This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Wang, D. H. Articles by Hu, Z. Search for Related Content PubMed PubMed Citation Articles by Wang, D. H. Articles by Hu, Z. Pubmed/NCBI databases Compound via MeSH Substance via MeSH

(Hypertension. 1998;32:65-70.)
© 1998 American Heart Association, Inc.

Scientific Contributions

Differential Regulation of Angiotensin II Receptor Subtypes in the Adrenal Gland

Role of Aldosterone

Donna H. Wang; Jingxin Qiu; ; Zhaoyong Hu

From the Department of Internal Medicine, University of Texas Medical Branch, Galveston, Texas.

Correspondence to Donna H. Wang, MD, Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555-1065. E-mail dwang{at}utmb.edu

	Abstract

Top Abstract Introduction Methods Results Discussion References

 
Abstract—It has been shown that aldosterone potentiates the action of angiotensin II (Ang II) in cultured rat vascular smooth muscle cells solely by increasing the number of Ang II receptors. The mechanisms responsible for aldosterone–Ang II interactions in the adrenal gland are unknown. The present study was designed to investigate the effect of aldosterone on expression of Ang II receptor subtypes (AT₁ and AT₂) in the adrenal gland. Seven-week-old male Wistar rats were treated for 2 weeks with either aldosterone (0.05 µg/h, n=14) or vehicle (n=14) by use of implanted osmotic minipumps. Systolic blood pressure was not altered by aldosterone treatment. Plasma aldosterone levels were higher in aldosterone-treated rats (181±53 pg/mL) compared with vehicle-treated rats (33±21 pg/mL, P<0.05). Northern blot analysis and radioligand binding assay showed that adrenal AT₁ mRNA levels and AT₁ receptor density in aldosterone-treated rats were not statistically different from those of vehicle-treated rats. However, immunohistochemical studies showed that the highest adrenal AT₁ receptor expression was shifted from the zona glomerulosa to the zona fasciculata after aldosterone treatment. In contrast, adrenal AT₂ mRNA and AT₂ receptor density in aldosterone-treated rats were decreased by approximately 50% and 40%, respectively, compared with vehicle-treated rats (P<0.05). Aldosterone-induced decrease in adrenal AT₂ receptor expression occurred mainly in the medulla. Thus, aldosterone differentially modulates the expression of AT₁ and AT₂ receptors in the adrenal gland. Although the function of the AT₂ receptor in the adrenal gland is largely unknown, our data indicate that aldosterone may modulate the effect of Ang II by altering the location of AT₁ receptors and by reducing the number of AT₂ receptors in the adrenal gland.

Key Words: aldosterone • receptors, angiotensin II • adrenal gland • Northern blot • radioligand assay • immunohistochemistry

	Introduction

Top Abstract Introduction Methods Results Discussion References

 
The renin-angiotensin-aldosterone system is one of the most potent systems that regulate blood pressure, electrolyte balance, and extracellular fluid volume. The effects of the principal substances of this system, angiotensin II (Ang II) and aldosterone, are triggered by their interaction with specific receptors in a variety of tissues. In the adrenal gland, Ang II receptors are classified into two major subtypes: AT₁ and AT₂. The AT₁ receptor subtype accounts for 80% of the Ang II receptors in the rat and bovine adrenal gland, with the other 20% being the AT₂ receptor subtype.^{1 2} It has been shown that Ang II–induced aldosterone production is mediated by activation of the AT₁ receptor in the adrenal zona glomerulosa cells.^{3 4} Although the physiological role of the AT₂ receptor in the adrenal gland is largely unknown, it has been reported that Ang II stimulates secretion of endogenous ouabain from bovine adrenocortical cells via activation of the AT₂ receptor.⁵

