Atrial Natriuretic Peptide Clearance Research Articles

A novel promoter variant of the natriuretic peptide clearance receptor gene is associated with lower atrial natriuretic peptide and higher blood pressure in obese hypertensives.

The clearance receptor for natriuretic peptides (NPRC), a candidate gene for essential hypertension, is highly expressed in adipose tissue, where is nutritionally regulated. The objectives of the present study were to sequence the human 5'-flanking regulatory region of NPRC, to identify allelic variants and their frequencies, and to study the genotype/phenotype correlation in hypertensive patients. Using polymerase chain reaction (PCR) and direct automated sequencing, a biallelic (A/C) polymorphism was detected at position -55 in a conserved promoter element named P1. The novel C(-55) variant makes the promoter sequence identical to the mouse gene and introduces a second Hgal site in the amplified DNA, allowing the genotyping of a large number of subjects. In a random sample of 232 white Caucasians the C(-55) allele was more commonly found (81.7% of all alleles) with 155 CC (66.8%), 69 AC (29.7%) and only eight AA (3.5%) genotypes. Atrial natriuretic peptide (ANP) levels were determined in 84 patients with essential hypertension. In the presence of obesity (body mass index (BMI) > or = 30 kg/m2) the homozygous CC hypertensives (n = 21) had significantly lower plasma ANP (33.6 +/- 11.1 pg/ml) compared with the AC patients (n = 11; 46.8 +/- 15.9 pg/ml; P = 0.01), whereas systolic blood pressure (SBP) and mean blood pressure (MBP) had the opposite association (SBP 163.9 +/- 18.7 versus 150.9 +/- 12.9 and MBP 123.3 +/- 12 versus 114.5 +/- 5.9 mmHg; P< 0.05). The difference in ANP levels were also present when overweight patients (BMI > or = 27 kg/m2) were considered. A common 'ancestral' C(-55) variant of the NPRC P1 promoter is associated with lower ANP levels and higher SBP and MBP in obese hypertensives. The C(-55) variant, in the presence of increased adiposity, might reduce plasma ANP through increased NPRC-mediated ANP clearance, contributing to higher blood pressure.

β-Adrenoceptor antagonist propranolol potentiates hypotensive action of natriuretic peptides

β-Adrenoceptor antagonists are known to increase plasma atrial natriuretic peptide (ANP) levels despite their hypotensive action. The aim of the present study was to examine the role of the ANP system in the antihypertensive effects of a β-adrenoceptor antagonist. We investigated the effects of propranolol (75 mg kg −1 day −1, p.o., 4 weeks) on the ANP system in stroke-prone spontaneously hypertensive rats. Plasma ANP levels were significantly higher in the propranolol group than in the control group. Both receptor densities and mRNA levels of ANP C receptor were significantly decreased in the lung as the major site of ANP clearance from the circulation. In contrast, both central venous pressure and ANP mRNA levels in the heart were not significantly different between the two groups. Under both basal and ANP-stimulated conditions, the cGMP content in the aorta was significantly greater in the propranolol group than in the control group, whereas the basal and stimulated cGMP content of the kidney was similar in the two groups. Inhibition of endogenous ANP action by a specific ANP receptor antagonist, HS-142-1, produced a greater increase of blood pressure in the propranolol group than in the control group. These results suggest potentiation of natriuretic peptide activity as a new antihypertensive mechanism of the β-adrenoceptor antagonist propranolol.

Receptor-specific ligands distinguish natriuretic peptide receptors-A and -C in primate tissues.

