Abstract The aim of this study was to examine the influence of the systemic renin-angiotensin system on the gene expression of atrial natriuretic peptide in rat hearts. The renin-angiotensin system was stimulated (1) by unilateral renal artery clipping (0.2-mm clip, 2 days), producing a fourfold increase of circulating plasma renin activity and increasing blood pressure; (2) by furosemide infusion with simultaneous salt substitu- tion, increasing plasma renin activity values to 45 ng angiotensin I/h per milliliter without changing blood pressure; or (3) by administration of the calcium antagonist amlodipine, which increased plasma renin activity values to 42 ng angiotensin I/h per milliliter and lowered blood pressure. Unilateral renal artery clipping increased atrial natriuretic peptide mRNA levels approximately 20-fold in the left ventricles and approximately twofold in the right ventricles and atria. Furosemide infusion had no effect on cardiac atrial natriuretic peptide mRNA levels, and in amlodipine-treated rats, cardiac atrial natriuretic peptide mRNA levels decreased to 30% of control values. The increase of atrial natriuretic peptide mRNA in the ventricles during renal artery clipping was blunted by the administration of the angiotensin-converting enzyme inhibitor ramipril, which also attenuated the blood pressure rise. In clipped rats amlodipine did not change elevated plasma renin activity values but abolished the rise of blood pressure and also attenuated the rise of atrial natriuretic peptide mRNA in the hearts. These findings indicate that an increase of the activity of the systemic renin-angiotensin system does not result in an obligatory change in cardiac atrial natriuretic peptide gene expression. Moreover, our results suggest that activation of the renin-angiotensin system by renal artery stenosis preferentially stimulates left ventricular atrial natriuretic peptide gene expression by an angiotensin II–dependent mechanism that could be associated with the induction of myocardial hypertrophy.
Cardiac ANP and the renal RAS are considered to be antagonistic physiological regulators of extracellular volume and BP.1 2 3 Apart from its natriuretic and diuretic effects, ANP also has an inhibitory effect on renin secretion from the kidneys,1 suggesting a direct control on its functional antagonist, the RAS. It is less clear whether the renal RAS in turn also controls the activity of the cardiac ANP system. There is ample information concerning changes of the RAS and ANP system during perturbations of sodium homeostasis or extracellular volume.1 These findings, however, do not distinguish between direct, renin-angiotensin–dependent, and renin-angiotensin–independent changes of the ANP system because salt load and extracellular volume may influence independently both the ANP system and the RAS. Only a few studies have suggested that the RAS may in fact have a direct effect on the ANP system by showing that infusion of subpressor doses of Ang II increases ANP secretion4 and that Ang II stimulates ANP gene expression in cultured cardiomyocytes.5 That Ang II may in fact influence ANP secretion and ANP gene expression directly is conceivable in view of the existence of Ang II receptors in the myocardium.6 7 8 The aim of the present study was therefore to investigate the influence of selective activation of the renal RAS on expression of the ANP gene in the heart.
We found that stimulation of the renal RAS by unilateral renal artery clipping stimulates cardiac ANP gene expression preferentially in the left ventricle. This stimulation requires Ang II and is best correlated with changes of BP rather than with circulating Ang II. Our findings show that induction of ANP mRNA in the 2K1C rat model is paralleled by induction of myocardial hypertrophy.
Animal experiments were conducted according to the National Institutes of Health Guide for the Care and Use of Laboratory Animals and German laws on the protection of animals. Male Sprague-Dawley rats weighing 180 to 220 g were used for the experiments. Rats had free access to normal food (Altromin C-1000, Lage) and tap water.
Amlodipine and Ramipril Treatments
Amlodipinebesilate (100 mg/kg, Norvasec, Pfizer GmbH) was administered by gavage each morning for 4 days. Ramipril (7.5 mg/kg) was administered by gavage each morning for 2 days.