Several lines of evidence have shown that modulation of the expression of the Ang II receptor is important in the regulation of Ang II action.^{6 7} We have recently shown that sodium deficiency increases the expression of genes encoding AT₁ receptor subtypes and elevates the AT₁ receptor density in the adrenal gland.⁸ The increase in AT₁ receptor density in the adrenal gland during sodium restriction may contribute to increased aldosterone production induced by Ang II. In addition, blockade of the binding of Ang II to the AT₁ receptor by losartan prevents the increase in mRNA expression of AT₁ receptor subtypes and AT₁ receptor density induced by sodium depletion, suggesting that these changes in the adrenal gland are mediated by activation of the renin-angiotensin system accompanying low sodium intake.⁸ Although it is known that sodium restriction also increases aldosterone production, and that aldosterone increases Ang II receptor density in cultured rat vascular smooth muscle cells,^{9 10} the effects of aldosterone on expression of Ang II receptor subtypes in the adrenal gland have not been defined. Therefore, in the present study, we used a combination of Northern blot, radioligand binding, and immunohistochemistry to test the hypothesis that infusion of aldosterone differentially regulates the expression of AT₁ and AT₂ receptors in the adrenal gland.

	Methods

Top Abstract Introduction Methods Results Discussion References

 
Animals and Treatment Groups
Seven-week-old male Wistar rats (Charles River Laboratories, Wilmington, Mass) weighing between 150 and 175 g were randomly divided into two groups: group 1 (Ald) was subcutaneously infused with aldosterone at 0.05 µg/h in saline plus 15% ethanol alcohol (n=14), and group 2 (Con) was subcutaneously infused with vehicle (saline plus 15% ethanol alcohol, n=14). It has been shown that aldosterone infused at 0.05 µg/h subcutaneously does not change systolic blood pressure in rats.¹¹ All the rats were anesthetized with a single intraperitoneal injection of ketamine hydrochloride (80 mg/kg) and xylazine (12 mg/kg). Alzet miniosmotic pumps (model 2002, Alza Corp) with a capacity of 226±6 µL and infusion rate of 0.5±0.02 µL/h were filled with either 0.9% saline plus 15% ethanol alcohol or aldosterone in 0.9% saline plus 15% ethanol alcohol at a concentration that was adjusted depending on the weight of the rat.¹² The pumps were implanted subcutaneously between the scapulae. Sterile technique was used, and the rats were given penicillin G (10 000 U IM) after the surgery. The rats were housed in pairs, fed regular food and tap water ad libitum, and maintained on a 12-hour light/dark cycle for a total of 14 days.

Systolic Blood Pressure
Indirect tail-cuff blood pressure measurements were routinely obtained in all rats to assess the blood pressure using a Narco Bio-Systems Electro-Sphygmomanometer. The pressures were measured in conscious rats every 3 days for 14 days, beginning 1 day before the placement of the minipumps. The blood pressure value for each rat was calculated as the average of 5 separated measurements at each session.

Plasma Aldosterone Levels
The rats were decapitated and bled into chilled EDTA tubes. Blood samples were centrifuged at low temperatures (4°C) to obtain plasma for aldosterone assay. Plasma aldosterone concentration was then determined by using a commercially available radioimmunoassay kit for aldosterone (Diagnostic Products Corp).

Tissue Preparation
To obtain tissue for Northern blot and radioligand binding assay, a midline abdominal incision was made, and adrenal glands were promptly removed, frozen in liquid nitrogen, and stored at -80°C. Adrenal glands from 7 rats in each group were used for RNA extraction for Northern blot analysis, and adrenal tissues from the other 7 rats were used for the radioligand binding assay.

AT₁ and AT₂ cDNA and Probe Preparation
cDNA probes were prepared as described previously.^{8 13} Briefly, a 0.8-kb fragment (-178 to +562) from the coding region of rat AT_1A cDNA¹⁴ (a generous gift from Dr Tadashi Inagami, Vanderbilt University, Nashville, Tenn) was used as a template to make AT₁ probes. Because this fragment contains the AT_1A coding region where AT_1A and AT_1B cDNAs exhibit high nucleotide sequence identity, these probes detect both AT_1A and AT_1B. A 1.23-kb fragment (+16 to +1249) from the coding region of rat AT₂ cDNA¹⁵ was used as a template to make AT₂ probes. The probes were labeled with ³²P-dCTP using a multiprimer DNA labeling system (Amersham Co) to a specific activity of 3x10⁸ cpm/µg. The labeled probes were separated from unincorporated nucleotides by Mini-Spin G-50 DNA purification spin columns (Worthington Biochemical).