Systemic clearance of atrial natriuretic peptide (ANP) is in part due to neutral endopeptidase (NEP) proteolysis and natriuretic peptide receptor-C (NPR-C) mediated endocytosis. Biological responses to ANP are primarily mediated by the membrane guanylyl cyclase-A/natriuretic peptide receptor-A (NPR-A). Analogs of ANP selective for NPR-A and/or resistant to NEP may have increased activity in those tissues where NPR-C and NEP are coexpressed with NPR-A. The analog of ANP termed vANP; [(R3D, G9T, R11S, M12L, G16R)ANP] is selective for human NPR-A with at least 10,000 fold reduction in affinity for human NPR-C. We report that rat NPR-A is insensitive to 10 nM vANP, demonstrating the limitations of this species in evaluating human therapeutic candidates. As an alternative approach we tested the binding and potency of receptor-selective and NEP-resistant ANP analogs in rhesus monkey tissues. Competition binding studies with a simplified version of vANP, sANP [(G9T, R11S, G16R)rANP], in rhesus monkey kidney and lung membrane preparations shows displacement of 125I-ANP from only a fraction of the total ANP receptor population, 30 and 85%, respectively. The remaining ANP binding sites can be occupied with the NPR-C selective ligand cANP(4-23). These data strongly suggest that only two classes of ANP receptor are present in these membrane preparations, NPR-A and NPR-C. The NEP resistant sANP derivative called sANP(TAPR) was 8 fold more potent (ED50 = 0.6 nM) than rANP (ED50 = 5 nM) in stimulating cGMP production in the lung membrane preparation. Our results demonstrate that the rhesus monkey natriuretic peptide receptors reflect the pharmacology of the human receptors, and that this species may be suitable to determine the role of NPR-C and NEP in peptide clearance and attenuating functional responses.

Effect of neutral endopeptidase inhibition on the natriuresis and renal clearance of atrial natriuretic peptide in perfused rat kidney.

Effect of neutral endopeptidase inhibition on the natriuresis and renal clearance of atrial natriuretic peptide in perfused rat kidney.

Effects of severe hemorrhage on plasma ANP and glomerular ANP receptors.

Atrial natriuretic peptide (ANP) plays an important role in blood volume and electrolyte homeostasis in normovolemia and in hypervolemic states. The currently available information on the effects of hypovolemia on plasma ANP is contradictory. Moreover, possible regulation of ANP receptors during severe hemorrhagic hypovolemia has not been investigated. This study evaluated the effects of severe hemorrhage on plasma ANP and on the regulation of glomerular ANP receptor subtypes in anesthetized rats. Constant rate bleeding of 50% of total blood volume within 2 h induced a reproducible shock state characterized by marked decreases in blood pressure, heart rate, and hematocrit and an increase in plasma renin activity and aldosterone. Hemorrhaged rats exhibited a gradual significant increase in plasma ANP from 39.3 +/- 2.9 to 114.7 +/- 20.0 pmol/l 1 h after the bleeding (P < 0.001 from the initial value and P < 0.02 from the final value of sham-shock rats). Hemorrhage induced a significant decrease in total glomerular ANP binding sites (172 +/- 25 vs. 363 +/- 39 fmol/mg protein in hemorrhaged and sham-shock rats, respectively, P < 0.05). This decrease was mainly due to a significant decrease in ANPC receptors (132 +/- 22 vs. 312 +/- 40 fmol/mg protein in hemorrhaged and sham-shock rats, respectively, P < 0.05). Hemorrhage did not change glomerular ANPA receptor density. No significant differences in the affinity of the glomerular receptor subtypes for ANP were detected. Our data indicate that plasma ANP increases after prolonged severe hemorrhage. It is suggested that downregulation of renal ANPC receptors leads to reduced clearance of ANP and contributes to elevation of its plasma level after severe hemorrhage.

Atrial Natriuretic Peptide (ANP)-Induced Inhibition of Glucagon Secretion: Mechanism of Action in Isolated Rat Pancreatic Islets

