ANP-stimulated Na+ secretion in the collecting duct prevents Na+ retention in the renal adaptation to acid load

We have recently reported that type A intercalated cells of the collecting duct secrete Na+ by a mechanism coupling the basolateral type 1 Na+-K+-2Cl− cotransporter with apical type 2 H+-K+-ATPase (HKA2) functioning under its Na+/K+ exchange mode. The first aim of the present study was to evaluate whether this secretory pathway is a target of atrial natriuretic peptide (ANP). Despite hyperaldosteronemia, metabolic acidosis is not associated with Na+ retention. The second aim of the present study was to evaluate whether ANP-induced stimulation of Na+ secretion by type A intercalated cells might account for mineralocorticoid escape during metabolic acidosis. In Xenopus oocytes expressing HKA2, cGMP, the second messenger of ANP, increased the membrane expression, activity, and Na+-transporting rate of HKA2. Feeding mice with a NH4Cl-enriched diet increased urinary excretion of aldosterone and induced a transient Na+ retention that reversed within 3 days. At that time, expression of ANP mRNA in the collecting duct and urinary excretion of cGMP were increased. Reversion of Na+ retention was prevented by treatment with an inhibitor of ANP receptors and was absent in HKA2-null mice. In conclusion, paracrine stimulation of HKA2 by ANP is responsible for the escape of the Na+-retaining effect of aldosterone during metabolic acidosis.

Atrial Natriuretic Peptide Receptor Upregulation in the Rat Inner Ear

The purpose of this study was to further examine whether fluid homeostasis in the endolymphatic system could be regulated by a locally effective paracrine system involving atrial natriuretic peptides (ANPs) and their receptors. We assessed the biologic activity of the 3 ANP receptors (ANP-A, ANP-B, ANP-C) in the rat inner ear by measuring receptor upregulation after inner ear administration of ANPs. After appropriate anesthesia, female Lewis rats were injected with ANP via the round window. The animals were sacrificed 24 hours later, and RNA was isolated for reverse transcription–polymerase chain reaction (RT-PCR). Electrophoresis of RT-PCR products showed the presence of all 3 ANP receptor genes in both injected and control animals. Gene expression was significantly higher 24 hours after injection. These findings demonstrate that ANP receptors in the inner ear can be upregulated after injection of ANPs.

Effects of water deprivation on atrial natriuretic peptide secretion and density of binding sites in adrenal glands and kidneys of maternal and fetal rats in late gestation

The effects of water deprivation for 3 days were studied in pregnant rats and their fetuses on day 21 of gestation. Maternal water deprivation induced a significant decrease of the body weight in both maternal and fetal rats. This weight loss was accompanied by significant increases in plasma osmolality and haematocrit in both maternal and fetal rats. Similarly, dehydration significantly decreased plasma atrial natriuretic peptide (ANP) concentrations and increased plasma aldosterone concentrations in maternal and fetal rats. Water-deprived maternal rats presented a significant increase in total ANP receptor density in isolated renal glomeruli and adrenal zona glomerulosa membranes. This increase was due to a significant increase in ANPc receptor density in both renal glomeruli and adrenal zona glomerulosa. The densities of total ANP, ANPb and ANPc receptors in fetal kidneys and adrenal glands were not affected by maternal dehydration. These results suggest that the dehydrated maternal rat is able to up-regulate the number of its ANP receptors in its kidneys and adrenal glands, in response to a decrease in plasma ANP concentrations. In contrast, the fetal rat does not seem to be able to regulate its own ANP receptors in response to maternal dehydration, in spite of a decrease in plasma ANP concentrations.