RNA Extraction and Northern Blots
Total RNA of adrenal gland was extracted using the guanidine thiocyanate-phenol-chloroform extraction protocol.¹⁶ Electrophoresis of 20 µg denatured RNA from each preparation was carried out in a 1% agarose gel containing 2.2 mol/L formaldehyde. RNA was transferred to a positively charged nylon membrane (Fisher Co). The membrane was baked at 80°C for 2 hours in a vacuum oven (Fisher Co). After prehybridization for 5 hours at 42°C in 50% deionized formamide, 5x Denhardt's solution, 5x SSC, 0.5% SDS, and 200 µg/mL denatured salmon sperm DNA, the membrane was hybridized with the ³²P-labeled probes for 18 to 20 hours at 42°C. The blot was then washed successively in 2x, 1x, and 0.5x SSC (two times, 10 minutes each) containing 0.1% SDS at 65°C. To control for differences in RNA loading, Northern blots were incubated at 90°C for 10 minutes in 20 mmol/L Tris-HCl (pH 8.0) to strip off the cDNA probes and rehybridized with ³²P-labeled probe for 18S rRNA. Blots were exposed to XAR-5 x-ray film (Eastman Kodak Co) with two intensifying screens. Autoradiographic signals were scanned with laser densitometry (Ultroscan XL, Pharmacia). Results of relative gene expression are expressed as the ratios of AT₁ mRNA and AT₂ mRNA to 18S rRNA.

Ligand Binding Assay
Whole adrenal tissues were homogenized in hypotonic buffer (20 mmol/L sodium phosphate, pH 7.1 to 7.2). Homogenates were then centrifuged at 48 000g for 20 minutes at 4°C. Cell membranes were resuspended in assay buffer (50 mmol/L sodium phosphate, 150 mmol/L NaCl, 1 mmol/L EDTA, 0.1 mmol/L bacitracin, pH 7.2) and recentrifuged. After resuspension in assay buffer, an aliquot of the cell membrane suspension was used for protein assay using a modified Bradford method (Bio-Rad).¹⁷ To measure the AT₁ receptor density, 20 µg protein was incubated with 125 pmol/L to 2 nmol/L ¹²⁵I-[Sar,Ile]Ang II (kindly provided by Dr Robert C. Speth, Washington State University) in a final volume of 200 µL assay buffer containing 0.1% bovine serum albumin in the presence or absence of the specific AT₂ receptor antagonist PD123319 (10 µmol/L).⁸ To measure the AT₂ receptor density, ¹²⁵I-[Sar,Ile]Ang II was used in the presence or absence of the specific AT₁ receptor antagonist losartan (10 µmol/L). Nonspecific binding was measured in the presence of 1 µmol/L unlabeled Ang II. Binding assays were performed for 120 minutes at room temperature and followed by immediate filtration through glass-fiber filters (Whatman GF/C). The filter-bound radioactivity was counted in a gamma spectrometer (Beckman, LS 3801). Receptor affinity and concentration were calculated by Scatchard analysis using GraphPAD InStat software.

