Effect of acute hyperglycemia on basal, secretin and secretin + cholecystokinin stimulated exocrine pancreatic secretion in humans

Pancreatico-biliary secretion is reduced during acute hyperglycemia. We investigated whether alterations in pancreatico-biliary flow or volume output are responsible for the observed reduction in duodenal output of pancreatic enzymes and bilirubin during hyperglycemia. Eight healthy subjects were studied on two occasions during normoglycemia and hyperglycemia (15 mmol/l). Pancreatico-biliary output was measured by aspiration using a recovery marker under basal conditions (60 min), during secretin infusion (0.1 CU/kg.h) for 60 min and during secretin + CCK (0.5 IDU/kg.h) infusion for 60 min. Secretin was infused to stimulate pancreatico-biliary flow and volume output. Secretin significantly (P<0.005-P<0.05) increased volume and bicarbonate output and CCK significantly (P<0.01) increased the output of bilirubin, pancreatic enzymes, bicarbonate and volume, both during normoglycemia and hyperglycemia. During hyperglycemia basal, secretin stimulated and secretin + CCK stimulated total pancreatico-biliary output were significantly (P<0.005-P<0.05) reduced compared to normoglycemia. The incremental outputs, however, were not significantly different between hyper- and normoglycemia. Pancreatic volume output was significantly (P<0.05) reduced during hyperglycemia compared to normoglycemia under basal conditions (31+/-16 m/h versus 132+/-33 m/h) during secretin infusion (130+/-17 ml/h versus 200+/-34 m/h) and during secretin + CCK infusion (370+/-39 ml/h versus 573+/-82 ml/h). Plasma PP levels were significantly (P<0.05) reduced during hyperglycemia. It is concluded that 1) hyperglycemia significantly reduces basal pancreatico-biliary output 2) the incremental pancreaticobiliary output in response to secretin or secretin + CCK infusion is not significantly affected during hyperglycemia, 3) a reduction in volume output contributes to the inhibitory effect of hyperglycemia on pancreatico-biliary secretion, 4) hyperglycemia reduces PP secretion suggesting vagal-cholinergic inhibition of pancreatico-biliary secretion and volume during hyperglycemia.     

Specific protein phosphorylation during stimulation of amylase secretion by beta-agonists or dibutyryl adenosine 3',5'-monophosphate in the rat parotid gland

The present study was undertaken in order to examine the possible involvement of protein phosphorylation during beta-adrenergic stimulation in the rat parotid gland. Isolated parotid gland slices were stimulated by either isoproterenol or dibutyryl adenosine 3',5'-monophosphate (Bt2cAMP) in the presence or absence of propranolol. Amylase output was measured as a parameter for the degree of stimulation of secretion. Stimulation of secretion by either isoproterenol or Bt2AMP was associated with phosphorylation of three protein bands as revealed by sodium dodecylsulfate/polyacrylamide gel electrophoresis and autoradiography. The apparent molecular weights of the three proteins were 35,100 (protein I), 25,700 (protein II) and 20,400 (protein III). After cell fractionation by differential and gradient centrifugation, protein I was enriched in a light membrane fraction most likely corresponding to the plasma membrane as revealed by marker enzyme analysis. Proteins II and III were recovered in a denser fraction containing mainly mitochondria and rough microsomes. The effect of isoproterenol but not that of Bt2cAMP on phosphorylation of all three protein bands was completely abolished by propranolol. The different time course in the stimulation of amylase secretion by isoproterenol and Bt2cAMP respectively was reflected by corresponding differences in the time course of protein phosphorylation.     

Developmental change in the ability of estradiol to suppress testosterone secretion by the testis of the rat

