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At least 37 records · Page 2Linked to original sources
PubMed1993

Neurophysin-containing pathway from the paraventricular nucleus of the hypothalamus to a sexually dimorphic motor nucleus in lumbar spinal cord.

A model that has been widely used in the study of steroid sensitive neurons, the spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus in the lower lumbar spinal cord that innervates the striated bulbocavernosus (BC) muscle. The BC is responsible for penile reflexes in the male rat, which are important in ensuring pregnancy in females. The characterization of afferents to the SNB aids in the understanding of the neural circuitry involved in reproductive behavior. We have recently identified the paraventricular nucleus (PVN) as a possible source of afferents to the SNB. Because the PVN is the major source of oxytocin/vasopressin within the central nervous system (CNS), the purpose of the present study was to examine and characterize a neurophysin (NP)-containing pathway from the PVN to the SNB. The results demonstrate that neurons of the lateral parvicellular subnucleus of PVN, which project to levels of spinal cord containing SNB motoneurons, contain NP, the coproduct of oxytocin and vasopressin. NP-containing fibers and putative terminals were found in the region of the SNB and appear to contact the soma and proximal dendrites of SNB motoneurons which were retrogradely identified as BC-innervating. Electrolytic lesions, which destroy the lateral parvicellular subnucleus of PVN, abolish NP-containing fibers in the region of the SNB, suggesting that the PVN is the source of these NP fibers. The results of this study indicate a NP-containing projection from the hypothalamus directly to SNB motoneurons. It is suggested that this pathway may play a role in the integration of penile reflexes with other aspects of male copulatory behavior that are under hypothalamic control.

Animals↗
PubMed1995

Expression and functional role of nitric oxide synthase in osteoblast-like cells.

Nitric oxide synthases (NOS) are enzymes that produce nitric oxide (NO) from L-arginine in a reaction yielding citrulline as a coproduct. Nitric oxide modulates the activity of a wide variety of cells, but little is known about its effects on bone cells. In the present study we report that the NOS inhibitor NG-monomethyl-L-arginine (NMMA) induced a dose-dependent inhibitory effect on the proliferation of the osteoblast-like cell lines MG63 and ROS 17/2.8. The inhibitory effect was prevented by increasing L-arginine concentrations in the medium and by the NO donor sodium nitroprusside. Likewise, NMMA inhibited interleukin-6 secretion, independently of its effect on cell number. NOS expression by MG63 cells was confirmed by measuring their ability to metabolize radiolabeled L-arginine to citrulline. NOS bioactivity was detected in unstimulated cells, but was markedly increased by stimulating the cells with cytokines, lipopolysaccharide, or 1,25-dihydroxyvitamin D3. NOS activity was partially dependent upon the presence of calcium in the medium. Furthermore, constitutive-type NOS (c-NOS) and inducible-type NOS (i-NOS) mRNA expression was detected in ROS 17/2.8 cells after reverse transcription and polymerase chain reaction amplification. In conclusion, osteoblast-like cells express c-NOS and i-NOS, and NOS activity seems to play an important role in the regulation of cell proliferation and function.

Amino Acid Oxidoreductases↗
PubMed1995

Mycoplasma stimulates the production of oxidative radicals by murine peritoneal macrophages.

Mycoplasmas and mycoplasma membranes have been shown to induce the production of inflammatory cytokines, including tumor necrosis factor alpha and interleukin-6, as well as nitric oxide, by mouse macrophages and rat brain astrocytes. Luminol-enhanced chemiluminescence was used as a sensitive method to show that Mycoplasma capricolum membranes induce mouse peritoneal macrophages to produce reactive oxygen radicals. Coincubation of the mycoplasma with a secondary stimulus, namely macrophage-activating factor or interferon-gamma, increased the chemiluminescence. The augmentation was abolished by the nitric oxide synthase inhibitor NG-methyl-L-arginine, indicating the involvement of nitric oxide. The coproduction of superoxide and nitric oxide by the same cell allows the formation of the powerful oxidant peroxynitrite, which could be responsible for the increased chemiluminescence. Induction of oxidizing radicals by mycoplasmas may contribute to the clinical pathology seen in mycoplasma infections.

Animals↗
PubMed1992

Phenotypic plasticity of avian embryonic sympathetic neurons grown in a chemically defined medium: direct evidence for noradrenergic and cholinergic properties in the same neurons.