Immunohistochemistry
Immunohistochemistry was performed as described previously.^{12 18 19} Briefly, the adrenal gland was fixed in 4% paraformaldehyde–0.1 mol/L phosphate buffer, pH 7.4, at 4°C overnight and was embedded in paraffin. Tissue sections (5 µm) were cut and mounted on slides. The endogenous peroxidase was blocked with 0.3% hydrogen peroxide in methanol, and nonspecific binding sites of secondary goat antibody were blocked with 3% normal goat serum and 1% nonfat dry milk in PBS. The sections were sequentially incubated at room temperature with primary antibody diluted in 1.5% normal goat serum and 0.5% nonfat dry milk in PBS [1:100 for AT₁ receptor antibody (Santa Cruz Biotechnologies Inc) and 1:100 for AT₂ receptor antibody (a generous gift of Dr Robert M. Carey, University of Virginia Health Sciences Center, Charlottesville)],¹⁹ biotinylated antibody, and avidin-peroxidase (ABC kit, Vector Laboratories). Staining was visualized with diaminobenzidine (Fast DAB tablets, Sigma). Slides were counterstained with hematoxylin.

Statistical Analysis
Results are expressed as mean±SE. The data were analyzed by either unpaired Student's t test (between 2 groups) or 2-way ANOVA followed by Tukey-Kramer multiple comparison test (for multiple groups). Differences were considered statistically significant at P<0.05.

	Results

Top Abstract Introduction Methods Results Discussion References

 
There was no significant difference in body weight between the two groups of rats at the end of the experiment (Con, 243±5 g; Ald, 240±4 g). Systolic blood pressures were not modified by the infusion of aldosterone compared with vehicle (Con, 112±6 mm Hg; Ald, 107±4 mm Hg at day 1; Con, 120±3 mm Hg; Ald, 125±2 mm Hg at the end of the experiment).

Plasma aldosterone levels were measured in each of the two groups to evaluate the effectiveness of aldosterone administration. Plasma aldosterone levels were approximately 5 times higher in aldosterone-infused rats (181±53 pg/mL) than in vehicle-treated rats (33±21 pg/mL) (P<0.05).

AT₁ and AT₂ receptor mRNA content in the adrenal gland of each of the two groups of rats was determined by Northern blot analysis. Blots were then stripped and rehybridized to 18S rRNA probes. Densitometric analysis (Figure 1) indicated that the ratio of AT₁ mRNA to 18S rRNA was not statistically different between aldosterone-infused (1.24±0.28) and vehicle-treated (0.92±0.14) rats. In contrast, densitometric analysis (Figure 2) indicated that the ratio of AT₂ mRNA to 18S rRNA in aldosterone-infused rats (0.70±0.14) was significantly decreased compared with vehicle-treated rats (1.36±0.15).

View larger version (26K): [in this window] [in a new window]   Figure 1. Northern blot analysis of AT1 mRNA levels in the adrenal gland in aldosterone- and vehicle-treated rats. Results are expressed as mean±SE; n=6 rats per group.

View larger version (20K): [in this window] [in a new window]   Figure 2. Northern blot analysis of AT2 mRNA levels in the adrenal gland in aldosterone- and vehicle-treated rats. Results are expressed as mean±SE; n=6 rats per group. *P<0.05 vs control.

The maximal binding (B_max) and dissociation constants (K_d) of the AT₁ and AT₂ receptors in the adrenal gland of each group were determined with the use of radioligand binding assay. The calculated AT₁ B_max (Figure 3) was not altered by aldosterone treatment (944±129 fmol/mg protein) compared with control (1016±134 fmol/mg protein). In contrast, the calculated AT₂ B_max (Figure 4) was decreased approximately 40% by aldosterone treatment (429±61 fmol/mg protein) compared with vehicle treatment (698±91 fmol/mg protein, P<0.05). There was no significant difference in the binding constants of both AT₁ and AT₂ receptors among adrenal preparations from two groups (Table).

View larger version (28K): [in this window] [in a new window]   Figure 3. AT1 receptor density in rats treated with aldosterone or vehicle. Protein (20 µg) was incubated with 125I-labeled [Sar,Ile]Ang II (125 pmol/L to 2 nmol/L) in the presence or absence of 10 µmol/L PD123319. Results are expressed as mean±SE; n=5 rats per group.