An intratesticular site of action has been proposed for the ability of estradiol (E2) to suppress testosterone secretion. Because testicular testosterone and E2 secretion as well as E2 receptors change during development, a physiologic role for E2 is possible. The present experiments compared the testes from 12-day-old and adult rats for the capacity of in vivo estradiol treatment to change in vitro androgen secretion in response to luteinizing hormone (LH) and dibutyryl cyclic AMP (Bt2cAMP). After 5 days in vivo treatment, in vitro responsiveness was estimated by radioimmunoassay (RIA) measurement of androgen secretion elicited by various doses of NIAMDD-LH-24 or 1.0 mM Bt2cAMP. Five days of E2 alone (500 ng/g BW s.c. once daily) markedly inhibited basal, LH-stimulated and Bt2cAMP-stimulated androgen production at both ages. Similar treatment of infant rats with LH (100 ng NIAMDD-LH-24/g BW) caused an increase in basal and LH-stimulated androgen secretion in vitro, but had no effect on the response to Bt2cAMP. The same pretreatment of adults with LH had no effect on basal, but inhibited LH- or Bt2cAMP-stimulated androgen secretion. Combined treatment of infants with E2 and LH for 5 days had no effect on basal or maximally stimulated androgen production; the in vitro response to submaximal stimulation with LH was significantly inhibited. Combined E2/LH treatment of adults significantly decreased the basal production of androgens and the response to LH or Bt2cAMP. These results suggest a major difference between the response to E2 of the Leydig cells from the rats of the two ages tested.(ABSTRACT TRUNCATED AT 250 WORDS)     

The secretion of adenosin 3',5'-monophosphate after hydrokinetic and ecbolic stimulation in the canine pancreas (author's transl)

The secretion of cAMP is studied in vivo and in the isolated perfused canine pancreas after administration of secretin and CCK or caerulein in comparison with hydrokinetic or ecbolic secretory events as well as with the magnitude and time course of changes in tissue cAMP. 1) The total output of cAMP and pancreatic juice shows a significant and positive correlation after stimulation with secretin. The linear correspondence between cAMP concentration and secretory rates of pancreatic juice beyond 3 ml/5 min and their non-linear, reciprocal correlation at lower rates of fluid secretion point to an active as well as to a passive secretory mechanism for cAMP. 2) CCK and caerulein increase secretion of cAMP too. The output of cAMP however neither corresponds to the time course of protein secretion nor correlates quantitatively with the latter. 3) The behaviour of cAMP secretion and concentration in the pancreatic juice after administration of secretin and CCK or caerulein as well as differs from the changes in tissue cAMP levels. The respective maximum of cAMP output after addition of secretin or ecbolic secretagogues during the greatest decrease in cellular cAMP levels yields on the average about 1% of the estimated reduction in total tissue cAMP content. The results indicate a functional coherence in secretion of pancreatic juice and cAMP but oppose the assumption, that essential amounts of cAMP are released during exocytosis of zymogen granules. The secretion of cAMP may be possibly influenced by cytoplasmatic cAMP levels, but neither reflects the present changes in cellular cAMP nor seems to be of a regulatory importance for the latter.     

Hormonal regulation of amino acid uptake by cultured 3T3-L1 adipocytes

Incubation of the adipocytes for 20 hours with insulin or with Bt2cAMP plus the theophylline stimulated adipocyte uptake of AIB and MeAIB but did not stimulate the uptake of glutamine or cycloleucine. MeAIB uptake by both 3T3-L1 preadipocytes and 3T3-C2 cells was relatively unresponsive to insulin. However, MeAIB uptake by 3T3-C2 cells was stimulated by treatment with Bt2cAMP plus theophylline. Incubation of 3T3 adipocytes for 60 min with insulin yielded maximal stimulation of 2-deoxyglucose uptake but no stimulation of the uptake of AIB, MeAIB or glutamine. Responsiveness of transport to Bt2cAMP does not appear to require adipocyte differentiation. By contrast, adipocyte differentiation may be required for the development of the insulin-responsive transport systems.     

Inhibition of lipolysis in hamster adipocytes by the cation ionophor X537A.

The present study reports the effects of the lipophylic ionophore X537A on lipolysis and accumulation of cAMP in isolated hamster epidiymal adipocytes. X537A inhibited lipolysis activated with norepinephrine, isoproterenol, dibutyryl cAMP or theophylline but failed to influence basal lipolysis. The minimum effective concentration of X537A required to inhibit lipolysis was between 1 and 3 micrograms/ml; at a concentration of 10 micrograms/ml, X537A inhibited lipolysis by approximately 50%. The antilipolytic effect of X537A does not result from decreased formation of cAMP because the accumulation of cAMP in response to isoproterenol or theophylline was significantly potentiated in the presence of the ionophore. Most of the additional cAMP that accumulated in the presence of X537A was found to be intracellelular, the distribution of cAMP between cells and incubation medium not being influenced by X537A. Neither the basal activity of cAMP dependent protein kinase nor the activity in the presence of isoproterenol or theophylline was influenced by X537A. The effects of X537A on lipolysis and on accumulation of cAMP were found to persist in the absence of extracellular calcium, but adipocytes that were preincubated in a calcium free media containing 4.0 mM EGTA failed to respond to X537A with an increase in cAMP levels. It is concluded that X537A inhibits lipolysis by uncoupling cAMP accumulation from activation of triglyceride lipase by a mechanism unrelated to activation of protein kinase.     