ANP increases insulin levels in vivo. Because in vitro an ANP-induced increase in cGMP levels of islets of Langerhans was observed but no simultaneous increase in insulin release, secreted glucagon may be a candidate for this second messenger affected by ANP. The inhibitory effect of glucose on glucagon secretion was pronounced by 1.0 nM ANP at 3.0 mM glucose as well as at 5.6 and 8.3 mM glucose. Because in other tissues cGMP (the specific second messenger of ANP) inhibits Ca2+ channels, the uptake of 45Ca2+ was investigated. ANP (1.0 nM) inhibited 45Ca2+ uptake, which was nearly completely abolished by a pertussis toxin (PT) pretreatment. The inhibition of 45Ca2+ uptake fits to inhibitory ANP effects on glucagon secretion but does not fit to insulin secretion. The glucagon secretion coupling cascade affected by ANP probably involves an increase in cGMP because 8-Br-cGMP (a membrane-permeable cGMP analogue) also decreased glucagon secretion. ANP (4–23), a truncated form of ANP, which is selective for the ANP clearance receptor, also inhibited glucagon secretion. HS-142-1, a guanylate cyclase receptor antagonist, tended to reverse the effect of ANP on glucagon release. The data indicate that in the presence of ANP, the in vivo homeostasis of glucose, though plasma insulin levels are increased, is not due to an ANP-mediated increase in glucagon secretion; ANP has a complex inhibitory effect on glucagon release. The data further indicate that the ANP-induced inhibition of glucagon secretion probably involves the cGMP system, an inhibition of Ca2+ uptake and the involvement of PT-sensitive G-proteins. Moreover, an involvement of the clearance receptor seems to be likely, ANP is a valuable tool for investigating glucagon secretion from pancreatic islets because paracrine effects of insulin can be excluded.

The metabolic clearance of atrial natriuretic peptide during human pregnancy

OBJECTIVE: Our purpose was to determine whether human pregnancy alters the metabolic clearance and natriuretic effect of atrial natriuretic peptide. STUDY DESIGN: The metabolic clearance rate of atrial natriuretic peptide (ANP 99-126) was measured serially in nine normotensive primigravid women studied in early and late pregnancy and again 4 months post partum (nonpregnant). Metabolic clearance of atrial natriuretic peptide was determined by use of a two-tier constant infusion technique (6 and 12 ng/kg/min, respectively). Sodium excretion was determined from 30-minute urine collections taken before and during infusion of atrial natriuretic peptide at both 6 and 12 ng/kg/min. RESULTS: Basal plasma atrial natriuretic peptide levels increased with gestation: in early pregnancy 18.0 ± 2.7 pg/ml, in late pregnancy 22.6 ± 4.2 pg/ml, and post partum 19.5 ± 3.6 pg/ml. Infusion of atrial natriuretic peptide at 6 and 12 ng/kg/min produced two distinct physiologic plasma levels of atrial natriuretic peptide. The metabolic clearance rates for nonpregnant women and those in early and late pregnancy at 6 and 12 ng/kg/min, respectively, were 3.4 ± 0.4 and 2.9 ± 0.4 L/min at plasma atrial natriuretic peptide levels of 86.2 ± 13.2 and 179.8 ± 42.5 pg/ml, respectively, 4.3 ± 0.5 and 4.3 ± 0.5 L/min at plasma atrial natriuretic peptide levels of 61.1 ± 4.9 and 131 ± 20.9 pg/ml ( p < 0.01, nonpregnant vs early pregnancy), and 3.8 ± 0.6 and 3.8 ± 0.5 L/min at plasma atrial natriuretic peptide levels of 72 ± 8.0 and 136 ± 18.3 pg/ml ( p < 0.05, nonpregnant vs late pregnancy), respectively. Infusion of atrial natriuretic peptide produced natriuresis in both pregnant and nonpregnant states; sodium excretion (basal to atrial natriuretic peptide infusion at 12 ng/kg/min) increased from 133 ± 19 to 207 ± 18 μmol/min, 129 ± 21 to 374 ± 35 μmol/min, and 128 ± 20 to 221 ± 33 μmol/min in nonpregnant women and those in early and late pregnancy, respectively. CONCLUSIONS: The metabolic clearance of atrial natriuretic peptide increased by 16 weeks' gestation and remained elevated thereafter. There appears to be no attenuation of the natriuretic effect of infused atrial natriuretic peptide in normotensive human pregnancy. (Am J Obstet Gynecol 1996;175:449-54.)

Clearance receptors and endopeptidase 24.11: equal role in natriuretic peptide metabolism in conscious sheep.

