Atrial natriuretic factor negatively modulates secretin intracellular signaling in the exocrine pancreas

We previously reported that atrial natriuretic factor (ANF) stimulates pancreatic secretion through NPR-C receptors coupled to PLC and potentiates secretin response without affecting cAMP levels. In the present study we sought to establish the intracellular signaling mechanism underlying the interaction between both peptides. In isolated pancreatic acini 100 nM ANF abolished cAMP accumulation evoked by any dose of secretin. Lower doses of ANF (1 fM, 1 pM, 1 and 10 nM) dose dependently reduced EC50 secretin-evoked cAMP. Although ANF failed to affect cAMP stimulated by amthamine (selective H2 agonist) or isoproterenol (beta-adrenergic agonist), it abolished VIP-induced cAMP formation. ANF inhibitory effect was prevented by U-73122 (PLC inhibitor) and GF-109203X (PKC inhibitor) but unaltered by PKG and nitric oxide synthase inhibition, supporting that the PLC/PKC pathway mediated the effect. ANF response was mimicked by cANP (4-23 amide) and abolished by pertussis toxin, strongly supporting NPR-C receptor activation. In vivo studies showed that ANF at 0.5 microg x kg(-1) x h(-1) enhanced secretion stimulated by 1 U x kg(-1) x h(-1) secretin but at 1 and 2 microg x kg(-1) x h(-1) it abolished secretin response. However, ANF at such doses failed to modify the secretion evoked by carbachol or CCK. Present results show that ANF negatively modulated secretin secretory response and intracellular signaling through the activation of NPR-C receptors coupled to the PLC/PKC pathway. Furthermore, the finding that ANF also inhibited VIP-evoked cAMP supports a selective modulation of class II G-protein coupled receptors by ANF. Present findings suggest that ANF may play a protective role by reducing secretin response to avoid overstimulation.     

Atrial natriuretic factor intracellular signaling in the rat submandibular gland

We previously reported that intravenously administered atrial natriuretic factor (ANF) induced no salivation but enhanced agonist-evoked secretion in submandibular glands. The gene expression of ANF and natriuretic peptide receptors (NPR) was later reported in the glands. In the present study we sought to establish the intracellular signalling mechanisms underlying ANF modulation of salivary secretion. Fasted rats were prepared with submandibular duct and femoral cannulation. Dose–response curves to methacholine (MC) and norepinephrine (NE) were performed in the presence of cANP (4–23 amide) (selective NPR-C agonist) and ANF. Local injection of the agonist or ANF-induced no salivation, but enhanced MC and NE-evoked secretion. ANF and cANP (4–23 amide) enhanced phosphoinositide turnover being the effect abolished by U73122 (PLC inhibitor). Further ANF and cANP (4–23 amide) decreased basal cAMP content but failed to affect isoproterenol or forskolin-evoked cAMP. ANF response was inhibited by pertussis toxin and mimicked by cANP (4–23 amide) strongly supporting NPR-C activation. ANF-induced cAMP reduction was abolished by PLC and PKC inhibitors. The content of cGMP was dose dependently stimulated by ANF but not modified by cANP (4–23 amide). These findings support that ANF through NPR-C receptors coupled to PLC activation and adenylyl cyclase inhibition interacts with sialogogic agonists in the submandibular gland to potentiate salivation.     

Roles of 5-HT receptors in the release and action of secretin on pancreatic secretion in rats