The effect of secretin on acid and pepsin secretion and gastrin release in the totally isolated vascularly perfused rat stomach

The effect of secretin on acid and pepsin secretion and gastrin release in the totally isolated vascularly perfused rat stomach was studied. With the phosphodiesterase inhibitor isobutyl methylxanthine (IMX) added to the vascular perfusate, baseline acid secretion was 4.7 +/- 1.1 (mean +/- S.E.M.) mumol/h and baseline pepsin output 1147 +/- 223 micrograms/h. Secretin significantly inhibited acid output to a minimum of 1.4 +/- 0.2 mumol/h at a concentration of 25 pM in the vascular perfusate (P less than 0.01). Pepsin output was not significantly different from baseline at any of the secretin doses tested. Threshold secretin concentration for acid inhibition was 5 pM. IMX stimulated gastrin output from 48 +/- 9 pM in the basal state to 95 +/- 13 pM after IMX (P less than 0.01). Secretin inhibited gastrin release only at the maximal dose of 625 pM, when gastrin concentration in the venous effluent decreased from 93 +/- 19 to 68 +/- 19 pM after secretin. Thus, in the totally isolated vascularly perfused rat stomach secretin in physiological concentrations inhibits acid secretion by a direct action on the acid secretory process and not via gastrin inhibition. The study also suggests that gastrin release at least in part is mediated via increased intracellular cAMP.     

Cellular mechanism of sodium oleate-stimulated secretion of cholecystokinin and secretin

Long-chain fatty acids are potent stimulants of secretin and CCK release. The cellular mechanisms of fatty acid-stimulated secretion of these two hormones are not clear. We studied the stimulatory effect and mechanism of sodium oleate (SO) on secretin- and CCK-producing cells. SO stimulated the release of secretin or CCK from isolated rat mucosal cell preparations enriched in either secretin- or CCK-producing cells, respectively. SO also time- and dose-dependently stimulated secretin and CCK release from STC-1 cells. In STC-1 cells, SO-stimulated secretin and CCK release was potentiated by IBMX and inhibited by a protein kinase A-selective inhibitor and a cAMP-specific antagonist. SO-stimulated releases of the two hormones were also inhibited by downregulation or inhibitors of protein kinase C, a calmodulin antagonist and an inhibitor of calmodulin-dependent protein kinase II. Chelating of extracellular Ca(2+) or addition of an L-type calcium channel blocker diminished SO-stimulated hormone releases. SO caused an increase in intracellular Ca(2+) concentration that was partially reversed by diltiazem but had no effect on production of cAMP, cGMP, or inositol-1,4,5-triphosphate. These results indicate that SO acts on secretin- and CCK-producing cells. Its stimulatory effect is potentiated by endogenous protein kinase A and mediated by activation of Ca(2+) influx through the L-type channels and of protein kinase C and Ca(2+)/calmodulin-dependent protein kinase II.     

Differential actions of phosphodiesterase inhibitors on secretin- and vasoactive intestinal peptide-induced increases in chief cell cAMP

The actions of three different phosphodiesterase inhibitors, theophylline, 3-isobutyl-1-methylxanthine (IBMX) and Ro 20-1274 (Ro), on cellular cAMP and pepsinogen secretion from dispersed chief cells prepared from guinea pig stomach were examined. The relative order of potency for increasing cAMP and pepsinogen secretion was Ro > IBMX > theophylline. Ro, the most efficacious agent, caused a 17-fold increase in basal cAMP and a similar augmentation of the increase in cAMP caused by secretin or vasoactive intestinal peptide (VIP). Differential actions of these agents on the dose-response curves for secretin- and VIP-induced increases in cAMP suggest that chief cell receptors for these peptides are coupled to pools of cAMP that are acted upon by heterogeneous phosphodiesterases with varying sensitivities to inhibitors. Moreover, Ro, a selective inhibitor of low K_m cAMP-specific phosphodiesterases, is the most potent and efficacious agent tested in this cell system.     