Rats were kept in metabolic cages and fed standard chow. Two bottles of drinking solution were equally accessible, one containing tap water and the other a salt solution of 0.8% NaCl and 0.1% KCl. For the first 4 days of the experiment all rats were kept under the same conditions. Osmotic pumps (2 ML1, Alzet) were then implanted subcutaneously under the skin of the neck (day 0). In control rats the pumps were filled with 0.9% NaCl and in furosemide-treated rats with 2 mL Dimazon (Hoechst AG), equivalent to 50 mg/mL furosemide. The pumps delivered 10 μL/h, yielding a daily furosemide dose of 12 mg per rat. Furosemide treatment lasted 6 days. To guarantee a constant drug effect during the experiment, the pumps were primed in vitro for at least 4 hours before implantation.
For clipping, rats were anesthetized with methohexital (50 mg/kg), and the left kidney was exposed by an abdominal incision. Sterile silver clips (Degussa) with an inner diameter of 0.2 mm were placed on the left renal arteries. In sham-clipped rats the left artery was only touched with a forceps. The combination of renal artery clipping with amlodipine or ramipril treatment was performed by either starting both treatment regimens simultaneously (ramipril) or by pretreatment with the drugs for 2 days (amlodipine).
During the time of the experiment systolic BP was measured with a BP recorder (Rhema) twice daily, in the morning and afternoon.
After 48 hours (clipped, ramipril-treated, and sham-clipped rats), 96 hours (amlodipine-treated rats and corresponding control rats), or 144 hours (furosemide-treated rats) rats were killed by decapitation. Blood was collected from the carotid arteries for determination of PRA. Total hearts were rapidly removed and immediately frozen in liquid nitrogen until RNA extraction. For selected experiments, hearts were removed, and right and left atria as well as right and left ventricles were separated. The four different parts of the separated hearts were weighed and frozen in liquid nitrogen. In all other experiments, total hearts were used for mRNA analysis.
Total RNA was extracted from frozen hearts stored at −70°C according to the protocol of Chomczynski and Sacchi.9 After homogenization in 10 mL solution D (guanidine thiocyanate [4 mol/L] containing 0.5% N-lauryl-sarcosinate, 10 mmol/L EDTA, 25 mmol/L sodium citrate, and 700 mmol/L β-mercaptoethanol), 1 mL of 2 mol/L sodium acetate (pH 4), 10 mL phenol (water saturated), and 2 mL chloroform were added sequentially to the homogenate. After cooling on ice for 15 minutes samples were centrifuged at 10 000g for 15 minutes at 4°C. RNA in the supernatant was precipitated with an equal volume of isopropanol at −20°C for at least 1 hour. The resulting RNA pellets were resuspended in 0.5 mL solution D and again precipitated with an equal volume of isopropanol at −20°C. Pellets were finally dissolved in diethylpyrocarbonate-treated water and stored at −80°C until further processing.
RNase Protection Assay for ANP and Actin
ANP mRNA and cytoplasmic β-actin mRNA were measured by an RNase protection assay as described for erythropoietin.10 A partial cDNA clone of ANP was subcloned in the transcription vector pAM18 (Amersham International). The ANP clone was a generous gift from W.-G. Forssmann (Hannover, Germany). It contains a 145-bp-long, PCR-derived sequence (bp 322 to 466) of rat preproANP mRNA.11 After linearization with HindIII and in vitro transcription with SP6 RNA polymerase, the resulting 190-bp cRNA fragment was used for detection of ANP mRNA in the protection assay. Performing an RNase protection assay with this cRNA produces a 145-bp-long protected fragment of the ANP gene transcript.
For the β-actin protection assay the resulting PCR fragment described below was cloned in the vector pSP64 (Promega-Serva). Linearization with EcoRI and following in vitro transcription with SP6 RNA polymerase produces a 306-bp-long RNA transcript complementary to rat cytoplasmic β-actin. Hybridization conditions and autoradiography were as described previously.10 Transcripts were continuously labeled with [α-32P]GTP (400 Ci/mmol, Amersham) and purified on a Sephadex G50 spin column. For hybridization, heart total RNA was dissolved in a buffer containing 80% formamide, 40 mmol/L piperazine-N,N′-bis(2-ethanesulfonic acid), 400 mmol/L NaCl, and 1 mmol/L EDTA (pH 8). RNA (5 μg for ANP, 1 μg for β-actin) was hybridized in a total volume of 50 μL at 60°C overnight with 8.3×103 Bq radiolabeled probe. RNase digestion with RNase A and T1 was carried out at room temperature for 30 minutes and terminated with proteinase K digestion for 30 minutes at 37°C (0.1 mg/mL containing 0.4% sodium dodecyl sulfate).