Avian embryonic sympathetic ganglia possess both catecholaminergic and cholinergic features and can synthesize noradrenaline (NAd) and acetylcholine (ACh) simultaneously. In the present study we sought to determine (1) whether or not this coproduction of NAd and ACh corresponds to the existence of two non-overlapping populations, and (2) to what extent the levels of synthesis are influenced by non-neuronal ganglion cells. We have focused on the correlation between the immunocytochemically demonstrable presence of the noradrenergic and cholinergic enzymes tyrosine hydroxylase (TH) and choline acetyltransferase (ChAT), respectively, and the synthesis of the corresponding neurotransmitters in embryonic quail sympathetic neuronal and non-neuronal cells purified by fluorescence-activated cell sorting. We show that (1) freshly sorted neurons synthesize both NAd and ACh, whereas non-neuronal cells produce neither; (2) the overwhelming majority of the sympathetic neurons display TH immunoreactivity; (3) about half of the TH-positive neurons are recognized by an anti-ChAT antibody in an artificial medium that selectively enhances synthesis and/or accumulation of ACh; (4) the non-neuronal cells are important for survival of the neurons and potentiate their synthesis of ACh in this medium, and (5) finally, we present evidence that expression of TH in noradrenergic neurons and in small intensely fluorescent cells of sympathetic ganglia is differentially regulated.

Animals↗
PubMed1997

Improved methods to measure end products of nitric oxide in biological fluids: nitrite, nitrate, and S-nitrosothiols.

The aim of this study was to compare and improve standard methods to determine nitrite (NO2-), nitrate (NO3-) and S-nitrosothiol (RSNO) levels in cell culture supernatants, sera, and urine. We modified the conventional Griess reaction by replacing sulfanilamide with dapsone (4,4'-diamino-diphenylsulfone) and compared the NO2- levels in our study samples with a commercially available NO2- assay kit. Our modification, along with ultrafiltration of the samples, resulted in an enhanced sensitivity to measure NO2- down to 0.2 microM. The detection limit was further improved to 0.02 microM when NO2- was identified by the fluorochrome 2,3-diaminonaphthalene (DAN). To measure the stable end product NO3- by the Griess reaction or the DAN method, this anion must be reduced to NO2-. We compared the capacity of bacterial nitrate reductase with the reducing metal cadmium to convert NO3- to NO2-. After reduction, NO2- levels were determined either by the DAN method or by our modified Griess reaction. We found that there was a high correlation (r2 = 0.998) in total NO2- concentrations in the study samples using both methods for reducing NO3- to NO2-. The simultaneous determination of NO2- and NO3- was achieved by using anion-exchange chromatography (HPLC; Polyspher IC AN-1 column). The detection limit of this assay for each anion is 0.5 microM, and it can be applied equally well to sera, urine, and culture media. We also adapted the DAN method to determine RSNO levels in our study samples. Using this approach, we were able to measure RSNO levels down to 0.15 microM. As result we discovered that RSNO levels were markedly increased in urine from septic patients and in supernatants from cytokine-stimulated human tumor cell lines. L-Citrulline, a coproduct of NO biosynthesis, was measured using a colorimetric assay with a sensitivity limit of 3.0 microM. Increased L-citrulline levels in media from cultured cells, but not in sera or urine, correlated with increased NO production. Although all methods studied were suitable for quantifying end products of NO in biological fluids and media, the use of bacterial reductase and the modified Griess reaction proved successful to provide the greatest sensitivity and linear range for routine measurements of NO2- and NO3-.

Body Fluids↗
PubMed1997

Self-assembly and protein-protein interactions between the SV40 capsid proteins produced in insect cells.

Soluble SV40 capsid proteins were obtained by expression of the three late genes, VP1, VP2, and VP3, in Sf9 cells using baculovirus expression vectors. Coproduction of the capsid proteins VP1, VP2, and VP3 was achieved by infecting Sf9 cells with the three recombinant baculovirus species at equal multiplicities. All three proteins were found to be localized in the nuclear fraction. Electron microscopy of nuclear extracts of the infected cells showed an abundance of SV40-like capsid structures and heterogeneous aggregates of variable size, mostly 20-45 nm. Under the same staining conditions wild-type SV40 virions are 45 nm. The capsid-like particles sedimented in glycerol gradients similarly to authentic wild-type SV40 virions. Pentamers of the major capsid protein VP1 were also seen. Protein analysis on sucrose gradients demonstrated that the capsid-like particles can be disrupted by treatment with the reducing agent dithiothreitol and the calcium chelator EGTA. The capsid-like particles were found to be significantly less stable than SV40 virions and were partially stabilized by calcium ions. Understanding the complex interactions between the capsid proteins is important for the development of an efficient in vitro packaging system for SV40 virions and pseudovirions.