The effect of secretin on pentagastrin-stimulated secretion of gastric pepsin and acid in rats.

Rats with chronic gastric fistulas were stimulated for 12 or 24 h with constant intravenous infusion of pentagastrin. When secretin was also infused for the last half period of the experiment, respectively, 6 or 12 h, the volume of gastric secretion and HCl output were significantly reduced but the concentration of pepsin was significantly increased. The dissociated effect of secretin on gastric acid and pepsin secretion reported previously in man, dog and cat was also found in the rat.     

Gs protein-coupled receptor agonists induce transactivation of the epidermal growth factor receptor in T84 cells: implications for epithelial secretory responses

We have previously shown that Gq protein-coupled receptor (GqPCR) agonists stimulate epidermal growth factor receptor (EGFr) transactivation and activation of mitogen-activated protein kinases (MAPK) in colonic epithelial cells. This constitutes a mechanism by which Cl- secretory responses to GqPCR agonists are limited. In the present study we examined a possible role for the EGFr in regulating Cl- secretion stimulated by agonists that act through GsPCRs. All experiments were performed using monolayers of T84 colonic epithelial cells grown on permeable supports. Protein phosphorylation and protein-protein interactions were analyzed by immunoprecipitation and Western blotting. Cl- secretion was measured as changes in short-circuit current (DeltaIsc) across voltage-clamped T84 cells. The GsPCR agonist, vasoactive intestinal polypeptide (VIP; 100 nM), rapidly stimulated EGFr phosphorylation in T84 cells. This effect was mimicked by a cell-permeant analog of cAMP, Bt2cAMP/AM (3 microM), and was attenuated by the protein kinase A (PKA) inhibitor, H-89 (20 microM). The EGFr inhibitor, tyrphostin AG1478 (1 microM), inhibited both Bt2cAMP/AM-stimulated EGFr phosphorylation and Isc responses. VIP and Bt2cAMP/AM both stimulated ERK MAPK phosphorylation and recruitment of the p85 subunit of phosphatidylinositol 3-kinase (PI3K) to the EGFr in a tyrphostin AG1478-sensitive manner. The PI3K inhibitor, wortmannin (50 nM), but not the ERK inhibitor, PD 98059 (20 microM), attenuated Bt2cAMP/AM-stimulated secretory responses. We conclude that GsPCR agonists rapidly transactivate the EGFr in T84 cells by a signaling pathway involving cAMP and PKA. Through a mechanism that likely involves PI3K, transactivation of the EGFr is required for the full expression of cAMP-dependent Cl- secretory responses.     

Regulation of aryl hydrocarbon (benzo-(A)-pyrene) hydroxylase activity in mammalian cells. Induction of hydroxylase activity by N6,O2'-dibutyryl8 adenosine 3':5'-monophosphate and aminophylline.

Treatment of hamster BHK cells with N6,O2'-dibutyryl adenosine 3':5'-monophosphate (Bt2cAMP), aminophylline, theophylline, or papaverine increased the level of aryl hydrocarbon (benzo(a)pyrene) hydrolxylase activity. The highese increase, 100-fold, was obtained with Bt2cAMP plus aminophylline or theophylline. N2,O2-Dibutyryl guanosine 3':5'-monophosphate gave a lower induction than Bt2cAMP. The level of hydroxylase activity started to decrease 6 hours after treatment with the inducer and was reduced to almost the uninduced level after 24 hours. Repeated addition of Bt2cAMP and aminophylline did not prevent this decrease. The hydroxylase can also be induced by treating cells with benz(a)anthracene, and the level of this induced activity was maintained for 24 hours. Aminophylline gave a 2- to 8-fold stimulation of the induction by benz(a)anthracene. The enzyme activity induced by Bt2cAMP, aminophylline, and benz(a)anthracene converted benzo(a)pyrene to similar alkali-extractable metabolities with a fluorescence spectra similar to that of 3-hydroxybenzo(a)pyrene. These induced enzyme activities also showed a similar heat stability. Induction by Bt2cAMP and aminophylline, like induction by benz(a)anthracene, required continued protein synthesis and only an initial period of RNA synthesis. Compared to the benz(a)anthracene-induced hydroxylase with a Km of 4.3 muM, the hydroxylase induced by Bt2cAMP and aminophylline showed a Km of 0.14 muM, and was 100-fold more sensitive to inhibition by 7,8-benzoflavone. Increasing the serum concentration in the culture medium stimulated the induction by aminophylline but did not stimulate induction by benz(a)anthracene. The results indicate that aryl hydrocaarbon (benzo(a)pyrene) hydroxylase can be induced by compounds that increase the level of adenosine 3':5'-monophosphate and that this induction and induced enzyme activity differs from that caused by benz(a)anthracene.     