Evidence for functional ANP receptors in cultured alveolar type II cells

Because atrial natriuretic peptide (ANP) is considered to play a role in lung physiology and pathology, our aim was to characterize natriuretic peptide receptors in cultured rat alveolar type II (ATII) cells. Guanylate cyclase A- and B-receptor but not clearance-receptor mRNAs were detected by reverse transcription-polymerase chain reaction. The absence of clearance-receptor expression in ATII cells was confirmed by competitive inhibition of ANP binding; ANP (0.1–100 nM) decreased the binding of 125I-ANP, whereas C-ANP-(4—23), a specific ligand of clearance receptors, was ineffective. ANP induced a dose-dependent increase in guanosine 3′,5′-cyclic monophosphate (cGMP) production, with a threshold of 0.1 nM, whereas the response to C-type natriuretic peptide was weak and was observed only at high concentrations (100 nM). In ATII cells cultured on filters, 1) ANP receptors were present on both the apical and basolateral surfaces and 2) cGMP egression was polarized, as indicated by the greater ANP-induced cGMP accumulation in the basolateral medium, and was partially inhibited by probenecid, an organic acid transport inhibitor. Influx studies demonstrated that ANP decreased the amiloride-sensitive component of22Na influx but did not change ouabain-sensitive 86Rb influx. In conclusion, ATII cells behave as a target for ANP. ANP activation of guanylate cyclase A receptors produces cGMP, which is preferentially extruded on the basolateral side of the cells and inhibits the amiloride-sensitive Na-channel activity.

The Natriuretic Peptide (Onp-39) From Platypus (Ornithorhynchus anatinus) Venom Promotes Mast Cell Histamine Release.

In this study we investigated the possibility that the C-type natriuretic peptide from platypus venom (ONP-39) contributes to the acute effects of envenomation, which include oedema, pain and erythema. These effects may result from the release of auto pharmacological mediators from mast cells. Using an in vitro assay we have demonstrated that both ONP-39 and the endogenous C-type natriuretic peptide (CNP-22) release histamine from rat peritoneal mast cells, an effect similar to the structurally homologous atrial natriuretic peptide (ANP) (Opgenorth et al., 1990, Peptides 11(5):1003-7). Two synthetic peptides, ONP-39(1-17) and ONP-39(18-39), corresponding to the N- and C- termini respectively, are equipotent, suggesting that ONP-39 and other natriuretic peptides do not act through conventional ANP receptors on mast cells. The ability of ONP-39 to promote histamine release suggests that it may contribute to the acute symptoms of envenomation.

Downregulation of adrenal atrial natriuretic peptide receptor mRNAs and proteins by pregnancy in rats

The main objective of this study was to find out if the reported changes in the aldosterone-suppressant activity of atrial natriuretic peptide (ANP) during different hormonal states in rats are due to a modulation of ANP receptors. In zona glomerulosa cells, ribonuclease protection assay detected mRNAs for guanylate cyclase (GC)-coupled ANP GC-A and GC-B receptors, and for ANP C receptors, which are not coupled to GC. Western analysis using polyclonal anti-GC-A and anti-GC-B receptor antibodies revealed the presence of GC-A but not GC-B receptor proteins in zona glomerulosa cells. Pregnancy (days 7, 16 and 21), oestradiol-17 beta and progesterone decreased mRNAs for all the three ANP receptors in zona glomerulosa cells. Pregnancy decreased GC-A receptor proteins in zona glomerulosa cells, but these recovered to virgin values on day 2 postpartum. ANP receptor mRNAs in zona glomerulosa cells increased by postpartum day 2, but did not reach the values found in virgin rats. Zona fasciculata mainly contained GC-A receptor mRNA. It is concluded that ANP receptors in rat adrenal zona glomerulosa are modulated by pregnancy, oestrogen and progesterone; a decrease in ANP GC-A receptors during pregnancy might explain the accompanying decrease in the aldosterone-suppressant effects of ANP.

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 ANPAreceptor 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.

Comparison of ANP binding and sensitivity in brains from hypertensive and normotensive rats

We compared the abundance and sensitivity of atrial natriuretic peptides (ANP) receptors in the brains of spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats and examined the effect of blood pressure on the abundance of brain ANP receptors in several other experimental rat models. Brain slices from SHR generated more guanosine 3',5'-cyclic monophosphate in response to ANP than brain slices from WKY rats. No differences were found in brain particulate guanylate cyclase activity in both strains of rats. In rat brain homogenates, we observed that ANP bound in a specific and saturable fashion to samples from WKY rats, but not in samples from SHR. In vitro receptor autoradiography revealed that ANP binding was reduced in the subfornical organ, the choroid plexus, and the paraventricular nucleus of SHR compared with WKY rat brains. Correction of hypertension in SHR or induction of hypertension in other strains did not affect ANP binding in any of these brain regions. Altogether, our data suggest that the increased sensitivity of SHR brains to the action of ANP may be a consequence of factors other than the abundance of receptors and that it is not secondary to the elevation of blood pressure.