Although many studies have examined the effects of administering natriuretic peptide receptor C (NPR-C) ligands and endopeptidase 24.11 (EP 24.11) inhibitors on clearance and bioactivity of atrial natriuretic peptide (ANP), none have systematically compared their effects on the endogenous levels of both ANP and brain natriuretic peptide (BNP) under physiological conditions. Accordingly, we examined the hemodynamic, hormonal, and renal actions of an EP 24.11 inhibitor, SCH-32615, and an NPR-C ligand, C-ANP-(4-23), both alone and in combination in eight normal conscious sheep. NPR-C blockade and EP 24.11 inhibition induce similar rises in plasma ANP, BNP, and guanosine 3',5'-cyclic monophosphate (cGMP). Synergistic increments in plasma ANP, BNP, and cGMP observed during combined administration are likely to be due to the reduced clearance of C-ANP-(4-23) in the setting of EP 24.11 inhibition, leading to increased inhibition of the receptor pathway. Combined administration was also associated with enhanced hemodynamic actions and diuretic and natriuretic effects. Our findings show that both enzymatic and receptor clearance pathways contribute equally to natriuretic peptide clearance and induce potentially important hemodynamic and renal effects in normal conscious sheep.

Regulation of the renal response to atrial natriuretic peptide by sodium intake in preweaned rats.

The neonate conserves sodium avidly, and sodium intake is normally limited to that present in maternal milk. To evaluate the role of atrial natriuretic peptide (ANP) in this adaptation, preweaned rat pups were artificially reared and fed a formula with either normal sodium (25 mEq/L) or high sodium (145 mEq/L) for 7-8 d. To determine whether increased dietary sodium decreases ANP clearance receptor activity, animals were anesthetized, and the plasma ANP concentration (ANPp), urine flow (V), urinary sodium (UNa V), and cGMP excretion (UcGMP V) were measured before and after infusion of ANF(4-23), an ANP clearance receptor inhibitor (C-ANF), at 50 micrograms/kg/ min. Infusion of C-ANF increased ANPp 10-fold in both normal and high sodium groups, but V, UNa V, and UcGMP V increased only in animals receiving the high sodium diet (p < 0.05). Incubation of isolated glomeruli with 0.1 microM ANP increased extracellular cGMP more in high sodium than normal sodium groups (p < 0.05). We conclude that ANP clearance receptors in the neonate are highly activated regardless of sodium intake. Increased dietary sodium increases the renal diuretic and natriuretic response to circulating ANP through enhanced generation of cGMP.

Structure of the 5′‐Flanking Regulatory Region of the Mouse Gene Encoding the Clearance Receptor for Atrial Natriuretic Peptide

A full-length cDNA, encoding the mouse atrial natriuretic peptide clearance receptor (ANP-CR), was isolated from a mouse lung cDNA library. The deduced amino acid sequence of the mouse ANP-CR, showing a typical tripartite organization which lacks a guanylyl cyclase domain, was extremely well conserved compared with the ANP-CR homologs. To understand the molecular mechanisms underlying the regulation of mouse ANP-CR gene expression and to define the essential DNA sequences for the transcriptional activity, a genomic clone containing over 9 kb of the 5'-flanking region of the mouse ANP-CR gene has been isolated from a mouse genomic library. Sequence analysis revealed that the 2.3-kb region upstream from an ATG codon of the mouse ANP-CR gene contained a number of putative regulatory elements; TATA box, CAAT box, cAMP response element, AP-1 and two shear stress responsive elements. Additionally, an unusual feature was the presence of the tandem-repeated AP-2-like elements, which were closely overlapped with SP-1 element. Promoter analysis using deletion plasmids in mouse Balb/3T3 cells, highly producing ANP-CR mRNA, demonstrated that deletion of the sequence from -144 to +46 relative to the transcription start point caused a dramatic decrease of the transcriptional activity and that the TATA box at -269 was not essential for the basal transcriptional activity. Primer extension analysis indicated that transcription of the mouse ANP-CR gene starts from at least two major sites, suggesting that the sequence from -144 to +46, which was shown to involve a novel sequence composed of tandem-repeated TATA-box-like elements, contained promoter sequences. Furthermore, cis-acting negative elements were shown to be situated in three regions (from -1178 to -708, from -707 to -625 and from -248 to -145) of the mouse ANP-CR gene promoter.