Animals↗
PubMed1994

Ectopic ACTH secretion due to a bronchopulmonary carcinoid localized by somatostatin receptor scintigraphy.

We present the case of a 65-year-old woman with an adrenocorticotropic hormone (ACTH) secreting bronchopulmonary carcinoid. This patient showed the typical long history of Cushing's syndrome, including hypokaliemia, impaired glucose tolerance, high levels of ACTH and beta-endorphin, and coproduction of other peptides. At the onset of clinical symptoms in 1979 an adrenal adenoma was suspected, and left-sided adrenalectomy was performed. The symptoms soon recurred, and the diagnosis of ACTH-dependent Cushing's syndrome was made. As no ACTH-secreting tumor was found, the right adrenal was resected, and the patient was followed up regularly. Fourteen years later chest roentgenography and computed tomography revealed a para-aortic pulmonary lesion, which was suspicious for a bronchopulmonary carcinoid. ACTH and beta-endorphin were excessively, pancreatic polypeptide slightly elevated at that time. The final diagnosis was made using somatostatin receptor scintigraphy which confirmed the hormonal activity of the suspicious lesion; no additional focus was found. This method turned out to be not only a useful additional localization technique but also a promising tool for characterization and staging of a suspected ACTH-producing carcinoid. The tumor was resected curatively, and the diagnosis was confirmed histologically.

ACTH Syndrome, Ectopic↗
PubMed1987

The inhibition and disposition of intestinal alkaline phosphatase.

A primary mechanism of amino acid inhibition of intestinal alkaline phosphatase is postulated to be the formation of a dissociable enzyme-amino acid complex at an allosteric zinc site. The degree of inhibition was highly correlated with the Zn2+ stability constant of each amino acid and the inhibition was reversible by the addition of exogenous Zn2+ or by dialysis. This allosteric amino acid inhibition proved to be a useful probe of the membrane arrangement of the enzyme in the intact tissue. The catalytic site appears to face the lumen based on the poor permeability of the substrate, the accumulation of the coproducts in the luminal bath, and the response of the enzyme to luminal pH. Amino acid inhibition of alkaline phosphatase in the intact tissue was only effective in the presence of sodium; whereas sodium was not required in butanol extracted preparations which lacked the sidedness of the intact tissue. Since amino acid uptake from the intestine is sodium dependent, the allosteric inhibitory site is probably intracellular. The results suggest that the intestinal alkaline phosphatase spans the apical membrane with the catalytic site accessible from the lumen and the allosteric inhibitory site from the cytoplasm.

Alkaline Phosphatase↗
PubMed1993

Production of parathyroid hormone and parathyroid-hormone-related protein by breast cancer cells in culture.

Parathyroid-hormone-related protein (PTHrP) has been implicated in the origin of malignant hypercalcaemia. However, PTHrP production is not restricted to neoplastic cells, it is widespread among a variety of normal cell types and tissues. A physiological role for PTHrP has not been well defined. We describe a case of breast cancer with bone metastases and humoral hypercalcaemia of malignancy, with high levels of plasma C-terminal parathyroid hormone (PTH), mid-molecule PTH and PTHrP. Cells from breast cancer biopsies were cultured and medium samples assayed for the C-terminal and mid-molecule fragments, intact PTH and PTHrP. The data indicate a progressive increase in both PTH fragments and PTHrP levels, over a period of 30 days. No temporal parallelism exists between PTH fragments and PTHrP concentrations, the former being maximum at the 14th day, and the latter at the 30th day from the beginning of the culture. Our results indicate a coproduction of PTH and PTHrP by the breast cancer cells both in vivo and in vitro.

Aged↗
PubMed1994

Free oxygen radicals in restraint-induced stress gastritis in the rat.

