–9 to 10–7 M) with and without losartan in NG-nitro-L-arginine methyl ester (L-NAME)-treated aortic rings of lean and obese Zucker rats. Data are expressed as means ± SE (n = 15 for lean and 17 for obese). *P < 0.05, significantly different compared with lean rats (1-way ANOVA followed by Newman-Keuls post hoc test for multiple comparisons).

 
Effect of losartan on ANG II-induced contraction in aortic rings. For this purpose, we used endothelium-intact rings with L-NAME so that various endothelium-derived contracting factors remain intact, and NO production, which affects ANG II response, is blocked. Losartan (1 µM) completely blocked the ANG II-induced contraction in the aortic rings from both lean and obese rats (Fig. 2). This suggested the involvement of AT1 receptor in the ANG II-induced contractile response in both rat strains.

Effect of endothelin-1- and KCl-induced contraction. Because we observed that blocking the NOS with L-NAME causes greater contractile response to ANG II in aortic rings of obese compared with lean rats, we also examined endothelin-1-induced contraction in the aortic rings in the presence of L-NAME. In the aortic rings treated with L-NAME, endothelin-1 produced concentration-dependent contractions that were similar in both lean and obese rats (Fig. 3A).


Figure 3
View larger version (7K):
[in this window]
[in a new window]

  		Fig. 3. Concentration-response curve of endothelin-1 (10–9 to 10–7 M; A) and KCl (10–75 mM; B) in the aortic rings of lean and obese Zucker rats. Data are expressed as means ± SE (n = 18 for endothelin-1, n = 19 for lean and 34 for obese for KCl).

 
KCl was used as an additional vasoconstrictor to ascertain that the vascular rings from obese rats do not have an inherent tendency for greater vasoconstriction. KCl in a concentration (10–75 mM)-dependent manner also produced contractions to a similar extent in the endothelium-intact aortic rings of both lean and obese rats (Fig. 3B). The similar responses of aortic rings to endothelin-1 and KCl in both rat strains suggest that obese rat aortic rings do not posses inherent tendency to higher contractility compared with lean rats.

Relaxation responses to SNP and ACh in aortic rings. Relaxation responses to SNP and ACh were obtained in PE precontracted endothelium-intact aortic rings from lean and obese rats. SNP, an NO donor, produced a concentration-dependent relaxation (10–9 to 10–7 M) to similar extent in the aortic rings of lean and obese rats (Fig. 4A). However, ACh, a hormone that produces endothelium-dependent vasorelaxation, produced significantly greater relaxation in the aortic rings of obese than lean rats (Fig. 4B).


Figure 4
View larger version (8K):
[in this window]
[in a new window]

  		Fig. 4. Concentration-response curve of sodium nitroprusside (SNP, 10–9 to 10–7 M; A) and ACh (10–9 to 10–4 M; B) in the aortic rings of lean and obese Zucker rats. Data are expressed as means ± SE (n = 13 for lean and 16 for obese for SNP and n = 10 for lean and 20 for obese for ACh). *P < 0.05, significantly different compared with lean rats (1-way ANOVA followed by Newman-Keuls post hoc test for multiple comparisons).

 
Expression of the protein and mRNA for AT1 receptor and eNOS in aorta. The expression of the AT1 receptor and the eNOS in the aorta of lean and obese rats was determined by Western blotting. The antibody used for AT1 receptors recognized a single band of ~45 kDa (Fig. 5A), which has earlier been characterized for its specificity (3). Similarly, the antibody used for eNOS detected a single band of ~133 kDa (Fig. 6A). The mRNA for the AT1 receptor and eNOS in the aorta of lean and obese Zucker rats was determined by RT-PCR. GAPDH mRNA was used as an internal control. The specific primers produced expected fragments of 305, 450, and 723 bp for AT1, eNOS, and GAPDH, respectively (Figs. 5B and 6B). The normalized values for AT1 with GAPDH show a significantly greater expression of the AT1 receptor protein and mRNA (Fig. 5, A and B) in the endothelium-intact vascular rings, whereas, in the endothelium-denuded vascular rings, the AT1 receptor protein and mRNA was modestly but not significantly higher in obese compared with lean Zucker rats (Fig. 5, A and B). On the other hand, the expression of both the protein and the mRNA of eNOS was significantly increased in the aorta of obese compared with lean Zucker rats (Fig. 6, A and B).