View larger version (17K): [in this window] [in a new window]   Figure 4. AT2 receptor density in rats treated with aldosterone or vehicle. Protein (20 µg) was incubated with 125I-labeled [Sar,Ile]Ang II (125 pmol/L to 2 nmol/L) in the presence or absence of 10 µmol/L losartan. Results are expressed as mean±SE; n=5 rats per group. *P<0.05 vs control.

View this table: [in this window] [in a new window]   Table 1. Kd Values of AT1 and AT2 Receptors in 2 Experimental Groups

Immunohistochemical localization of the AT₁ and AT₂ receptors in the adrenal gland was determined in both groups (Figure 5). The AT₁ receptor immunohistochemical signal was detected strongly in the zona glomerulosa in vehicle-treated rats (Figure 5A and 5C). AT₁ receptor signal also was detected in the zona fasciculata, zona reticularis, and medulla of vehicle-treated rats (Figure 5A). In contrast, heavy immunohistochemical staining for the AT₁ receptor was observed in the zona fasciculata in aldosterone-infused rats (Figure 5B and 5D). Adrenal staining for the AT₁ receptor was reduced but remained detectable in the zona glomerulosa, zona reticularis, and medulla of aldosterone-infused rats (Figure 5B).

View larger version (116K): [in this window] [in a new window]   Figure 5. Representative photomicrographs show immunostaining of the AT1 (A, B, C, D) and AT2 (E, F, G, H) receptors in the adrenal gland of vehicle-treated (A, C, E, G, I) and aldosterone-treated (B, D, F, H, J) rats. Magnification x40 for A, B, E, F, I, and J; x200 for C, D, G, and H. I and J show method controls in which AT1 (I) and AT2 (J) receptor antibody was replaced by normal rabbit IgG. CO indicates cortex; ME, medulla; ZG, zona glomerulosa; and ZF, zona fasciculata. Positive staining of the AT1 receptor is observed mainly in the zona glomerulosa in vehicle-treated rats (C) and in the zona fasciculata in aldosterone-treated rats (D). Positive staining of the AT2 receptor is observed mainly in the medulla of both vehicle-treated and aldosterone-treated rats (G and H). Method controls (I and J) are negative.

The AT₂ receptor immunohistochemical signal was detected strongly in the adrenal medulla in vehicle-treated rats (Figure 5E and 5G). AT₂ receptor signal also was detected in the adrenal zona glomerulosa and zona fasciculata but was absent from the zona reticularis of vehicle-treated rats (Figure 5E). Aldosterone treatment reduced immunohistochemical staining for the AT₂ receptor in the medulla (Figure 5F and 5H). Method controls in which AT₁ (Figure 5I) and AT₂ (Figure 5J) receptor antibody was replaced by normal rabbit IgG were negative.

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
Although it is well known that Ang II is a principal regulator of aldosterone production in the adrenal gland, knowledge of the feedback regulation of aldosterone on the action of Ang II in the adrenal gland remains incomplete. The present experiment was designed to test the hypothesis that infusion of aldosterone differentially regulates the expression of AT₁ and AT₂ receptors in the adrenal gland. We found that aldosterone shifts adrenal AT₁ receptor expression without altering total AT₁ receptor density and significantly decreases adrenal AT₂ receptor mRNA content and AT₂ receptor density. This appears to be the first indication of a role for aldosterone as a regulator of the action of Ang II through modulation of AT₁ and AT₂ receptor expression in the adrenal gland.

The dose of aldosterone used was carefully chosen from analysis of the literature so that blood pressure responses in our experiment would be comparable to those of other reports. Accordingly, we chose a 0.05-µg/h dose of aldosterone because, in agreement with other reports,¹¹ this dose did not cause detectable changes in systolic blood pressure in aldosterone-infused rats compared with rats infused with vehicle only. In addition, this dose allowed discrimination of effects of aldosterone on AT₁ and AT₂ receptor expression in the adrenal gland because plasma aldosterone levels were approximately 5 times higher in aldosterone-infused compared with vehicle-infused rats.