Atrial natriuretic peptide (ANP)-induced inhibition of glucagon secretion: mechanism of action in isolated rat pancreatic islets.

ANP increases insulin levels in vivo. Because in vitro an ANP-induced increase in cGMP levels of islets of Langerhans was observed but no simultaneous increase in insulin release, secreted glucagon may be a candidate for this second messenger affected by ANP. The inhibitory effect of glucose on glucagon secretion was pronounced by 1.0 nM ANP at 3.0 mM glucose as well as at 5.6 and 8.3 mM glucose. Because in other tissues cGMP (the specific second messenger of ANP1 inhibits Ca2+ channels, the uptake of 45Ca2+ was investigated. ANP (1.0 nM) inhibited 45Ca2+ uptake, which was nearly completely abolished by a pertussis toxin (PT) pretreatment. The inhibition of 45Ca2+ uptake fits to inhibitory ANP effects on glucagon secretion but does not fit to insulin secretion. The glucagon secretion coupling cascade affected by ANP probably involves an increase in cGMP because 8-Br-cGMP (a membrane-permeable cGMP analogue) also decreased glucagon secretion. ANP(4-23), a truncated form of ANP, which is selective for the ANP clearance receptor, also inhibited glucagon secretion. HS-42-1, a guanylate cyclase receptor antagonist, tended to reverse the effect of ANP on glucagon release. The data indicate that in the presence of ANP, the in vivo homeostasis of glucose, though plasma insulin levels are increased, is not due to an ANP-mediated increase in glucagon secretion; ANP has a complex inhibitory effect on glucagon release. The data further indicate that the ANP-induced inhibition of glucagon secretion probably involves the cGMP system, an inhibition of Ca2+ uptake and the involvement of PT-sensitive G-proteins. Moreover, an involvement of the clearance receptor seems to be likely. ANP is a valuable tool for investigating glucagon secretion from pancreatic islets because paracrine effects of insulin can be excluded.

Fasting inhibits natriuretic peptides clearance receptor expression in rat adipose tissue.

The early phase of weight loss induced by fasting is associated with diuresis, natriuresis and reduction in blood pressure through unclear mechanisms. Atrial natriuretic peptide (ANP) is a cardiac hormone with potent natriuretic, diuretic and hypotensive effects mediated by 'biologically active' receptors (NPr-A). A second type of receptor mediates the clearance of ANP (the clearance receptor, NPr-C). Since NPr-C appears to be abundant in adipose tissue, we analysed NPr-C and NPr-A gene expression in white and brown adipose tissue (WAT and BAT) as well as in renal cortex of fasting rats. Plasma ANP, cyclic GMP and aldosterone were also measured. Twelve male Wistar rats were deprived of food for about 50 h, and 12 other rats were fed ad libitum. Periepididymal WAT, interscapular BAT and left renal cortex were used for RNA extraction and northern blot analysis with rat NPr-C and NPr-A complementary DNA probes labelled with 32P-dCTP. Densitometric analysis of hybridization signals was corrected by beta actin expression before statistical analysis. Blood was drawn for ANP, cyclic GMP (cGMP) and aldosterone radioimmunoassays, which were also measured in a group of six rats deprived of food for 25 h. A dramatic decrease in NPr-C steady-state messenger RNA levels was observed both in WAT (about 3.6-fold, P < 0.001) and in BAT (about threefold, P < 0.01), but fasting did not affect the expression of NPr-A in adipose tissues. In the renal cortex NPr-C and NPr-A messenger RNA levels were unaffected by fasting. ANP and aldosterone levels were reduced after fasting whereas cyclic GMP was increased at 25 h, but the differences did not reach statistical significance. Fasting exerts a tissue-specific and gene-specific suppression of NPr-C gene expression in adipose tissue that appears to be accompanied by an increased biological activity of ANP. The natriuresis and diuresis and reduction of blood pressure induced by fasting might result from a reduced expression of NPr-C in adipose fat pads.