Protected mRNA fragments were purified by phenol/chloroform extraction, ethanol precipitation, and subsequent electrophoresis on a 10% denaturing polyacrylamide gel. After autoradiography of the dried gel at −80°C overnight, bands representing protected mRNA fragments were excised from the gel, and radioactivity was counted with a liquid scintillation counter (1500 Tri-carb, Packard Instruments). The number of counts per minute obtained from each sample was expressed relative to an external ANP mRNA standard included in each hybridization procedure. The external standard consisted of 5 μg pooled RNA extracted from six hearts of normal Sprague-Dawley rats. Fig 1⇓ shows the results of an RNase protection assay for ANP mRNA with the use of equal amounts of total RNA of the six control rats used for generating a pool of RNA.
RT-PCR for α-Myosin, β-Myosin, and β-Actin
The abundance of rat cytoplasmic β-actin, α-myosin heavy chain, and β-myosin heavy chain in total mRNA from isolated hearts was determined by PCR. Primers for actin were act1 (5′-GGAATTCCCAACTGGGACGACATGG-3′), binding at bp 227 to 245, and act2 (5′-CGGGATCCTGGCGTGAGGGAGAGCAT-3′), binding at bp 514 to 532 of the β-actin mRNA.12 The resulting fragment was 306 bp long. To avoid coamplification of genomic DNA, the sequence amplified by the primers contained an intron.
Primer myo1 (5′-CGGGATCCACACCAACCTGTCCAAGT-3′), the binding site of which is located in the coding region near the stop codon, is identical for α-myosin (binding at bp 5697 to 5714)13 and β-myosin (binding at bp 5667 to 5684).14 The downstream primers myo2 for β-myosin (5′-GGAATTCACAGGCATCCTTAGGGTT-3′, binding at bp 5821 to 5843)14 and myo4 for α-myosin (5′-GGAATTCACAGGTTATTCCTCATCG-3′, binding at bp 5830 to 5847)13 are located immediately behind the stop codon and are specific for the corresponding isoform. The combination of myo1 and myo4 thus produces a 150-bp fragment of α-myosin, whereas myo1 combined with myo2 yields a 176-bp fragment of β-myosin. To facilitate cloning, all primers were constructed with EcoRI sites at their 3′ ends and with BamHI sites at their 5′ ends. For PCR reaction 1 μg total RNA of control or clipped rats was reverse-transcribed with 200 U of M-MLV reverse transcriptase (Gibco/BRL) with the use of standard protocols. To allow the quantification of actin and the two myosin isoforms from one cDNA sample, oligo(dT) (12-18) was used for priming the reverse transcriptase reaction (500 ng per reaction). From a total reaction volume of 20 μL, a 1:200 dilution was made, and 3 μL of 20 μL was used for PCR. Measurement of one cDNA for actin and the two isoforms of myosin PCR was as follows. To three cups with 3 μL of diluted reverse transcriptase reaction were added 1 μL of each primer (10 pmol/μL), 2 μL dNTP (2.5 mmol/L), 2 μL 10× concentrated buffer (supplied with enzyme), 1 U Taq polymerase (Boehringer), and 1 μCi [3H]dCTP (64 Ci/mmol). The reaction was performed in a final volume of 20 μL. PCR conditions were 26 cycles consisting of denaturation at 94°C (30 seconds), annealing at 55°C (1 minute), and extension at 72°C (30 seconds). PCR was completed by a final extension step of 10 minutes at 72°C.