In this experimental study, the role of free oxygen radicals (FOR) in stress gastritis (SG) was investigated in a restraint stress model for rats. Allopurinol, dimethyl sulfoxide (DMSO), and superoxide dismutase (SOD) were tested as single agents in controlled groups. The portal blood pH values, the ratio of the mucosal erosion area to the gastric mucosal area (EA/MA), the ratio of the depth of mucosal erosions to the concomitant gastric mucosal wall (ED/MD), and concentrations of malondialdehyde (MDA)--a lipid peroxidation coproduct--in the gastric mucosa were used as parameters in the experiment. The EA/MA between the treated and untreated control groups were found to have no statistically significant difference (P > 0.05). ED/MD, a crucial determinant for bleeding probability, was found to be decreased in the SOD group (P < 0.05). SOD, allopurinol, and DMSO reduced the mucosal MDA concentration to lower values than the untreated group (P < 0.05). We concluded that although FOR may not play a dominant role in stress-induced gastric lesions, SOD may be a good candidate for a clinical trial on SG prophylaxis.

Allopurinol↗
PubMed1992

Determination of glutamic acid decarboxylase activity and inhibition by an H2O2-sensing glutamic acid oxidase biosensor.

The catalytic activity of the enzyme L-glutamic acid decarboxylase (GAD) is determined by an amperometric method based on a recently developed glutamate-selective biosensor. The biosensor is composed of an amperometric H2O2 electrode and a biocatalytic membrane containing the enzyme glutamic acid oxidase (GAO). The biosensor allows the direct and continuous measurement of GA levels by monitoring the H2O2 produced at the electrode interface as a coproduct of the GAO-catalyzed GA oxidation to alpha-ketoglutaric acid. Since GA is transformed to gamma-aminobutyric acid and CO2 under the catalytic activity of GAD, the rate of GA consumption in solution, monitored by the GAO biosensor, represents a reliable measure of GAD catalytic activity. Additional experiments performed in the presence of different concentrations of the GAD inhibitor valproic acid have shown the suitability of the proposed approach for the study of GAD inhibitors also. Discussion of the main experimental characteristics of this new analytical method is given in terms of sensitivity, reproducibility, and reliability of the experimental results and ease, time, and cost of operation.

Amino Acid Oxidoreductases↗
PubMed1996

Induction of neuronal nitric oxide after hypothermic circulatory arrest.

BACKGROUND: Although hypothermic circulatory arrest (HCA) has become routine practice in cardiac surgery, it is associated with substantial neurotoxicity. We tested the hypothesis that increased nitric oxide production during HCA participates in neuronal death. We previously described a canine survival model of HCA that produces a consistent neurologic deficit and histopathologic pattern of selective neuronal death. METHODS: Adult male hound dogs (n = 17) were subjected to 2 hours of HCA at a brain temperature of 18 degrees C and reperfused to normothermia; they were sacrificed at various intervals up to 74 hours. Using in vivo cerebral microdialysis, dogs (n = 5) were given a simultaneous infusion of artificial cerebrospinal fluid containing L-[14C]arginine or L-[14C]arginine and L-nitroarginine methyl ester (a nitric oxide synthase inhibitor) in contralateral hemispheres while undergoing 2 hours of HCA and reperfusion to normothermia. RESULTS: L-[14C]citrulline recovery, a coproduct of nitric oxide, significantly increased during HCA in the hemisphere without the inhibitor (at 300 minutes: control, 236 +/- 94 fmol/min versus L-nitroarginine methyl ester, 6 +/- 6 fmol/min; p < 0.05). Citrulline production in vitro from canine cortical homogenates in the presence of calcium (n = 12) was significantly greater 8 and 20 hours after reperfusion (5.11 +/- 0.54 x 10(-7) mmol.mg-1.min-1 and 7.52 +/- 0.59 x 10(-7) mmol.mg-1.min-1, respectively) than before HCA (1.51 +/- 0.09 x 10(-7) mmol.mg-1.min-1; p < 0.05). Nitric oxide metabolites in the serum were also increased significantly early after reperfusion (baseline, 6.72 +/- 0.95 mmol/L; at 4 hours, 17.58 +/- 1.46 mmol/L; p < 0.05). Immunocytochemical staining of the cortex with neuronal nitric oxide synthase-specific monoclonal antibodies (Transduction Labs) revealed increased neuronal nitric oxide synthase expression 6 to 18 hours after HCA. Darkfield analysis demonstrated neuronal nitric oxide synthase localization to neuronal processes with widespread formation of dense plexi of nitric oxide synthase fibers. CONCLUSIONS: We conclude that neurotoxicity after HCA involves a significant, early induction in neuronal nitric oxide synthase expression in neuronal processes leading to widespread augmented nitric oxide production in the brain.