5-Hydroxytryptamine (serotonin, 5-HT) is a hormone and neurotransmitter regulating gastrointestinal functions. 5-HT receptors are widely distributed in gastrointestinal mucosa and the enteric nervous system. Duodenal acidification stimulates not only the release of both 5-HT and secretin but also pancreatic exocrine secretion. We investigated the effect of 5-HT receptor antagonists on the release of secretin and pancreatic secretion of water and bicarbonate induced by duodenal acidification in anesthetized rats. Both the 5-HT(2) receptor antagonist ketanserin and the 5-HT(3) receptor antagonist ondansetron at 1-100 microg/kg dose-dependently inhibited acid-induced increases in plasma secretin concentration and pancreatic exocrine secretion. Neither the 5-HT(1) receptor antagonists pindolol and 5-HTP-DP nor the 5-HT(4) receptor antagonist SDZ-205,557 affected acid-evoked release of secretin or pancreatic secretion. None of the 5-HT receptor antagonists affected basal pancreatic secretion or plasma secretin concentration. Ketanserin or ondansetron at 10 microg/kg or a combination of both suppressed the pancreatic secretion in response to intravenous secretin at 2.5 and 5 pmol x kg(-1) x h(-1) by 55-75%, but not at 10 pmol x kg(-1) x h(-1). Atropine (50 microg/kg) significantly attenuated the inhibitory effect of ketanserin on pancreatic secretion but not on the release of secretin. These observations suggest that 5-HT(2) and 5-HT(3) receptors mediate duodenal acidification-induced release of secretin and pancreatic secretion of fluid and bicarbonate. Also, regulation of pancreatic exocrine secretion through 5-HT(2) receptors may involve a cholinergic pathway in the rat.     

C-type natriuretic peptide enhances amylase release through NPR-C receptors in the exocrine pancreas

Several studies show that C-type natriuretic peptide (CNP) has a modulatory role in the digestive system. CNP administration reduces both jejunal fluid and bile secretion in the rat. In the present study we evaluated the effect of CNP on amylase release in isolated pancreatic acini as well as the receptors and intracellular pathways involved. Results showed that all natriuretic peptide receptors were expressed not only in the whole pancreas but also in isolated pancreatic acini. CNP stimulated amylase secretion with a concentration-dependent biphasic response; maximum release was observed at 1 pM CNP, whereas higher concentrations gradually attenuated it. The response was mimicked by a selective natriuretic peptide receptor (NPR-C) agonist and inhibited by pertussis toxin, strongly supporting NPR-C receptor activation. CNP-evoked amylase release was abolished by U-73122 (PLC inhibitor) and 2-aminoethoxydiphenyl borate (2-APB) [an inositol 1,4,5-triphosphate (IP(3)) receptor antagonist], partially inhibited by GF-109203X (PKC inhibitor), and unaltered by ryanodine or protein kinase A (PKA) and protein kinase G (PKG) inhibitors. Phosphoinositide hydrolysis was enhanced by CNP at all concentrations and abolished by U-73122. At 1 and 10 pM, CNP did not affect cAMP or guanosine 3',5'-cyclic monophosphate (cGMP) levels, but at higher concentrations it increased cGMP and diminished cAMP content. Present findings show that CNP stimulated amylase release through the activation of NPR-C receptors coupled to the PLC pathway and downstream effectors involved in exocytosis. The attenuation of amylase release was likely related to cAMP reduction. The augmentation in cGMP supports activation of NPR-A/NPR-B receptors probably involved in calcium influx. Present findings give evidence that CNP is a potential direct regulator of pancreatic function.     

G(i-1)/G(i-2)-dependent signaling by single-transmembrane natriuretic peptide clearance receptor

Single-transmembrane natriuretic peptide clearance receptor (NPR-C), which is devoid of a cytoplasmic guanylyl cyclase domain, interacts with pertussis toxin (PTx)-sensitive G proteins to activate endothelial nitric oxide synthase (eNOS) expressed in gastrointestinal smooth muscle cells. We examined the ability of NPR-C to activate other effector enzymes in eNOS-deficient tenia coli smooth muscle cells; these cells expressed NPR-C and NPR-B but not NPR-A. Atrial natriuretic peptide (ANP), the selective NPR-C ligand cANP-(4-23), and vasoactive intestinal peptide (VIP) inhibited (125)I-ANP and (125)I-VIP binding to muscle membranes in a pattern indicating high-affinity binding to NPR-C. Interaction of VIP with NPR-C was confirmed by its ability to inhibit (125)I-ANP binding to membranes of NPR-C-transfected COS-1 cells. In tenia muscle cells, all ligands selectively activated G(i-1) and G(i-2); VIP also activated G(s) via VIP(2) receptors. All ligands stimulated phosphoinositide hydrolysis, which was inhibited by ANP-(1-11), PTx, and antibodies to phospholipase C-beta3 (PLC-beta3) and Gbeta. cANP-(4-23) contracted tenia muscle cells; contraction was blocked by U-73122 and PTx and by antibodies to PLC-beta3 and Gbeta in intact and permeabilized muscle cells, respectively. VIP and ANP contracted muscle cells only after inhibition of cAMP- and cGMP-dependent protein kinases. ANP and cANP-(4-23) inhibited forskolin-stimulated cAMP in a PTx-sensitive fashion. We conclude that NPR-C is coupled to activation of PLC-beta3 via betagamma-subunits of G(i-1) and G(i-2) and to inhibition of adenylyl cyclase via alpha-subunits.     