After PCR the amplification products originating from actin or myosin mRNA were separated by polyacrylamide gel electrophoresis. N,N′-Methylene-bis-acrylamide was replaced by dihydroxyethylene-bis-acrylamide to allow dissolution of the bands. The bands were excised, solubilized in 0.025 mol/L periodic acid at 80°C for 2 hours, and counted in a β-counter. The radioactivity incorporated in the bands of α- and β-myosin was expressed as the relation of counts per minute β-myosin to counts per minute α-myosin measured in the same cDNA sample.
Determination of PRA, Immunoreactive ANP, and Plasma Aldosterone
PRA was determined with a commercially available radioimmunoassay kit for Ang I (Sorin Biomedica). Immunoreactive ANP was determined with a radioimmunoassay kit directed against rat α-ANF (Paesel and Lorei). Plasma aldosterone was measured with a radioimmunoassay (ALDOCTK-2, Sorin Biomedica).
ANOVA followed by Student’s unpaired t test was used for interindividual comparisons. A value of P<.05 was considered significant.
ANP gene expression in rat hearts was semiquantitated by RNase protection assay. To allow comparison, the hybridization signals obtained from hearts from the different experimental groups were related to an external standard obtained by pooling total RNA from the hearts of six untreated rats. Fig 1⇑ shows the hybridization signals for rat ANP mRNA of these hearts before and after pooling. The coefficient of variation for ANP mRNA for the normal hearts was 54%.
The endogenous RAS was stimulated with three different maneuvers: unilateral renal artery stenosis, treatment with the loop diuretic furosemide, and treatment with the calcium antagonist amlodipine. Clipping of left renal arteries with 0.2-mm clips for 2 days increased PRA from 8 to 32 ng Ang I/h per milliliter; systolic BP increased by 29 mm Hg; and ANP mRNA increased fivefold in the ventricles compared with control rats (Fig 2⇓). Both clipped and furosemide-treated rats showed a significant increase in PRA compared with control rats. Two days after clipping, ANP mRNA was obviously more abundant in atria and ventricles, the most prominent increase occurring in the left ventricle (Fig 3⇓). For semiquantitation of the absolute amount of ANP mRNA present in the different regions of the hearts, the hybridization signal obtained with 1 μg total RNA, expressed as a percentage of the external standard, was multiplied by the yield of total RNA from the different heart regions. This calculation revealed 1.6- to 1.8-fold increases of ANP mRNA in the atria and right ventricle, respectively, and a 19-fold increase in the left ventricles 2 days after renal artery clipping (Fig 4⇓).
Rats infused with furosemide to stimulate the RAS had free access to food, salt, and water to compensate for the enormous salt and water loss induced by the loop diuretic. This maneuver prevented volume contraction, as indicated by the unchanged body weights (control: 197±4.9 g; furosemide: 190±22 g), hematocrit (control: 38.5±0.4%; furosemide: 39.2±1.5%), and plasma sodium concentration (control: 143±3 mmol/L; furosemide: 148±5 mmol/L). In these rats PRA was increased to 45 ng Ang I/h per milliliter, but neither BP nor cardiac ANP mRNA levels were changed (Fig 2⇑).
A third means of stimulating the endogenous RAS was to treat rats with a calcium antagonist. Administration of the dihydropyridine derivative amlodipine (100 mg/kg per day) for 4 days increased PRA values to 42 ng Ang I/h per milliliter and decreased basal BP by 9 mm Hg. Amlodipine treatment lowered cardiac ANP mRNA levels significantly to 40% of control values. Because PRA was equally high in the 2K1C and amlodipine groups, this experiment shows a strong divergence between circulating Ang II and cardiac ANP mRNA levels (Fig 5⇓.
We performed further experiments to characterize the mechanism by which renal artery clipping led to the early enhancement of ANP gene expression. In particular, the roles of Ang II and BP were considered in this context. To investigate the possible involvement of Ang II in the enhancement of ANP gene expression after renal artery clipping, we examined the influence of the Ang I–converting enzyme inhibitor ramipril (Fig 6⇓). Effective inhibition of angiotensin-converting enzyme by ramipril was indicated by the strong compensatory increase of PRA from 8 to 70 ng Ang I/h per milliliter. In sham-clipped rats ramipril significantly decreased BP without changing cardiac ANP mRNA levels. In rats with renal artery clips ramipril attenuated the increase of BP and almost completely blocked the increase of ANP mRNA levels (Fig 6⇓).