Renal clearance of atrial natriuretic peptide during acute unilateral complete ureteral obstruction in the pig.

Renal extraction and renal plasma clearance of atrial natriuretic peptide from pigs with complete unilateral ureteral obstruction (UUO) and from intact anaesthetized pigs were determined from arteriovenous differences in plasma atrial natriuretic peptide and measured renal plasma flow. The effect of administration of either a cyclooxygenase inhibitor or an angiotensin converting enzyme inhibitor was examined during UUO. Renal extraction ratio and renal clearance rate of plasma atrial natriuretic peptide (ANP) in the intact pig was stable during the 15 h observation period. UUO resulted in a significant (P < 0.05) temporary increase in renal extraction ratio and a significant (P < 0.05) reduction in the renal clearance rate of atrial natriuretic peptide. During cyclooxygenase inhibition there was a significant increase in the renal extraction ratio of ANP. During angiotensin II converting enzyme inhibition, renal handling of atrial natriuretic peptide did not differ from that observed in control animals. The present data demonstrate that atrial natriuretic peptide is extracted by the obstructed kidney. Despite the significant reduction in renal blood flow during indomethacin administration, renal clearance of ANP was unaltered. The increase in ipsilateral renal extraction of atrial natriuretic peptide immediately after ureteral obstruction and indomethacin administration could be explained either by a direct influence of PGE2 on the renal haemodynamics altering renal extraction of ANP, or by a compensatory mechanism attempting to preserve renal function.

Pharmacodynamics of atrial natriuretic peptide in isolated perfused Dahl rat kidneys.

Atrial natriuretic peptide (ANP) has been implicated in the development of hypertension in Dahl R and S rats. To test the responses of DR and DS kidneys in the absence of the influence of neural and humoral mechanisms, we investigated the pharmacokinetics and pharmacodynamics of ANP in isolated perfused DR and DS kidneys, obtained from rats given a high or low sodium diet, after a bolus injection of ANP (1 microgram) or after a bolus injection plus infusion of ANP to maintain the perfusate concentration at 1000 pg/ml. The elimination rate constant was not different between the groups (DR, 0.044 min-1 vs. DS, 0.050 min-1). Clearance of ANP was 4 times greater than the glomerular filtration rate, indicating that a receptor-mediated peritubular clearance is probably the primary route of elimination. DS kidneys excreted 50% less sodium than DR kidneys. However, ANP caused a 5-fold increase in fractional sodium excretion in both DR and DS. ANP also increased sodium excretion, creatinine clearance, and urine flow. No alteration in ANP kinetics occurred to account for the reportedly increased circulating concentrations of ANP seen in DS rats. We conclude that isolated DR and DS kidneys respond differently to ANP after bolus ANP administration to concentrations of 10,000 pg/ml. This difference in response is due to the sodium excretory defect inherent in the DS kidney and not to an alteration in the DS kidney's ANP responsiveness.

The changes in natriuretic peptide receptors (NP-R) in the lung and kidney in DOCA-salt hypertensive rats