The regulation of glucose transport by cAMP stimulators via three different mechanisms in rat and human adipocytes

The regulation of glucose transport by a beta-adrenergic agonist and other cAMP stimulators was assessed by kinetic analyses of 3-O-methylglucose (MG) transport in rat and human adipocytes and in isolated rat plasma membrane vesicles. Basal MG transport was biphasically affected by L-isoproterenol in rat adipocytes: lower concentrations (10-25 nM) of L-isoproterenol stimulated the basal rate by increasing the Vmax, but higher concentrations (0.5-2 microM) of L-isoproterenol inhibited the basal rate. On the other hand, the maximum insulin-stimulated MG transport rate was not affected by 25 nM L-isoproterenol, but was suppressed by 2 microM L-isoproterenol in rat adipocytes. In the presence of adenosine deaminase plus L-isoproterenol (25 nM and 2 microM), dibutyryl cyclic AMP (Bt2cAMP), 3-isobutyl-1-methylxanthine, or forskolin, both basal and the maximum rates of MG transport were suppressed in rat adipocytes. However, from kinetic experiments, both L-isoproterenol plus adenosine deaminase and Bt2cAMP decreased the Vmax. On the other hand, isobutymethylxanthine and forskolin decreased the Vmax as well as increased the K8. MG transport in plasma membrane vesicles was directly inhibited by either forskolin or isobutylmethylxanthine. In contrast, both 25 nM and 2 microM L-isoproterenol with or without adenosine deaminase, Bt2cAMP, or cAMP had no effect on MG transport in rat plasma membrane vesicles. In human adipocytes, L-isoproterenol always stimulated basal MG transport and did not suppress the maximum rate of MG transport, even though cAMP production was maximally stimulated by L-isoproterenol. Both adenosine deaminase plus L-isoproterenol and Bt2cAMP did not suppress the basal rate, but did show a modest suppression (40%) of the maximum insulin effect on MG transport in human adipocytes. However, both isobutylmethylxanthine and forskolin remarkably suppressed (85%) both the basal and the maximum rate of MG transport by both increasing the K8 and decreasing the Vmax. These results indicate MG transport in both rat and human adipocytes is regulated by 3 different mechanisms: (I) L-isoproterenol, a beta-adrenergic agonist, stimulates basal MG transport by increasing the Vmax, (II) cAMP mediates a decrease in MG transport by decreasing the Vmax, and (III) both forskolin and isobutylmethylxanthine also decrease MG transport by directly inhibiting the binding of MG molecules to transporters, resulting in a decrease in the Vmax and an increase in the K8.     

Cyclic adenosine 3',5'-monophosphate suppresses interleukin 1-induced synthesis of matrix metalloproteinases but not of tissue inhibitor of metalloproteinases in human uterine cervical fibroblasts.