To investigate the role of BP changes in the stimulation of ANP mRNA levels by renal artery clipping, we further examined the influence of the calcium antagonist amlodipine in rats with renal artery clips (Fig 5⇑). In these rats amlodipine prevented an increase of BP upon renal artery clipping, whereas PRA values were unchanged compared with sham-clipped, amlodipine-treated rats (Fig 5⇑). Clipping increased cardiac ANP mRNA levels approximately threefold in amlodipine-treated rats compared with rats from the same treatment group (Fig 5⇑). These data indicate that a combined influence of BP and Ang II is necessary for stimulation of ANP mRNA expression.
Since expression of the ANP gene in the ventricles is a marker for ventricular hypertrophy,15 16 we considered the possibility that induction of left ventricular hypertrophy by unilateral renal artery clipping leads to the enhancement of ANP gene expression. We therefore compared heart weights of sham-clipped and unilaterally clipped rats and found they did not differ (sham: 0.79±0.02 g; clipped: 0.78±0.05 g). An early marker for ventricular hypertrophy is changes in α- and β-myosin gene expression, such that β-myosin gene expression is upregulated and that for α-myosin is downregulated already in the early phase of ventricular hypertrophy.17 18 We therefore determined the ratio of β-myosin mRNA to α-myosin mRNA by RT-PCR in the hearts of clipped and sham-clipped rats (Fig 7⇓). As shown in Fig 8⇓, there was a fourfold increase of the ratio in rats with left renal artery clips. Ramipril treatment prevented the changes of α- and β-myosin gene expression. As shown in Fig 5⇑ for ANP mRNA expression, amlodipine treatment similarly attenuated but did not totally block the shift in the expression of the myosin isoforms in the clipped rats (Fig 8⇓).
For comparison we also analyzed the abundance of β-actin mRNA, which is considered to be a “housekeeping gene.” As shown in Figs 9⇓ and 10⇓ the abundance of β-actin mRNA in the hearts did not differ between treatment groups.
To establish whether changes in ANP mRNA correlate with circulating peptide we performed a radioimmunoassay directed against the biologically active, 28–amino acid α-ANF of rat. The Table⇓ shows that the mRNA values do indeed reflect plasma ANP level. Clipped rats showed a 3.5-fold increase, and clipped and amlodipine-treated rats showed a 2.1-fold increase of immunoreactive ANP. As shown in Fig 5⇑ the level of ANP mRNA was reduced in 2K1C and amlodipine plus 2K1C rats. In contrast, circulating ANP was increased in amlodipine plus 2K1C rats.
To elucidate the role of aldosterone in this study we determined plasma aldosterone concentrations with a radioimmunoassay. In rat atrial myocytes aldosterone modestly increases ANP mRNA.19 The Table⇑ indicates that plasma aldosterone correlated with the mRNA of ANP and circulating ANP except in the amlodipine plus 2K1C rats. Amlodipine plus 2K1C treatment attenuated ANP mRNA expression compared with 2K1C rats, but plasma aldosterone was increased approximately twofold in amlodipine plus 2K1C rats. Under furosemide treatment aldosterone levels were low, probably because of hypokalemia, despite an increased PRA.20 Plasma aldosterone and plasma ANP showed good agreement between ANP mRNA on the one hand and plasma ANP and aldosterone on the other. An exception was the amlodipine plus 2K1C group, in which ANP mRNA was reduced but circulating ANP and aldosterone levels were increased.