To elucidate the pathophysiological significance and the regulation of natriuretic peptide receptors (NP-R) in hypertension, we investigated the changes of NP-R in the lung, renal cortex and medulla using radioreceptor assay. We also examined the concentrations of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) in the atria and ventricles and plasma ANP concentration by specific radioimmunoassays. Elevated plasma ANP level, decreased atrial ANP concentration and increased ventricular ANP and BNP contents were observed in the DOCA-salt group when compared with the control group (p < 0.01). The ratio of BNP/ANP in the ventricle of the DOCA-salt rats was 50% of the control rats. The elevated plasma ANP secreted from the heart seems to reflect a defensive compensatory mechanism to counteract hypertension, and that ANP is the major natriuretic peptide secreted from the cardiac ventricle in DOCA-salt hypertensive rats. Scatchard plot analysis revealed that the maximal binding capacities (Bmax) of NP-R of the lung and renal cortex in DOCA-salt rats were significantly decreased from 71.0 +/- 10.4 to 38.4 +/- 5.9 (p < 0.05) and from 32.7 +/- 1.8 to 21.7 +/- 0.4 (fmol/mg. protein) (p < 0.01) compared with those in the control rats. The values of Bmax of the renal medulla between the two groups were not different. There was no significant change in the apparent dissociation constant (Kd) in the lung, renal cortex and medulla between the two groups. A competitive binding study using 125I- alpha-rANP1-28 and C-ANF4-23, a biologically silent clearance receptor (C-receptor) specific ligand, revealed that C-receptors are abundantly present in the renal cortex, while a relatively small quantity of C-receptor was detected in the renal medulla. In the lung, a substantial amount of C-receptor was detected. In the DOCA-salt treated rats, C-receptors were decreased in the lung and renal cortex compared with the control rats. These results indicate that the down-regulation of NP-R, especially C-receptor, was induced in the lung and renal cortex when plasma ANP levels were elevated in DOCA-salt hypertensive rats. In conclusion, our results suggest that down-regulation of C-receptor in the lung and kidney contributes to maintaining higher plasma ANP levels and maybe responsible for the counter-regulatory role of endogenous ANP in DOCA-salt rats. Our results show that the down-regulation of NP-R in the lung was larger than that in the kidney, suggesting that the lung may play a dominant role in the regulation of the clearance of ANP through C-receptors in vivo.

Interaction of a neutral endopeptidase inhibitor with an ANP-C receptor ligand in anesthetized dogs.

Inhibition of important degradative pathways of atrial natriuretic peptide (ANP) in vivo could be a valuable therapeutic tool for regulating endogenous levels of ANP. The aim was to investigate the in vivo effects of both blockade of atrial natriuretic peptide clearance receptor and inhibition of neutral endopeptidase 24.11, an enzyme shown to be involved in ANP breakdown. Therefore, we infused a specific neutral endopeptidase inhibitor ((S)-thiorphan) and an ANP-C receptor ligand (AP 811) alone or in combination into anaesthetized beagle dogs. Compared with vehicle controls, coadministration of (S)-thiorphan and AP 811 (100 micrograms/kg/min and 10 micrograms/kg/min, resp.) had greater effects on endocrine and renal parameters than administration of either substance alone. Coadministration of both compounds increased urinary excretion of volume and sodium, cGMP and ANP. We found also increased plasma cGMP, plasma ANP and decreased plasma renin activity. No effects were observed with respect to blood pressure, left ventricular pressure or heart rate during the infusion period of 2 h. We conclude from these investigations, that blocking both degrading pathways of ANP with the ANP-C receptor ligand AP 811 and the neutral endopeptidase inhibitor (S)-thiorphan is more effective than inhibition of either system alone. Such a combination might therefore be a useful therapeutic tool in cardiovascular diseases.

Downregulation of pulmonary atrial natriuretic peptide receptors in rats exposed to chronic hypoxia.

We hypothesized that a downregulation in pulmonary atrial natriuretic peptide (ANP) receptors helps raise plasma ANP levels during chronic hypoxia. We measured in vivo pulmonary uptake and plasma clearance of 125I-ANP and in vitro pulmonary binding kinetics of 125I-ANP in normoxic and chronically hypoxic rats. Exposure to 21 days of hypobaric (0.5 atm) hypoxia did not decrease specific binding of 125I-ANP in the kidney, but pulmonary binding decreased 35 and 75% after 1 and 3 days of hypoxia, respectively, and increased 200% after 3 days of normoxic recovery from 21 days of hypoxia. The total binding capacity for ANP to lung membrane fractions from normoxic rats, chronically hypoxic rats, and rats that had recovered from hypoxia was 488 +/- 59, 109 +/- 17, and 338 +/- 48 fmol/mg, respectively (P < 0.05 for hypoxic vs. normoxic or recovered lung membranes). The area under the 125I-ANP plasma concentration curve for normoxic and hypoxic rats and normoxic rats that were infused with the ANP C-receptor ligand C-ANF-(4-23) was 3,292 +/- 216, 5,022 +/- 466, and 8,205 +/- 1,059 disintegrations.min-1.ml-1, respectively [P < 0.05 for hypoxic vs. normoxic or C-ANF-(4-23)-infused rats]. We conclude that pulmonary ANP clearance is reduced during chronic hypoxia secondary to a downregulation in pulmonary ANP clearance receptors. Reduced pulmonary clearance of ANP may represent an adaptation that contributes to increased plasma ANP levels during chronic hypoxia.