Northern blot analysis and radioligand binding study show that adrenal AT₁ receptor mRNA levels and receptor density are not altered by aldosterone infused at the specific dose used in this experiment. We have previously shown that sodium deficiency increases both AT₁ receptor mRNA content and AT₁ receptor density in the adrenal gland.⁸ These increases appear to be mediated by activation of the AT₁ receptor by Ang II, since blockade of the AT₁ receptor with losartan prevents these increases induced by low sodium intake.⁸ Likewise, Lehoux et al²⁰ have shown that low sodium or high potassium intake increases adrenal AT₁ receptor expression, and they suggested that increased aldosterone secretion induced by low sodium or high potassium intake involves increases in AT₁ receptor mRNA levels in the adrenal gland. Furthermore, Ullian et al⁹ have shown that in cultured rat vascular smooth muscle cells, aldosterone potentiates Ang II–stimulated, phospholipase C–dependent intracellular signals solely by coupling to an increased number of Ang II receptors. Although adrenal AT₁ receptor density is not altered by aldosterone infusion in the present experiment, it appears that there is a shift in adrenal AT₁ receptor expression in aldosterone-infused rats, indicated by immunohistochemical studies; ie, the highest AT₁ receptor expression is relocated from the zona glomerulosa to the zona fasciculata after aldosterone treatment. Because AT₁ receptors in the zona glomerulosa mediate Ang II–induced secretion of aldosterone,^{3 4} it is tempting to speculate that attenuated AT₁ receptor expression in the zona glomerulosa induced by aldosterone infusion represents a means of negative feedback regulation in aldosterone secretion.

The suppressive effects of aldosterone on AT₂ receptor mRNA content and receptor density in the adrenal gland have not been previously reported and deserve comment. It is well known that aldosterone alters plasma electrolyte concentrations and levels of neurohormonal factors.²¹ It is possible that aldosterone-induced changes in these parameters have direct or indirect effects on adrenal AT₂ receptor expression.

It has been shown in cultured bovine adrenal cells and PC12W cells that Ang II downregulates both AT₁ and AT₂ receptors through different mechanisms; AT₁ receptor is regulated through internalization/degradation of the occupied receptor and inhibition of transcription, whereas AT₂ receptor is regulated mainly by decrease in the stability of its mRNA.²² Moreover, results from the same group showed that the phorbol ester phorbol 12-myristate 13-acetate decreases both AT₂ mRNA and receptor binding on PC12W cells, suggesting that the hormonal regulation of AT₂ receptors is mediated through protein kinase C activation.²² Because it is difficult to separate the effects of aldosterone-induced changes in plasma electrolyte concentrations and levels of neurohormonal factors on AT₂ receptor expression in vivo, future studies using in vitro models may provide insight into the mechanisms by which aldosterone inhibits adrenal AT₂ receptor expression.

Although the function of the AT₁ receptor in the adrenal gland is relatively clear,¹ the function of the adrenal AT₂ receptor is largely unknown. It has been shown that Ang II stimulates secretion of endogenous ouabain from cultured bovine adrenocortical cells via activation of the AT₂ receptor.⁵ Furthermore, in the rat zona glomerulosa and PC12W cells, stimulation of AT₂ receptors appears to reduce guanylate cyclase activity.²³ Because modulation of the expression of the Ang II receptor is associated with changes in the Ang II action,^{6 7} it is conceivable that aldosterone may modulate Ang II action by suppressing the expression of the AT₂ receptor in the adrenal gland. This possibility awaits further in vivo and in vitro investigation.

In conclusion, we have demonstrated that infusion of aldosterone shifts adrenal AT₁ receptor expression without decreasing AT₁ receptor density. Moreover, aldosterone infusion decreases adrenal AT₂ receptor mRNA and AT₂ receptor density. Aldosterone-induced modulation of AT₁ and AT₂ receptor expression in the adrenal gland may be important in the adaptation to low salt diets and other conditions in which aldosterone is increased.