This study aimed to assess the influence of the systemic RAS on ANP gene expression in the heart. A classic maneuver to activate the RAS is unilateral renal artery stenosis. This is characterized by increased levels of circulating renin and Ang II caused by enhancement of renin secretion from the affected kidney.21 Our findings now show that unilateral renal artery clipping for 2 days markedly stimulates the expression of the ANP gene, in particular in the left ventricle (Fig 4⇑). However, stimulation of the RAS by the loop diuretic furosemide with maintenance of a normal sodium balance20 had no influence on cardiac ANP mRNA levels (Fig 2⇑), and stimulation of the RAS by the calcium antagonist amlodipine was even associated with a decrease of ANP mRNA levels (Fig 5⇑). Assuming that elevated PRA values reflect concomitant changes of plasma Ang II concentrations, these findings suggest that an increase of systemic renin and Ang II levels per se has no general effect on cardiac ANP gene expression. A decrease of ANP gene expression by calcium antagonists in vitro has previously been reported and explained by the hypothesis that calcium influx into cardiomyocytes positively influences ANP gene expression.22
The question thus arises by which pathway unilateral renal artery clipping causes an early and potent stimulation of ventricular ANP gene expression. Since the stimulation of ANP gene expression during renal artery clipping was blunted by the converting enzyme inhibitor ramipril (Fig 6⇑), it appears likely that Ang II plays an essential role in the stimulation of ANP gene expression. Prolonged unilateral renal artery stenosis is known to cause severe hypertension and cardiac hypertrophy.23 Therefore, it appears likely that the induction of hypertension is a major reason for the enhancement of ANP gene expression. Our findings suggest that the enhancement of ANP gene expression is associated with early induction of ventricular hypertrophy, because we found characteristic signs for ventricular hypertrophy, such as relative changes in the expression of the α- and β-myosin genes, at a time when heart weights were not different (Figs 7⇑ and 8⇑). The relative changes of α- and β-myosin gene expression seen in our experiments were smaller than those reported for manifest ventricular hypertrophy, indicating a very early stage of ventricular hypertrophy 2 days after unilateral renal artery clipping.
The transcellular pathways inducing ventricular hypertrophy are not well understood. There is both in vivo and in vitro evidence that Ang II plays an essential role in this process, leading to transcription of a special subset of genes in the myocardium.24 25 Our findings show that converting enzyme inhibition in fact prevents a change of myosin gene expression during renal artery stenosis and also prevents the enhancement of ANP gene expression (Fig 8⇑). Since an increase of systemic renin activity by furosemide or amlodipine did not stimulate ANP gene expression, it would appear that both Ang II and BP are necessary for the stimulation of ANP gene expression during renal artery stenosis. Moreover, our findings suggest the existence of a local heart RAS that appears to act independently of the systemic RAS. Plasma ANP determinations showed that changes in ANP mRNA are paralleled by changes in circulating ANP. An activated RAS results in high aldosterone levels. However, in our study plasma aldosterone was not strongly correlated with ANP expression. ANP levels were reduced in amlodipine plus 2K1C rats, which in fact showed elevated plasma aldosterone levels. In furosemide-treated rats aldosterone values were low because of hypokalemia.20 It has been found in in vitro experiments that cardiomyocytes are capable of elaborating Ang II in response to mechanical stress. Ang II in the myocyte has been shown to induce the hypertrophic changes.5 Our results now show that circulating Ang II is not directly involved in the development of left ventricular hypertrophy. The trigger for an enhanced intracardial formation of Ang II may include the increase of BP and additional factors that need to be identified.
Selected Abbreviations and Acronyms
|2K1C||=||two-kidney, one clip|
|Ang I, II||=||angiotensin I, II|
|ANP||=||atrial natriuretic peptide|
|PCR||=||polymerase chain reaction|
|PRA||=||plasma renin activity|
|RT-PCR||=||reverse transcriptase–polymerase chain reaction|
We thank Wolf-Georg Forssmann, Hannover, Germany, for providing us with rat ANP cDNA. The expert technical and graphic assistance provided by Wiebke de Haan, Karl Heinz Götz, Marlies Hamann, and Marie Luise Schweiger is gratefully acknowledged. For editorial help we thank Dr John Michael Davis, Munich, Germany.
- Received June 6, 1995.
- Revision received July 11, 1995.
- Accepted August 18, 1995.