Figure 5
View larger version (16K):
[in this window]
[in a new window]

  		Fig. 5. A, top: representative Western blot showing ANG II type 1 (AT1) receptor and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) protein expression in the aorta of lean and obese Zucker rats. A, bottom: bar graph showing the densitometric analysis of AT1 receptor normalized with GAPDH. Values are means ± SE; n = 5. *P < 0.05 compared with lean rats (Student's t-test). B, top: representative semiquantitative RT-PCR of AT1 receptors and GAPDH in the aorta of lean and obese Zucker rats. B, bottom: bar graph showing the densitometric analysis of AT1 receptor normalized with GAPDH. Values are expressed as means ± SE of aortic rings from 3 lean and 5 obese rats. *P < 0.05 compared with lean Zucker rats (Student's t-test).

 

Figure 6
View larger version (14K):
[in this window]
[in a new window]

  		Fig. 6. A, top: representative Western blot showing endothelial nitric oxide synthase (eNOS) and GAPDH protein expression in the aorta of lean and obese Zucker rats. A, bottom: bar graph showing the densitometric analysis of eNOS normalized with GAPDH. Values are means ± SE; n = 4 for lean and 5 for obese. *P < 0.05 compared with lean Zucker rats (Student's t-test). B, top: representative semiquantitative RT-PCR of eNOS and GAPDH in the aorta of lean and obese Zucker rats. B, bottom: bar graph showing the densitometric analysis of eNOS normalized with GAPDH. Values are expressed as means ± SE of aortic rings from 3 lean and 5 obese rats. *P < 0.05 compared with lean Zucker rats (Student's t-test).

 

   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
ANG II is a potent vasoconstrictor that, through the activation of the AT1 receptors, regulates vascular tone and thereby regulates blood pressure (28, 38). An enhanced function of the AT1 receptors is implicated in the development of hypertension (1). Obesity is a high risk factor for the development of hypertension (18), and enhanced activity of RAS is reported in obese animals (1, 17) as well as in humans (36, 39). The obese Zucker rat is a model of obesity and insulin resistance, a characteristic of type 2 diabetes. There are conflicting reports as to the effects of ANG II on the vascular tone in obese compared with lean rats. Some reports suggested no difference (19) while other studies showed that ANG II caused a greater vasoconstriction in obese than in lean rats (27).

Endothelium dysfunction is associated with various diseases, including obesity, diabetes, and hypertension (29, 31). Because ANG II is shown to induce the release of NO (5, 30, 32), an endothelium-derived relaxing factor, and endothelin-1 (13, 15), an endothelium-derived contraction factor, a role of endothelium and endothelium-derived factors was determined on ANG II-induced vasoconstriction in the isolated aorta of obese and lean Zucker rats.

ANG II-induced vasoconstriction was modestly higher, but not significant, in endothelium-intact aortic rings of obese compared with lean rats. When the endothelium was removed, ANG II elicited greater vasoconstriction in the obese rat aortic rings compared with lean rats. Endothelial cells are known to express AT1 receptors (8), which upon activation stimulate NO production and affect ANG II-elicited vascular tone (5, 30, 32). It is interesting to note that the removal of endothelium or addition of L-NAME, an NOS inhibitor, not only increased ANG II-induced contraction (Emax) per se in both the lean and obese rats, but the increase was more pronounced in obese than lean rats. Additionally, removal of endothelium or addition of NOS inhibitor caused an increase in vascular sensitivity to ANG II that was evident by an increase in pD2 of ANG II in obese rats, whereas the vascular sensitivity to ANG II in lean rats was decreased (decreased pD2) after endothelium removal or NOS inhibition. These findings suggest that an enhanced NO production mediated by the activation of the endothelial AT1 receptor may be responsible for protecting against the increased ANG II-induced vasoconstriction in obese Zucker rats.

Because AT1 receptor expression in the endothelium-denuded aortic rings was only modestly increased in obese compared with lean rats, the enhanced AT1 receptor function in obese rats could be attributed, in part, to the postreceptor cellular mechanisms. Other studies have shown significantly greater AT1 receptor function, although there was modest to no change in the AT1 receptor levels in hypertensive rat models compared with control rats (7, 23, 34). ANG II produces significantly greater stimulation of the Na-K-ATPase activity in renal proximal tubules of obese compared with lean rats, but the AT1 receptor levels were only modestly higher in obese rats (34). Similarly, the renal AT1 receptor levels in the spontaneously hypertensive (SHR) and normotensive rats were similar (7), yet renal sensitivity to ANG II was greater in SHR (23). However, in the present study, we found that expression of the AT1 receptor and eNOS in endothelium-intact vascular rings is increased. The coupling of the endothelial AT1 receptors with eNOS may cause an enhanced production of NO in response to ANG II (30, 32). Therefore, it is likely that the enhanced NO production by endothelial AT1 receptor compensates the higher sensitivity of the smooth muscle AT1 receptors to ANG II, resulting in similar contractile response to ANG II in endothelium-intact vascular rings of obese compared with lean rats.