Natriuretic peptides increase cAMP production in human thyrocytes via the natriuretic peptide clearance receptor (NPR-C)

The relationship between natriuretic peptides and adenylyl cyclase/cAMP signal transduction has generally been shown to be an inhibitory one, mediated via the NPR-C receptor coupled to adenylyl cyclase by inhibitory G proteins (Gi). In the present studies, we have investigated the modulation of cAMP by natriuretic peptides in a long-term culture of human thyroid cells. Competition of [125I] rat ANF binding to human thyrocytes (HTU-5) by rat ANF (99-126) and by the NPR-C-specific analog C-ANF (4-23) indicated that greater than 97% of the ANF binding sites on HTU-5 cells are of the NPR-C type. However, rather than inhibiting intracellular cAMP in these cells, ANF increased maximal cAMP to 200-300% of control value. The ANF-induced increase in cAMP was duplicated by C-ANF (4-23). Basal cAMP content was reduced, and the response to ANF was abolished when the cells were grown in low (0.5%) serum without the addition of pituitary and hypothalamic extracts. CNP-22 also increased cAMP above control in HTU-5 cells identically to ANF. Neither ANF nor C-ANF (4-23) had any effect on cAMP in a culture of rat aortic smooth muscle cells. These results provide the first evidence for a positive effect of natriuretic peptides on cAMP mediated through the NPR-C, suggesting the possibility of an alternative mode of signaling by this receptor subtype.     

Secretin is not necessary for exocrine pancreatic development and growth in mice

Adaptive exocrine pancreatic growth is mediated primarily by dietary protein and the gastrointestinal hormone cholecystokinin (CCK). Feeding trypsin inhibitors such as camostat (FOY-305) is known to induce CCK release and stimulate pancreatic growth. However, camostat has also been reported to stimulate secretin release and, because secretin often potentiates the action of CCK, it could participate in the growth response. Our aim was to test the role of secretin in pancreatic development and adaptive growth through the use of C57BL/6 mice with genetic deletion of secretin or secretin receptor. The lack of secretin in the intestine or the secretin receptor in the pancreas was confirmed by RT-PCR. Other related components, such as vasoactive intestinal polypeptide (VIP) receptors (VPAC(1) and VPAC(2)), were not affected. Secretin increased cAMP levels in acini from wild-type (WT) mice but had no effect on acini from secretin receptor-deleted mice, whereas VIP and forskolin still induced a normal response. Secretin in vivo failed to induce fluid secretion in receptor-deficient mice. The pancreas of secretin or secretin receptor-deficient mice was of normal size and histology, indicating that secretin is not necessary for normal pancreatic differentiation or maintenance. When WT mice were fed 0.1% camostat in powdered chow, the pancreas doubled in size in 1 wk, accompanied by parallel increases in protein and DNA. Camostat-fed littermate secretin and secretin receptor-deficient mice had similar pancreatic mass to WT mice. These results indicate that secretin is not required for normal pancreatic development or adaptive growth mediated by CCK.     