Robertson JIS, Nicholls MG, eds. The Renin-Angiotensin System. London, UK: Gower Medical Publishing; 1993:36.1-36.17.
Volpe M, Atlas SA, Marion DE, Mueller FB, Sealey JE, Laragh JH. Angiotensin II-induced atrial natriuretic factor release in dogs is not related to hemodynamic responses. Circ Res. 1990;67:774-779.
Scott AL, Chang RSL, Lotti VJ, Siegl PKS. Cardiac angiotensin receptors: effects of selective angiotensin II receptor antagonists, DUP 753 and PD 121981 in rabbit heart. J Pharmacol Exp Ther. 1992;261:931-935.
Sechi LA, Griffin CA, Grady EF, Kalinyak YE, Schambelan M. Characterization of angiotensin II receptor subtypes in rat heart. Circ Res. 1992;71:1482-1489.
Suzuki J, Matsubara H, Urakami M, Inada M. Rat angiotensin II (type 1A) receptor mRNA regulation and subtype expression in myocardial growth and hypertrophy. Circ Res. 1993;73:439-447.
Ratcliffe PJ, Jones RW, Philips RE, Nicholls LG, Bell JI. Oxygen-dependent modulation of erythropoietin mRNA levels. J Exp Med. 1991;172:657-660.
Nudel U, Zakut R, Neumann S, Levy Z, Yaffe D. The nucleotide sequence of the rat cytoplasmatic beta-actin gene. Nucleic Acids Res. 1983;11:1759-1771.
McNally EM, Gianola KM, Leinwand LA. Complete nucleotide sequence of full length cDNA for rat α cardiac myosin heavy chain. Nucleic Acids Res. 1989;17:7527-7528.
Kraft R, Bravo-Zehnder M, Taylor DA, Leinwand LA. Complete nucleotide sequence of full length cDNA of rat β cardiac myosin heavy chain. Nucleic Acids Res. 1989;17:7529-7530.
Rockman HA, Ross RS, Harris AN, Knowlton KU, Steinhelper ME, Field LJ, Ross J Jr, Chien KR. Segregation of atrial-specific and inducible expression of an atrial natriuretic factor transgene in an in vivo murine model of cardiac hypertrophy. Proc Natl Acad Sci U S A. 1991;88:8277-8281.
Saito Y, Nakao K, Arai H, Sunagawa A, Morii N, Yamada T, Itoh H, Shioho S, Mukoyama M, Obata K, Yasue H, Ohkubo H, Nakanishi S, Imura H. Atrial natriuretic polypeptide (ANP) in human ventricle: increased gene expression of ANP in dilated cardiomyopathy. Biochem Biophys Res Commun. 1987;148:211-217.
Chassagne C, Wisnewsky C, Schwartz K. Antithetical accumulation of myosin heavy chain but not α-actin mRNA isoforms during early stages of pressure-overload−induced rat cardiac hypertrophy. Circ Res. 1993;72:857-864.
Chien KR, Knowlton KU, Zhu H, Chien AS. Regulation of cardiac gene expression during myocardial growth and hypertrophy: molecular studies of an adaptive physiologic response. FASEB J. 1991;5:3037-3046.
Stanton BA, Kaissling B. Adaptation of distal tubule and collecting duct to increased Na delivery, II: Na+ and K+ transport. Am J Physiol. 1988;255:F1269-F1275.
Hackenthal E, Paul M, Ganten D, Taugner R. Morphology, physiology, and molecular biology of renin secretion. Physiol Rev. 1990;70:1067-1116.
LaPointe MC, Deschepper CF, Wu J, Gardner DG. Extracellular calcium regulates expression of the gene for atrial natriuretic factor. Hypertension. 1990;15:20-28.
Baker KM, Chernin MI, Wixson SK, Aceto JF. Renin-angiotensin system involvement in pressure-overload cardiac hypertrophy in rats. Am J Physiol. 1990;259:H324-H332.
Everett AD, Tufro-McReddie A, Fisher A, Gomez RA. Angiotensin receptor regulates cardiac hypertrophy and transforming growth factor β1 expression. Hypertension. 1994;23:587-592.