	Acknowledgments

 
This study was supported in part by National Institutes of Health grant HL-52279 (Dr Wang). We thank DuPont Merck and Parke-Davis Pharmaceuticals for providing losartan and PD123319. The authors are indebted to Drs Tadashi Inagami of Vanderbilt University for provision of the AT₁ receptor cDNA, Robert C. Speth of Washington State University for ¹²⁵I- [Sar,Ile]Ang II, and Robert M. Carey of the University of Virginia for AT₂ receptor antibody.

Received February 3, 1998; first decision February 23, 1998; accepted February 27, 1998.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Balla T, Baukal AJ, Eng S, Catt KJ. Angiotensin II receptor subtypes and biological responses in the adrenal cortex and medulla. Mol Pharmacol. 1991;40:401–406.[Abstract]

2. Quali R, LeBrethon MC, Saez JM. Identification and characterization of angiotensin II receptor subtypes in cultured bovine and human adrenal fasciculata cells and PC12W cells. Endocrinology. 1993;133:2766–2772.[Abstract/Free Full Text]

3. Gupta P, Franco-Saenz R, Mulrow PJ. Locally generated angiotensin II in the adrenal gland regulates basal, corticotropin-, and potassium-stimulated aldosterone secretion. Hypertension. 1995;25:443–448.[Abstract/Free Full Text]

4. Timmermans PBMWM, Benfield P, Chiu AT, Herblin WF, Wong PC, Smith RD. Angiotensin II receptors and functional correlates. Am J Hypertens. 1992;5:221S–235S.[Medline] [Order article via Infotrieve]

5. Laredo J, Shah JR, Lu ZR, Hamilton BP, Hamlyn JM. Angiotensin II stimulates secretion of endogenous ouabain from bovine adrenocortical cells via angiotensin type 2 receptors. Hypertension. 1997;29(pt 2):401–407.

6. Ullian ME, Linas SL. Angiotensin II surface receptor coupling to inositol triphosphate formation in vascular smooth muscle cells. J Biol Chem. 1990;265:195–200.[Abstract/Free Full Text]

7. Ullian ME, Schelling JR, Linas SL. Aldosterone enhances angiotensin II receptor binding and inositol phosphate responses. Hypertension. 1992;20:67–73.[Abstract/Free Full Text]

8. Du Y, Guo DF, Inagami T, Speth RC, Wang DH. Regulation of ANG II-receptor subtype and its gene expression in adrenal gland. Am J Physiol. 1996;271:H440–H446.[Abstract/Free Full Text]

9. Ullian ME, Fine JJ. Mechanisms of enhanced angiotensin II-stimulated signal transduction in vascular smooth muscle by aldosterone. J Cell Physiol. 1994;161:201–208.[Medline] [Order article via Infotrieve]

10. Ullian ME, Hutchison FN, Hazen-Martin DJ, Morinelli TA. Angiotensin II-aldosterone interactions on protein synthesis in vascular smooth muscle cells. Am J Physiol. 1993;264:C1525–C1531.[Abstract/Free Full Text]

11. Garwitz ET, Jones AW. Aldosterone infusion into the rat and dose-dependent changes in blood pressure and arterial ionic transport. Hypertension. 1982;4:374–381.[Abstract/Free Full Text]

12. Wang DH, Prewitt RL, Beebe SJ. Regulation of PDGF-A: a possible mechanism for angiotensin II-induced vascular growth. Am J Physiol. 1995;269:H356–H364.[Abstract/Free Full Text]

13. Elijovich F, Zhao HW, Laffer CL, Du Y, DiPette DJ, Inagami T, Wang DH. Regulation of growth of the adrenal gland in DOC-salt hypertension: role of angiotensin II receptor subtypes. Hypertension. 1997;29(pt 2):408–413.