Animals↗
PubMed1991

Purification and characterization of the monoterpene cyclase gamma-terpinene synthase from Thymus vulgaris.

The monoterpene cyclase, gamma-terpinene synthase, from Thymus vulgaris (thyme) leaves was purified to apparent homogeneity by isoelectric focusing and dye-ligand, anion-exchange, hydrophobic interaction, and gel permeation chromatography. The enzyme has a native molecular weight of 96,000 as determined by gel permeation chromatography, and exhibited a specific activity of 538 nmol/h.mg protein (turnover number of approximately 0.01/s). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed the enzyme to be composed of two apparently identical subunits of Mr approximately 55,000. The protein was very hydrophobic, and possessed a pI value of 4.85 as determined by isoelectric focusing. Maximum activity was observed at pH 6.8 in the presence of 20 mM Mg2+; 5 mM Mn2+ could support catalysis, albeit at a much lower rate. The Km value for the substrate, geranyl pyrophosphate, was 2.6 microM. Cyclase activity was inhibited by cysteine- and histidine-directed reagents. Purified gamma-terpinene synthase also possessed the ability to cyclize geranyl pyrophosphate to small amounts of alpha-thujene and to lesser quantities of myrcene, alpha-terpinene, limonene, linalool, terpinen-4-ol, and alpha-terpineol, all of which appear to be coproducts of the reaction sequence leading to gamma-terpinene. In general properties, the gamma-terpinene synthase from thyme leaves resembles other monoterpene cyclases as well as sesquiterpene and diterpene cyclases.

Alkyl and Aryl Transferases↗
PubMed1993

NADPH-diaphorase activity in neurons of the mammalian pancreas: coexpression with vasoactive intestinal polypeptide.

BACKGROUND: To provide a morphological basis for understanding the role of nitric oxide in the pancreas, the present study was designed to clarify the localization and distribution of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) activity, a marker of NO synthase, in the pancreas of several mammalian species, including humans. METHODS: NADPH-d activity was examined in the rat, guinea pig, dog, and human pancreas by histochemistry. In addition, the possibility of coproduction of NO and vasoactive intestinal polypeptide (VIP) was investigated by a combined use of histochemistry and immunohistochemistry. RESULTS: In the pancreas, NADPH-d activity was localized in nerve fibers, nerve cell bodies, and the vascular endothelium. Nerve fibers with the enzyme activity were chiefly distributed in the exocrine pancreas and showed species differences in the distribution. Nerve fibers stained for NADPH-d were also observed in the endocrine pancreas, but the enzyme activity was not detected in the islet cells. Part of the nerve fibers and nerve cell bodies coexpressed NADPH-d activity and VIP-immunoreactivity. CONCLUSIONS: These results suggest that NO may act as a neuronal mediator and an endothelium-derived relaxing factor and may physiologically interact with VIP in the mammalian pancreas.

Amino Acid Oxidoreductases↗
PubMed1995

Nitric oxide modulates pepsinogen secretion induced by calcium-mediated agonist in guinea pig gastric chief cells.

BACKGROUND & AIMS: Nitric oxide, a putative cellular messenger synthesized from L-arginine, is a powerful modulator of gastric motility and secretions. The aim of this study was to investigate whether (1) guinea pig gastric chief cells express NO synthase, (2) NO modulates the pepsinogen secretion and guanosine 3',5'-cyclic monophosphate (cGMP) generation induced by calcium (Ca2+)-mediated agents, and (3) NO donors and cGMP analogues stimulate pepsinogen release. METHODS: Chief cells were prepared by sequential digestion with collagenase and Ca2+ chelation. NO generation was measured by determining the NO coproduct citrulline. RESULTS: NO synthase immunoreactivities were constitutively expressed in approximately 70% chief cells. Carbachol (10 mumol/L) caused a 4- 6-fold increase in pepsinogen release, citrulline generation, intracellular Ca2+ concentration ([Ca2+]i) and cGMP concentration. These effects were concentration dependently inhibited by NG-monomethyl-L-arginine (L-NMMA). As gastrin, cholecystokinin, thapsigargin, and Ca2+ ionophore increased NO generation, [Ca2+]i seemed to regulate NO synthase activity. [Ca2+]i chelator and calmodulin antagonist inhibited the carbachol-induced pepsinogen secretion and NO generation. Preincubating the cells with L-NMMA had no effect on carbachol-stimulated inositol triphosphate generation or [Ca2+]i or Ca(2+)-dependent adenosine triphosphatase levels. Nitrovasodilator agents and 8-bromo-cGMP stimulated pepsinogen release. CONCLUSIONS: Gastric chief cells express a Ca2+/calmodulin-dependent NO synthase. NO modulates the stimulatory effect of Ca(2+)-mediated agonists on pepsinogen release.