Interleukin 1 (IL-1) mediates many cellular functions, but the signal transduction mechanisms of its actions are not clearly understood. Here, we have examined the exact participation of cAMP in the IL-1-induced production of the precursors of matrix metalloproteinase (MMPs) and their specific inhibitor, tissue inhibitor of metalloproteinases (TIMP) in human uterine cervical fibroblasts. IL-1 significantly augmented the production of proMMP-1 (vertebrate procollagenase), proMMP-3 (prostromelysin), and TIMP without detectable changes in the intracellular level of cAMP. Dibutyryl cAMP (Bt2cAMP) and the cAMP elevating agent (forskolin) did not replace IL-1 as MMP inducers. On the contrary, the IL-1-mediated induction of proMMP-1 and proMMP-3 was significantly suppressed by treatment of the cells with Bt2cAMP, forskolin, or theophylline. The suppressive effect of Bt2cAMP on the IL-1-induced production of proMMP-1 and -3 was not due to the inhibition of zymogen secretion, but resulted from the decrease in the steady-state levels of proMMP-1 and proMMP-3 mRNAs. In contrast, Bt2cAMP slightly enhanced the IL-1-induced production of TIMP. The synthesis of proMMP-2 (72-kDa progelatinase/type IV procollagenase) was not altered by IL-1 and/or Bt2cAMP. These results suggest, first, that induction of proMMP-1 and -3 synthesis may share similar transduction pathways but they are distinct from those for proMMP-2 and TIMP synthesis and, second, that cAMP does not function as a second messenger in the MMPs' induction upon IL-1 stimulation in human uterine cervical fibroblasts. Thus, it is further suggested that the system that increases the intracellular cAMP level may be involved in negative regulation of proMMP-1 and -3 production.     

Behavior of cellular cyclic nucleotides after hydrokinetic or ecbolic stimulation of the canine pancreas

The studies deal with the influence of secretin and various ecbolic secretagogues on tissue levels of cAMP and cGMP in vivo and in the isolated perfused canine pancreas. The mutual behaviour of cellular cAMP and cGMP is observed and compared with the time course of the respective secretory events. Synthetic secretin as well as CCK, acetylcholine or Caerulein likewise elevate tissue cAMP and cGMP simultaneously. There exists no difference in the magnitude of increase and in the time course of changes in tissue cyclic nucleotide levels between hydrokinetic and ecbolic stimulation. The rise in cAMP and cGMP coincides with the onset of the respective secretory events and reaches peak values contemporarily to the excretory maxima. The following decrease in tissue cyclic nucleotides approximatively parallels juice or enzyme secretion in the isolated perfused pancreas but differs widely in vivo. Under this condition cAMP and cGMP rapidly fall to basal levels during undiminished excretory function and show a second rise after cessation of the latter. Secretin and various ecbolic secretagogues do not increase tissue content of cyclic nucleotides in the same dose-dependent manner as can be observed with pancreatic secretion. The behaviour of cAMP and cGMP after addition of secretin and CCK or acetylcholine remains widely unchanged during calcium-free perfusion in spite of an extensive excretory inhibition. The corresponding rise in cellular cAMP and cGMP in the sequence of hydrokinetic as well as of ecbolic stimulation points to an analogous intracellular mediation of various secretagogues in different target cells of the exocrine canine pancreas.     

LDL and cAMP cooperate to regulate the functional expression of the LRP in rat ovarian granulosa cells

Rat ovarian granulosa rely heavily on lipoprotein-derived cholesterol for steroidogenesis, which is principally supplied by the LDL receptor- and scavenger receptor class B type I (SR-BI)-mediated pathways. In this study, we characterized the hormonal and cholesterol regulation of another member of the LDL receptor superfamily, low density lipoprotein receptor-related protein (LRP), and its role in granulosa cell steroidogenesis. Coincubation of cultured granulosa cells with LDL and N6,O2'-dibutyryl adenosine 3',5'-cyclic monophosphate (Bt2cAMP) greatly increased the mRNA/protein levels of LRP. Bt2cAMP and Bt2cAMP plus human hLDL also enhanced SR-BI mRNA levels. However, there was no change in the expression of receptor-associated protein, a chaperone for LRP, or another lipoprotein receptor, LRP8/apoER2, in response to Bt2cAMP plus hLDL, whereas the mRNA expression of LDL receptor was reduced significantly. The induced LRP was fully functional, mediating increased uptake of its ligand, alpha2-macroglobulin. The level of binding of another LRP ligand, chylomicron remnants, did not increase, although the extent of remnant degradation that could be attributed to the LRP doubled in cells with increased levels of LRP. The addition of lipoprotein-type LRP ligands such as chylomicron remnants and VLDL to the incubation medium significantly increased the progestin production under both basal and stimulated conditions. In summary, our studies demonstrate a role for LRP in lipoprotein-supported ovarian granulosa cell steroidogenesis.     

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.