14. Iwai N, Yamano Y, Chaki S, Konishi F, Bardhan S, Tibbetts C, Sasaki K, Hasegawa M, Matsuda Y, Inagami T. Rat angiotensin II receptor: cDNA sequence and regulation of the gene expression. Biochem Biophys Res Commun. 1991;177:299–304.[Medline] [Order article via Infotrieve]

15. Kambayashi Y, Bardhan S, Takahashi K, Tsuzuki S, Inui H, Hamakubo T, Inagami T. Molecular cloning of a novel angiotensin II receptor isoform involved in phosphotyrosine phosphatase inhibition. J Biol Chem. 1993;268:24543–24546.[Abstract/Free Full Text]

16. Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987;162:156–159.[Medline] [Order article via Infotrieve]

17. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–254.[Medline] [Order article via Infotrieve]

18. Paxton WG, Runge M, Horaist C, Cohen C, Alexander RW, Bernstein WG. Immunohistochemical localization of rat angiotensin II AT1 receptor. Am J Physiol. 1993;264:F989–F995.[Abstract/Free Full Text]

19. Ozono R, Wang ZQ, Moore AF, Inagami T, Siragy HM, Carey RM. Expression of the subtype 2 angiotensin (AT₂) receptor protein in rat kidney. Hypertension. 1997;30:1238–1246.[Abstract/Free Full Text]

20. Lehoux JG, Bird IM, Rainey WE, Tremblay A, Ducharme L. Both low sodium and high potassium intake increase the level of adrenal angiotensin II receptor type 1, but not that of adrenocorticotropin receptor. Endocrinology. 1994;134:776–782.[Abstract/Free Full Text]

21. Young DB. Quantitative analysis of aldosterone's role in potassium regulation. Am J Physiol. 1988;255:F811–F822.[Abstract/Free Full Text]

22. Ouali R, Berthelon MC, Begeot M, Saez JM. Angiotensin II receptor subtypes AT1 and AT2 are down-regulated by angiotensin II through AT1 receptor by different mechanisms. Endocrinology. 1997;138:725–733.[Abstract/Free Full Text]

23. Bottari SP, King IN, Reichilin S, Dahlstroem I, Lydon N, de Gasparo M. The angiotensin AT2 receptor stimulates protein tyrosine phosphatase activity and mediates inhibition of particulate guanylate cyclase. Biochem Biophys Res Commun. 1992;183:206–211.[Medline] [Order article via Infotrieve]

This article has been cited by other articles:

				F. Xiao, J. R. Puddefoot, S. Barker, and G. P. Vinson Mechanism for Aldosterone Potentiation of Angiotensin II-Stimulated Rat Arterial Smooth Muscle Cell Proliferation Hypertension, September 1, 2004; 44(3): 340 - 345. [Abstract] [Full Text] [PDF]

				A. F. Moore, N. T. Heiderstadt, E. Huang, N. L. Howell, Z.-Q. Wang, H. M. Siragy, and R. M. Carey Selective Inhibition of the Renal Angiotensin Type 2 Receptor Increases Blood Pressure in Conscious Rats Hypertension, May 1, 2001; 37(5): 1285 - 1291. [Abstract] [Full Text] [PDF]

				S. Gallinat, S. Busche, M. K. Raizada, and C. Sumners The angiotensin II type 2 receptor: an enigma with multiple variations Am J Physiol Endocrinol Metab, March 1, 2000; 278(3): E357 - E374. [Abstract] [Full Text] [PDF]

				Z.-Q. Wang, L. J. Millatt, N. T. Heiderstadt, H. M. Siragy, R. A. Johns, and R. M. Carey Differential Regulation of Renal Angiotensin Subtype AT1A and AT2 Receptor Protein in Rats With Angiotensin-Dependent Hypertension Hypertension, January 1, 1999; 33(1): 96 - 101. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Wang, D. H. Articles by Hu, Z. Search for Related Content PubMed PubMed Citation Articles by Wang, D. H. Articles by Hu, Z. Pubmed/NCBI databases Compound via MeSH Substance via MeSH