Animals↗
PubMed1993

Occurrence and partition of the beta-carboline norharman in rat organs.

The beta-carboline norharman was determined in plasma, brain, liver, kidney, spleen, heart and lung of the rat using HPLC with fluorescence detection. In order to improve the speed and sensitivity of this assay an earlier published sample clean-up extraction procedure and HPLC method were adjusted. Norharman was found to be present in plasma as well as in all organs tested, concentrations in organs being about 80 times higher than those in plasma. Intraperitoneal injections of 2 and 100 mg/kg norharman showed that the partition of norharman between organs and plasma is about 3. Only the highest dose was found to have behavioural effects, viz. alerting reactions, a decrease in motor and exploratory activity, sedation, loss of righting reflex and after 30 min complete muscle relaxation, but no catatonia was observed. Norharman was found to be metabolized by the liver with a half live of about 20 min, whereas all other organs tested did not show any norharman clearing capacity. The results suggest that norharman is not likely the cause of psychosis, but a natural sedative and by- or coproduct of a more primary biochemical derangement.

Animals↗
PubMed1991

Citrulline in the rat brain: immunohistochemistry and coexistence with NADPH-diaphorase.

The presence in the brain of the urea cycle intermediate citrulline in the absence of a complete urea cycle has never been adequately explained. In an attempt to clarify this problem, we developed antibodies to citrulline and determined the distribution of citrulline-immunoreactivity in fixed sections of rat brain using immunoperoxidase and indirect immunofluorescence techniques. Citrulline-positive neurons were found to have a restricted distribution within the brain. A few cells were present in the cortex and corpus callosum. A large population of strongly stained cells was diffusely scattered throughout the striatum, nucleus accumbens and olfactory tubercle. Less strongly stained cells were detected in the supraoptic and paraventricular nuclei of the hypothalamus, the dorsal raphe, and the laterodorsal and pedunculopontine tegmental nuclei of the pons. The citrulline-immunoreactive cells were similar to those previously shown to contain NADPH-diaphorase activity, and double staining experiments indicated that citrulline-immunoreactivity was present in a subpopulation of NADPH-diaphorase-positive neurons. We have recently identified NADPH-diaphorase as a nitric oxide synthase. Thus the presence of citrulline in these cells suggests that it is formed within the brain as a coproduct during nitric oxide formation from arginine.

Amino Acid Oxidoreductases↗
PubMed1996

Epidermal growth factor modulates pepsinogen secretion in guinea pig gastric chief cells.

BACKGROUND & AIMS: Although epidermal growth factor (EGF) inhibits gastric acid secretion, the effects it exerts on gastric chief cells are unknown. The aim of this study was to investigate whether EGF modulates pepsinogen release and intracellular Ca2+ concentrations ([Ca2+]i) and whether the effect involves mitogen-activated protein (MAP) kinase, eicosanoid generation, and nitric oxide. METHODS: Chief cells were obtained by sequential digestion with collagenase and Ca2+ chelation. [Ca2+]i was measured in cells loaded with Fura-2 and NO generation by the NO coproduct citrulline. RESULTS: In situ hybridization, immunohistochemistry, and immunoblotting showed that EGF receptor and MAP kinases were constitutively expressed in chief cells. EGF caused a concentration-dependent stimulation of pepsinogen secretion and MAP kinase activity and determined a 2.5-7.0-fold increase in [Ca2+]i, inositol 1,4,5-tryphosphate, prostaglandin E2, and leukotriene B4. Tyrosine kinase inhibitors and cyclooxygenase and lipoxygenase inhibitors reduced pepsinogen secretion and eicosanoid generation induced by EGF. EGF increased citrulline generation and guanosine 3',5'-cyclic monophosphate accumulation sixfold; the effect was blocked by NG monomethyl-L-arginine, which is an NO synthase inhibitor. CONCLUSIONS: EGF stimulates pepsinogen secretion by activating eicosanoid generation, tyrosine kinases, MAP kinases, Ca2+, NO, and guanosine 3',5'-cyclic monophosphate.

4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyraz↗