ANG II is also reported to stimulate the production of endothelin-1 (13), which could affect the ANG II-induced vascular response. Because endothelin-1 is released predominantly from endothelial cells, the enhanced ANG II vasoconstrictor response in endothelium-denuded rings suggests that endothelin-1 may not be differentially affecting the ANG II-induced response. This notion is further supported by the observation that endothelin-1-induced contractions were similar in lean and obese Zucker rats. It is known that NO produced in the endothelial cells diffuses into the smooth muscle cells and induces vasorelaxation via a cGMP-mediated pathway (2). To test whether it was the increased NO production or post-NO signaling, we studied the relaxation of PE precontracted rings to ACh, a hormone known to cause endothelium-dependent relaxation by activating the NOS (40), and SNP, an NO donor. We found that, although while ACh-elicited vasodilatation was greater in obese aortic rings, the SNP-induced response was similar in both rat strains. An enhanced vasorelaxation response to ACh in the obese Zucker rats of the same age group that we have used in our studies has also been reported by other workers (11, 33). This further suggested that it is the enhanced NO production, and not the downstream signaling components, that may be responsible for the greater endothelium-mediated functions in obese rats.

Although a higher endothelial function through the production of NO may provide a buffer against the higher AT1 receptor function, a compromised function of endothelium in the advance stage of obesity/diabetes may exacerbate the AT1 receptor function for its vasoconstriction activity and thereby contribute to arterial hypertension. The enhanced function of AT1 receptors in the smooth muscles may also have a consequence on insulin resistance in obese rats. Treatment of obese rats with AT1 receptor antagonist has been shown to improve insulin sensitivity (20). As it relates to a correlation between the RAS hyperactivity and obesity-related hypertension in various animal models, including obese Zucker rats (1, 18), the present findings suggest that the overall contractile response to ANG II is same, and the arterial AT1 receptor, at least in this age group of obese Zucker rats, may not be a contributing factor to hypertension. Other mechanisms, such as enhanced anti-natriuretic function of renal ANG II via enhanced tubular AT1 receptor function as shown in obese Zucker rats, may be responsible for contributing to hypertension in these animals (1, 4, 16, 34, 37).

In summary, ANG II-induced vasoconstriction in denuded aortic rings was higher in obese than lean Zucker rats. The higher ANG II response may be because of an increase in the postreceptor cellular signaling associated with AT1 receptor in the endothelium-denuded rings. However, an enhanced endothelial AT1 receptor function possibly via greater NO production seems to counteract the enhanced ANG II-induced vasoconstriction in the aortic rings of obese Zucker rats.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant RO1 DK-061578.