C-type natriuretic peptide receptor expression in pancreatic alpha cells

Atrial natriuretic peptide (ANP), brain type natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) comprise a family of natriuretic peptides that mediate their biological effects through three natriuretic peptide receptor subtypes, NPR-A (ANP, BNP), NPR-B (CNP) and NPR-C (ANP, BNP, CNP). Several reports have provided evidence for the expression of ANP and specific binding sites for ANP in the pancreas. The purpose of this study was to identify the ANP receptor subtype and to localize its expression to a specific cell type in the human pancreas. NPR-C immunoreactivity, but neither ANP nor NPR-A, was detected in human islets by immunofluorescent staining. No immunostaining was observed in the exocrine pancreas or ductal structures. Double-staining revealed that NPR-C was expressed mainly in the glucagon-containing alpha cells. NPR-C mRNA and protein were detected in isolated human islets by RT-PCR and Western blot analysis, respectively. NPR-C expression was also detected by immunofluorescent staining in glucagonoma but not in insulinoma. ANP, as well as BNP and CNP, stimulated glucagon secretion from perifused human islets (1,111 ± 55% vs. basal [7.3 fmol/min]; P < 0.001). This response was mimicked by cANP(4–23), a selective agonist of NPR-C. In conclusion, the NPR-C receptor is expressed in normal and neoplastic human alpha cells. These findings suggest a role for natriuretic peptides in the regulation of glucagon secretion from human alpha cells.     

Factors regulating amylase secretion from chicken pancreatic acini in vitro

In mammals, cholecystokinin regulates pancreatic exocrine secretion under physiological conditions. We have shown, however, that cholecystokinin at physiological concentrations does not induce pancreatic amylase secretion in birds. Therefore, we investigated the effects of various neurotransmitters and gut hormones on the pancreatic amylase secretory response in isolated chicken pancreatic acini. Acetylcholine (half-maximal stimulation at 800 nM) and vasoactive intestinal polypeptide (half-maximal stimulation at 40 pM) produced a concentration-dependent increase in amylase secretion at physiological concentrations. The combination of acetylcholine and vasoactive intestinal polypeptide produced an additive response in amylase secretion. Sodium nitroprusside, a spontaneous nitric oxide releaser, and bombesin, induced amylase secretion at concentrations greater than 10 nM and 100 nM, respectively. Gastrin and secretin increased amylase secretion at pharmacological concentrations (10 to 100 nM). Our findings suggest that neural regulation is important for pancreatic enzyme secretion in birds and the contribution of gut hormones seems to be physiologically unimportant.     

MEK inhibits secretin release and pancreatic secretion: roles of secretin-releasing peptide and somatostatin

We investigated the mechanism of action of methionine enkephalin (MEK) on HCl-stimulated secretin release and pancreatic exocrine secretion. Anesthetized rats with pancreatobiliary cannulas and isolated upper small intestinal loops were perfused intraduodenally with 0.01 N HCl while bile and pancreatic juice were diverted. The effect of intravenous MEK on acid-stimulated secretin release and pancreatic exocrine secretion was then studied with or without coinfusion of naloxone, an anti-somatostatin (SS) serum, or normal rabbit serum. Duodenal acid perfusate, which contains secretin-releasing peptide (SRP) activity, was collected from donor rats with or without pretreatment with MEK, MEK + naloxone, or MEK + anti-SS serum, concentrated by ultrafiltration, and neutralized. The concentrated acid perfusate (CAP), which contains SRP bioactivity, was infused intraduodenally into recipient rats. MEK increased plasma SS concentration and inhibited secretin release and pancreatic fluid and bicarbonate secretion dose-dependently. The inhibition was partially reversed by naloxone and anti-SS serum but not by normal rabbit serum. In recipient rats, CAP increased plasma secretin level and pancreatic secretion. CAP SRP bioactivity decreased when it was collected from MEK-treated donor rats; this was partially reversed by coinfusion with naloxone or anti-SS serum. These results suggest that in the rat, MEK inhibition of acid-stimulated pancreatic secretion and secretin release involves suppression of SRP activity release. Thus the MEK inhibitory effect appears to be mediated in part by endogenous SS.     