No effect of the oral neutral endopeptidase inhibitor candoxatril, on bronchomotor tone and histamine reactivity in asthma

Neutral endopeptidase (NEP) is found in many tissues in man, including the lung. Metabolism by NEP is one of the main mechanisms for the clearance of atrial natriuretic peptide (ANP), a hormone that causes bronchodilation and reduces nonspecific bronchial reactivity in man. Candoxatril, an oral NEP inhibitor has been shown to elevate circulating ANP levels. We have sought to determine whether the administration of candoxatril will alter bronchomotor tone (forced expiratory volume in one second (FEV1)) and histamine reactivity. Ten male asthmatic patients with stable asthma were enrolled (mean (SD) age 32 (10) yrs; FEV1 92 (11)% predicted) in a randomized, double-blind, placebo-controlled study. On each study day, after baseline spirometry, patients received 200 mg of candoxatril or placebo. Spirometry was repeated at half hourly intervals. After 2 h a histamine inhalation test was performed. There was no significant difference in FEV1 values at baseline or at 2 h post-dosing between active and placebo study days, with mean (SEM) FEV1 at baseline and 2 h of 3.71 (0.29) l and 3.85 (0.29) l on the placebo day, and 3.89 (0.27) l and 4.05 (0.82) l on the active day, respectively. The geometric mean (range) provocative concentration of histamine producing a 20% fall in FEV1 (PC20) on the placebo day and active day did not differ significantly, being 1.17 (0.25-25.8) and 0.93 (0.13-32) mg.ml-1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Atrial natriuretic peptide metabolism during pregnancy in the rat

OBJECTIVE: Our purpose was to determine whether plasma clearance rates and production rates of atrial natriuretic peptide 99-126 are altered during pregnancy in the rat. STUDY DESIGN: Twelve virgin and 12 late-pregnant chronically instrumented, conscious, unrestrained Sprague-Dawley rats were studied. Mean arterial pressure, heart rate, and plasma atrial natriuretic peptide levels were measured before and during a 40-minute continuous infusion of atrial natriuretic peptide (10 ng/kg/min). RESULTS: Control mean arterial pressure was 106 ± 5 mm Hg in virgin rats versus 97 ± 4 mm Hg in pregnant rats. Atrial natriuretic peptide infusion did not significantly affect mean arterial pressure in either group of animals but decreased heart rate in virgin rats. Basal plasma atrial natriuretic peptide levels were significantly higher in virgin than in pregnant rats (107 ± 10 vs 78 ± 7 pg/ml, respectively, p < 0.05). Atrial natriuretic peptide infusion significantly increased plasma levels in both groups to similar (183 ± 19 and 154 ± 14 pg/ml, virgin vs pregnant rats). Calculated plasma clearance rates were similar in virgin and pregnant rats (166 ± 27 vs 155 ± 17 ml/kg/min). Estimated production rates of atrial natriuretic peptide were higher in virgin then in pregnant rats (15.1 ± 1.4 vs 11.4 ± 1.1 ng/kg/min, p < 0.05). CONCLUSIONS: Plasma atrial natriuretic peptide levels are lower in chronically instrumented near-term pregnant rats compared with levels in virgin rats. This is not related to differences in plasma atrial natriuretic peptide clearance rates but rather to a decrease in production rates in late pregnancy. (AM J OBSTET GYNECOL 1994;170:1184-8.)

214 Renal clearance of atrial natriuretic peptide is possibly enhanced in renal artery stenosis and may be of diagnostic value

Tunny, Terry; Klemm, Shelley; Finn, Wendy; Stowasser, Michael; Storie, W.; Gordon, Richard Author Information