   FOOTNOTES
 

Address for reprint requests and other correspondence: T. Hussain, Dept. of Pharmacological and Pharmaceutical Sciences, Science and Research Bldg. 2, Univ. of Houston, 4800 Calhoun, Houston, TX 77204-5037 (e-mail: Thussain2{at}uh.edu)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Alonso GM, Brands MW, Zappe DH, Hall JE. Hypertension in obese Zucker rats. Role of angiotensin II and adrenergic activity. Hypertension. 28: 1047–1054, 1996.[Abstract/Free Full Text]
  2. Archer SL, Huang JM, Hampl V, Nelson DP, Shultz PJ, Weir EK. Nitric oxide and cGMP cause vasorelaxation by activation of a charybdotoxin-sensitive K channel by cGMP dependent protein kinase. Proc Natl Acad Sci USA 91: 7583–7587, 1994.[Abstract/Free Full Text]
  3. Banday AA, Siddiqui AH, Menezes MM, Hussain T. Insulin treatment enhances AT1 receptor function in OK cells. Am J Physiol Renal Physiol 288: F1213–F1219, 2005.[Abstract/Free Full Text]
  4. Becker M, Umrani D, Lokhandwala MF, Hussain T. Increased renal angiotensin II AT1 receptor function in obese Zucker rat. Clin Exp Hypertens 25: 35–47, 2003.[CrossRef][ISI][Medline]
  5. Boulanger CM, Caputo L, Levy BI. Endothelilal AT1-mediated release of nitric oxide decreases angiotensin II contractions in rat carotid artery. Hypertension 26: 752–757, 1995.[Abstract/Free Full Text]
  6. Chen L, McNeill JR, Wilson TW, Gopalakrishnan V. Heterogeneity in vascular smooth muscle responsiveness to angiotensin II. Role of endothelin. Hypertension 26: 83–88, 1995.[Abstract/Free Full Text]
  7. Cheng HF, Wang JL, Vinson GP, Harris RC. Young SHR express increased type 1 angiotensin II receptors in renal proximal tubule Am J Physiol Renal Physiol 274: F10–F17, 1998.[Abstract/Free Full Text]
  8. Chiu AT, McCall DE, Ardecky RJ, Duncia JV, Nguyen TT, Timmermans PB. Angiotensin II receptor subtypes and their selective nonpeptide ligands. Receptor 10: 33–40, 1990.
  9. Chung O, Csikos T, Unger T. Angiotensin II receptor pharmacology and AT1-receptor blockers. J Hum Hypertension, Suppl 1: S11–S20, 1999.
  10. Cooper R, McFarlane AN, Bennett FI, Wilks R, Puras A, Tewksbury D, Ward R, Forrester T. ACE, angiotensinogen and obesity: a potential pathway leading to hypertension. J Hum Hypertens 11: 107–111, 1997.[CrossRef][ISI][Medline]
  11. Cox RH, Kikta DC. Age related changes in thoracic aorta of obese Zucker rats. Am J Physiol Heart Circ Physiol 262: H1548–H1556, 1992.[Abstract/Free Full Text]
  12. de Gasparo M, Catt KJ, Inagami T, Wright JW, Unger TH. International Union of Pharmacology XXIII: the angiotensin receptors. Pharmacol Rev 52: 415–472, 2000.[Abstract/Free Full Text]
  13. Emori T, Hirata Y, Ohta K, Shichiri M, Marumo F. Secretory mechanism of immunoreactive endothelin in cultured bovine endothelial cells. Biochem Biophys Res Commun 160: 93–100, 1989.[CrossRef][ISI][Medline]
  14. Gruetter CA, Ryan CA, Lemke SM, Bailly DA, Fox MK, Schoepp DD. Endothelium-dependent modulation of angiotensin II-induced contraction in blood vessels. Eur J Pharmacol 146: 85–95, 1988.[CrossRef][ISI][Medline]
  15. Hahn AW, Resink TJ, Scott-Burden T, Powell J, Dohi Y, Buhler FR. Stimulation of endothelin mRNA and secretion in rat vascular smooth muscle cells: a novel autocrine function. Cell Regul 1: 649–659, 1990.[ISI][Medline]
  16. Hakam AC, Hussain T. Renal angiotensin II type-2 receptors are upregulated and mediate the candesartan-induced natriuresis/diuresis in obese Zucker rats. Hypertension 45: 270–275, 2005.[Abstract/Free Full Text]
  17. Hall JE, Brands MW, Dixon WN, Smith MJ. Obesity-induced hypertension: renal function and systemic hemodynamics. Hypertension 22: 292–299, 1993.[Abstract/Free Full Text]
  18. Hall JE, Zappe DH, Alonso-Galicia M, Granger JP, Brands MW, Kassab SE. Mechanisms of obesity-induced hypertension. News Physiol Sci 11: 255–261, 1996.[Abstract/Free Full Text]
  19. Harker CT, O'Donnell MP, Kasiske BL, Keane WF, Katz SA. The renin-angiotensin system in the type II diabetic obese Zucker rat. J Am Soc Nephrol 4: 1354–1361, 1993.[Abstract]
  20. Henriksen EJ, Jacob S, Kinnick TR, Teachey MK, Krekler M. Selective angiotensin II receptor antagonism reduces insulin resistance in obese Zucker rats. Hypertension 38: 884–890, 2001.[Abstract/Free Full Text]
  21. Hussain T, Leigh JH, Mustafa SJ. Regulation of adenosine receptor function by theophylline in rat aorta. J Cardiovasc Pharmacol 24: 95–99, 1994.[ISI][Medline]