Hepatic bile and pancreatic exocrine secretions evoked by gastrointestinal peptides in sheep

The secretory response of hepatic bile and exocrine pancreas to gastrointestinal peptides has been studied in chronically cannulated sheep. Pancreatic juice flow and protein output were evoked dose dependently by intraportal injection of secretin, CCK-8, caerulein, VIP and neurotensin. However, biliary secretion was evoked by only secretin. Biliary and pancreatic exocrine secretions were enhanced by delivered gastric juice into the duodenum as followed by the increased plasma concentration of immunoreactive secretin (IRS). Results suggest that secretin is the major peptide that regulates pancreatic exocrine secretion and hepatic bile production in the sheep.     

Effect of nitric oxide synthase inhibitors on the temporal coordination of duodenal contractions and pancreatic exocrine secretion in sheep

The present study evaluated the role of nitric oxide in the regulation of duodenal motility and pancreatic exocrine secretion in conscious sheep. Intravenous infusions of nitric oxide synthase inhibitors, Nω-nitro-l-arginine-methyl ester (l-NAME) and Nω-nitro-l-arginine, induced clusters of duodenal contractions like phase III of migrating motor complexes and simultaneously inhibited flow rate, bicarbonate ion and enzyme outputs of pancreatic juice. The effects of l-NAME were inhibited by simultaneous infusion of l-arginine, but not altered by adrenergic blockade using a combined infusion of phentolamine and propranolol. Inhibition of the pancreatic secretion occurred in coincidence with initiation of the duodenal contractions, while the pancreatic secretion was not inhibited when the premature duodenal contractions were abolished by the l-arginine infusion. The initiation of the cluster of duodenal contractions by l-NAME was not abolished by background infusion of atropine, whereas the amplitude of contractions was significantly inhibited by atropine. These results suggest that intrinsic nitric oxide plays a crucial role in the regulation of duodenal tone and maintenance of continuous secretion by the exocrine pancreas in sheep. These results also implied that inhibition of pancreatic exocrine secretion by the nitric oxide synthase inhibitor is presumably mediated in part through the contractile effect on the duodenum. Accepted: 27 June 2000     

A secretin releasing peptide exists in dog pancreatic juice

Canine pancreatic juice has been shown to stimulate exocrine pancreatic secretion in the dog. In the present study we investigated whether there is a secretin-releasing peptide in canine pancreatic juice. Pancreatic juice was collected from the dogs with Thomas gastric and duodenal cannulas while pancreatic secretion was stimulated by intravenous administration of secretin at 0.5 microg/kg/h and CCK-8 at 0.2 microg/kg/h, respectively. The pancreatic juice was separated into three different molecular weight (MW) fractions (Fr) by ultrafiltration (Fr 1; MW > 10,000, Fr 2; MW=10,000-4,000 and Fr 3; MW < 4,000), respectively. All the fractions were bioassayed in anesthetized rats. Fraction 3 dose-dependently and significantly stimulated pancreatic juice flow volume from 78.0% to 99.4% (p<0.05) and bicarbonate output from 128.9% to 202.1% (p<0.01), respectively. Plasma secretin concentration also increased from 1.2 +/- 0.5 pM to 5.0 +/- 0.8 pM and 6.0 +/- 1.0 pM (p<0.05). None of these fractions increased pancreatic protein secretion or plasma CCK level. The stimulatory effect of Fraction 3 on pancreatic secretion and the release of secretin was completely abolished by treatment with trypsin (1 mg/ml for 60 min at 37 degrees C) but not by heating (100 degrees C, 10 min). Intravenous injection of a rabbit anti-secretin serum, which rendered plasma secretin almost undetectable in rat plasma, also abolished Fr 3-stimulated pancreatic secretion of fluid and bicarbonate secretion. These observations suggest that a secretin-releasing peptide exists in the canine pancreatic juice. It is trypsin-sensitive and heat-resistant. This peptide may play a significant physiological role on the release of secretin and regulation of exocrine pancreatic secretion.     

Neuronal regulation and function of natriuretic peptide receptor C

The neuromodulatory effect of natriuretic peptides is probably the best example of a whole organ response mediated by natriuretic peptide C receptors (NPR-Cs). Both NPR-C specific agonists and ablation experiments utilizing NPR-C specific antibodies or antisense oligonucleotides demonstrated the essential signaling role of the NPR-C in these neuromodulatory responses. Our most recent studies utilize peptides representing the NPR-C intracellular region to elucidate the specific signaling region of the NPR-C. These studies have identified an inhibitory influence of NPR-C on adrenergic neurotransmission by a calcium-dependent process.     

NPR-C receptors are involved in C-type natriuretic peptide response on bile secretion

C-type natriuretic peptide (CNP) is a member of the natriuretic peptide family. Previous studies reported the presence of natriuretic peptide receptors and mRNA CNP in the liver. In the present work, we sought to establish the role of CNP in the regulation of bile secretion in the rat and the possible pathways involved.CNP diminished basal as well as bile salt-evoked bile flow and bile acid output in a dose-dependent manner. It also reduced the excretion of sodium, chloride, and potassium but did not modify bile pH or the excretion of phospholipids, total proteins, and glutathione. Neither parasympathetic nor sympathetic blockade abolished CNP inhibitory response on bile secretion. The selective NPR-C agonist, C-ANP-(4-23) amide, diminished bile flow and the co-administration of both peptides did not further decrease it. CNP did not alter mean arterial pressure or portal venous pressure at any given doses.CNP decreased bile acid-dependent flow without affecting bile acid-independent flow. The inhibitory effect of CNP did not involve the participation of the autonomic nervous system or hemodynamic changes. The participation of NPR-C receptors in CNP response is strongly supported by present findings. The present study shows that CNP modulates bile secretion in the rat, suggesting that CNP may be part of the large family of peptides involved in the regulation of gastrointestinal physiology.     

Gastrointestinal secretory, motor and circulatory effects of corticotropin releasing factor (CRF)

This study was designed to determine the effects of CRF on the gastrointestinal functions such as secretion, motility and circulation in dogs. CRF was found to inhibit dose-dependently gastric acid response to pentagastrin but not to histamine. CRF stimulated pancreatic bicarbonate and protein secretion under basal conditions and in response to secretin or cholecystokinin (CCK). This stimulation was accompanied by an increase in plasma levels of pancreatic polypeptide (PP), but not of secretin or gastrin. CRF caused a partial inhibition of the migrating motor complexes in fasted dogs and increased spike activity of the small bowel. These motor effects of CRF probably resulted from the action of the released PP on the intestinal smooth muscle. CRF is also a potent and selective stimulant of the mesenteric blood flow. This effect may be secondary to the stimulation of intestinal motility and metabolism.     

Effects of gamma-aminobutyric acid on secretagogue-induced exocrine secretion of isolated, perfused rat pancreas

Because GABA and its related enzymes have been determined in beta-cells of pancreas islets, effects of GABA on pancreatic exocrine secretion were investigated in the isolated, perfused rat pancreas. GABA, given intra-arterially at concentrations of 3, 10, 30, and 100 microM, did not exert any influence on spontaneous or secretin (12 pM)-induced pancreatic exocrine secretion. However, GABA further elevated CCK (10 pM)-, gastrin-releasing peptide (100 pM)-, or electrical field stimulation-induced pancreatic secretions of fluid and amylase dose dependently. The GABA (30 microM)-enhanced CCK-induced pancreatic secretions were completely blocked by bicuculline (10 microM), a GABA(A) receptor antagonist, but were not affected by saclofen (10 microM), a GABA(B) receptor antagonist. The enhancing effects of GABA (30 microM) on CCK-induced pancreatic secretions were not changed by tetrodotoxin (1 microM) but were partially reduced by cyclo-(7-aminoheptanonyl-Phe-D-Trp-Lys-Thr[BZL]) (10 nM), a somatostatin antagonist. In conclusion, GABA enhances pancreatic exocrine secretion induced by secretagogues, which predominantly induce enzyme secretion, via GABA(A) receptors in the rat pancreas. The enhancing effect of GABA is partially mediated by inhibition of islet somatostatin release.     

Effects of putative galanin antagonists M35 and C7 on rat exocrine pancreas.