  22. Hussain T. Renal angiotensin II receptors, hyperinsulinemia and obesity. Clin Exp Hypertens 25: 395–403, 2003.[CrossRef][ISI][Medline]
  23. Kost CK Jr, Jackson EK. Enhanced renal angiotensin II subtype 1 receptor responses in the spontaneously hypertensive rat. Hypertension 21: 420–431, 1993.[Abstract/Free Full Text]
  24. Mark AL, Correia M, Morgan DA, Shaffer RA, Haynes WG. Obesity-induced hypertension: new concepts from the emerging biology of obesity. Hypertension 33: 537–541, 1999.[Abstract/Free Full Text]
  25. Matsubura H. Pathphysiological role of angiotensin II type 2 receptor in cardiovascular and renal diseases. Circ Res 52: 1182–1191, 1998.
  26. Mulrow PJ. The intrarenal renin-angiotensin system. Curr Opin Nephrol Hypertens 2: 41–44, 1993.[Medline]
  27. Nishimatsu H, Suzuki E, Satonaka H, Takeda R, Omata M, Fujita T, Nagai R, Kitamura T, Hirata Y. Endothelial dysfunction and hypercontractility of vascular myocytes are ameliorated by Fluvastatin in Obese Zucker Rats. Am J Physiol Heart Circ Physiol 288: H1770–H1776, 2005.[Abstract/Free Full Text]
  28. Oparil S, Haber E. The renin-angiotensin system. N Engl J Med 291: 389–401, 1974.[ISI][Medline]
  29. Panus C, Mota M, Vladu D, Vanghelie L, Raducanu CL. The endothelial dysfunction in diabetes mellitus. Rom J Intern Med 41: 27–33, 2003.[Medline]
  30. Patzak A, Lai EY, Mrowka R, Steege A, Persson PB, Persson EG. AT1 receptors mediate angiotensin II-induced release of nitric oxide in afferent arterioles. Kidney Int 66: 1949–1958, 2004.[CrossRef][ISI][Medline]
  31. Prasad A, Quyyumi A. Renin-angiotensin system and angiotensin receptor blockers in the metabolic syndrome. Circulation 110: 1507–1512, 2004.[Free Full Text]
  32. Pueyo ME, Arnal JF, Rami J, Michel JB. Angiotensin II stimulates the production of NO and peroxynitrite in endothelial cells. Am J Physiol Cell Physiol 274: C214–C220, 1998.[Abstract/Free Full Text]
  33. Sexl V, Manusci G, Raberger G, Schutz W. Age-related changes in vascular reactivity in genetically diabetic rats. Pharmacology 50: 238–246, 1995.[ISI][Medline]
  34. Shah S, Hussain T. Enhanced angiotensin II-induced activation of Na+, K+-ATPase in the proximal tubules of obese Zucker rats. Clin Exp Hypertens 28: 29–40, 2006.[CrossRef][ISI][Medline]
  35. Siragy HM, Inagami T, Ichiki T, Carey RM. Sustained hypersensitivity to angiotensin II and its mechanism in mice lacking the subtype-2 (AT2) angiotensin receptor. Proc Natl Acad Sci USA 96: 6506–6510, 1999.[Abstract/Free Full Text]
  36. Sowers JR, Nyby M, Stern N, Beck F, Baron S, Catania R, Vlachis N. Blood pressure and hormone changes associated with weight reduction in the obese. Hypertension 4: 686–691, 1982.[Abstract/Free Full Text]
  37. Tallam LS, Jandhyala BS. Significance of exaggerated natriuresis after angiotensin AT1 receptor blockade or angiotensin-converting enzyme inhibition in obese Zucker rats. Clin Exp Pharmacol Physiol 28: 433–440, 2001.[CrossRef][ISI][Medline]
  38. Timmermans PB, Wong PC, Chiu AT, Herblin WF, Benfiled P, Carini DJ, Lee RJ, Wexler RR, Saye JA, Smith RD. Angiotensin II receptors and angiotensin II receptors antagonists. Pharm Rev 45: 205–250, 1993.[ISI][Medline]
  39. Tuck ML, Sowers J, Dornfield L, Kledzik G, Maxwell M. The effect of weight reduction on blood pressure, plasma renin activity, and plasma aldosterone levels in obese patients. N Engl J Med 304: 930–933, 1981.[Abstract]
  40. Vanhoutte PM, Mombouli JV. Vascular endothelium: vasoactive mediators. Prog Cardiovasc Dis 39: 229–238, 1996.[CrossRef][ISI][Medline]



This article has been cited by other articles:


Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
J. H. Lee, S. Xia, and L. Ragolia
Upregulation of AT2 receptor and iNOS impairs angiotensin II-induced contraction without endothelium influence in young normotensive diabetic rats
Am J Physiol Regulatory Integrative Comp Physiol, July 1, 2008; 295(1): R144 - R154.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
292/4/H1722    most recent
00612.2006v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (2)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Siddiqui, A. H.
Right arrow Articles by Hussain, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Siddiqui, A. H.
Right arrow Articles by Hussain, T.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2007 by the American Physiological Society.