Galanin is a neuropeptide having a wide range of biological actions. Recently selective galanin receptor antagonists such as M35 [galanin(1-12)-Pro-bradykinin(2-9)-amide] and C7 [galanin(1-12)-Pro-spantide-amide] have been described. These antagonists have blocked the actions of galanin on flexor reflex, glucose-induced insulin secretion, and acetyicholine release from hippocampus. Our present aim was to investigate whether M35 and C7 can affect galanin-induced inhibition of pancreatic enzyme secretion in rats. Pancreatic enzyme secretion was studied in urethane-anesthetized rats supplied with jugular vein catheter and pancreatic cannula. Amylase secretion evoked by submaximal CCK-8 stimulation was inhibited dose-dependently by galanin in anesthetized rats. Surprisingly, neither M35 nor C7 was able to inhibit the responses of the exocrine pancreas to galanin. However, both putative galanin receptor antagonists behaved as agonists in our experimental models. Our data suggest that the effects of galanin on pancreatic enzyme secretion are not mediated by M35- or C7-sensitive galanin receptors. Therefore, these galanin receptors are different from those described in the central nervous system.     

Atrial natriuretic factor stimulates renal dopamine uptake mediated by natriuretic peptide-type A receptor

To determine the effects of atrial natriuretic factor (ANF) on renal dopamine (DA) metabolism, 3H-DA and 3H-L-DOPA uptake by renal tubular cells was measured in experiments carried out in vitro in Sprague-Dawley rats. The receptor type involved was also analyzed. The results indicate that ANF increased at 30 min, DA uptake in a concentration-response fashion having 10 pM ANF as the threshold concentration. Conversely, the uptake of the precursor L-DOPA was not modified by the peptide. ANF effects were observed in tissues from external and juxtamedullar cortex and inner medulla. On this basis, 100 nM ANF was used to continue the studies in external cortex tissues. DA uptake was characterized as extraneuronal uptake, since 100 microM hydrocortisone blocked ANF-induced increase of DA uptake. Renal DA uptake was decreased at 0 degrees C and in sodium-free medium. The effects of ANF in these conditions were not present, confirming that renal DA uptake is mediated by temperature- and sodium-dependent transporters and that the peptide requires the presence of the ion to exhibit its actions on DA uptake. The biological natriuretic peptide type A receptor (NPR-A) mediates ANF effects, since 100 nM anantin, a specific blocker, reversed ANF-dependent increase of DA uptake. The natriuretic peptide type C receptor (NPR-C) is not involved, since the specific analogous 100 nM 4-23 ANF amide has no effect on renal DA uptake and does not alter the effects of 100 nM ANF. In conclusion, ANF stimulates DA uptake by kidney tubular cells. ANF effects are mediated by NPR-A receptors coupled to guanylate cyclase and cGMP as second messenger. The process involved was characterized as a typical extraneuronal uptake, and characterized as temperature- and sodium-dependent. This mechanism could be related to DA effects on sodium reabsorption and linked to ANF enhanced natriuresis in the kidney. The increment of endogenous DA into tubular cells, as a consequence of increased DA uptake, would permit D1 receptor recruitment and Na+,K+-ATPase activity inhibition, which results in decreased sodium reabsorption and increased natriuresis.     

Effects of human atrial natriuretic polypeptide on pancreatic exocrine secretion in the dog

We examined the effects of atrial natriuretic polypeptide (hANP) on exocrine function in the isolated and blood-perfused dog pancreas in situ. Intra-arterial injection of hANP (1-10 micrograms) resulted in the dose-dependent increases of the pancreatic juice secretion. The secretory activity of 3 micrograms of hANP was approximately equal to one third of the secretory activity of 0.1 units of secretin. The use of hANP increased the concentration of bicarbonate but not that of sodium and protein in the pancreatic juice as compared with the basal values. These secretory responses to hANP were not inhibited by treatment with haloperidol, sulpiride, phentolamine, propranolol, atropine, cimetidine or ethacrynic acid. These results suggest that hANP acts directly on the pancreatic exocrine gland to stimulate pancreatic secretion; without, however, increasing sodium excretion. The mechanism of this effect remains to be elucidated.