Measurement of calcitonin and calcitonin gene-related peptide mRNA refines the management of patients with medullary thyroid cancer and may replace calcitonin-stimulation tests.

BACKGROUND: Serum calcitonin (sCT) is the main tumor marker for medullary thyroid cancer (MTC), but it has certain limitations. Various sCT assays may have important intra-assay or interassay variation and may yield different and sometimes conflicting results. A pentagastrin- or calcium-stimulation calcitonin (CT) test may be desirable in some situations. Alternatively, or in the absence of the stimulation test, mRNA detection offers the advantages of being more comfortable and less invasive; it only requires blood collection and has no side effects. The objective of this study was to investigate the applicability of measuring calcitonin-related polypeptide alpha (CALCA) gene transcripts (CT-CALCA and calcitonin gene-related peptide [CGRP]-CALCA) in patients with MTC and in relatives diagnosed with a RET mutation and to test mRNA as an alternative diagnostic tool for the calcitonin-stimulation test. METHODS: Twenty-three healthy controls and 26 individuals evaluated for MTC were selected, including patients with sporadic or hereditary MTC and RET mutation-carrying relatives. For molecular analysis, RNA was extracted from peripheral blood, followed by cDNA synthesis using 3.5 µg of total RNA. Quantitative real-time polymerase chain reaction (RT-qPCR) was performed with SYBR Green and 200 nM of each primer for the two specific mRNA targets (CT-CALCA or CGRP-CALCA) and normalized with the ribosomal protein S8 as the reference gene. RESULTS: We detected CALCA transcripts in the blood samples and observed a positive correlation between them (r=0.946, p<0.0001). Both mRNAs also correlated with sCT (CT-CALCA, r=0.713, p<0.0001; CGRP-CALCA, r=0.714, p<0.0001). The relative expression of CT-CALCA and CGRP-CALCA presented higher clinical sensitivity (86.67 and 100, respectively), specificity (97.06 and 97.06), positive predictive value (92.86 and 93.75), and negative predictive value (94.29 and 100), than did sCT (73.33, 82.35, 64.71, and 87.50, respectively). In addition, the CALCA transcript measurement mirrored the response to the pentagastrin test. CONCLUSION: We demonstrate that the measurement of CALCA gene transcripts in the bloodstream is feasible and may refine the management of patients with MTC and RET mutation-carrying relatives. We propose considering the application of this diagnostic tool as an alternative to the calcitonin-stimulation test.

Functional and histologic assessment of rat gastric mucosa after chronic treatment with sulphurous thermal water.

The effect of a chronic (4 weeks) administration of sulphurous thermal water on gastric acid secretion and mucosal defense was investigated in rats. Animals were randomized to receive daily intake of tap water or of thermal water obtained from a local spa center (Tabiano, Parma, Italy). Rats were followed for one month as for water and food consumption, body weight and general conditions. At the end of the watering period, the following study protocols were carried out: (a) study of basal and stimulated gastric acid secretion under general anesthesia, and (b) study of the gastric mucosal resistance against the damage induced by ethanol and indomethacin in conscious rats. Basal acid secretion and the acid response to pentagastrin or to histamine were similar in rats assuming ordinary drinking water or thermal water. As for resistance to gastric damage, histological, but not macroscopic, evaluation revealed that rats which assumed thermal water were slightly more resistant to the gastrolesive effect of ethanol (either absolute or diluted). Again, when indomethacin was used as a noxious stimulus, no difference was noted between the two groups as for macroscopic damage; only a nonsignificant reduction of damage was observed histologically in stomachs of rats assuming thermal water. In conclusion, these results indicate that chronic treatment of rats with thermal water, rich in sulphur compounds, may have only minimal effects on the rat gastric mucosa and did not significantly affect mucosal defense mechanisms. The observed tendency to gastroprotection would possibly need further investigation with longer periods of administration.

Characterization of gastrin-cholecystokinin 2 receptor interaction in relation to c-fos induction.

The interaction of gastrin with the cholecystokinin 2 (CCK2)/gastrin receptor has been studied extensively in relation to gastric acid secretion. However, not much is known about the contribution of individual amino acids of gastrin interacting with the CCK2 receptor, when gastrin is acting as a tumor growth factor. The purpose of the present study was to determine the significance of each individual amino acid residue of human gastrin-17 with respect to CCK2 receptor-mediated cell proliferation. Activation of this receptor was assessed using an in vitro bioassay based on gastrin-induced expression of a c-fos-luciferase reporter, transfected in AR42JB13 and Colo 320 cells, a rat pancreatic and human colorectal cell line respectively. Gastrin-17 dose dependently increased c-fos induction in both cancer cell lines. L365,260, a known CCK2 receptor antagonist, completely blocked the gastrin signal, demonstrating the specificity of this assay. We demonstrated for the first time that four carboxy-terminal amino acids of gastrin-17 are essential for activation of the CCK2 receptor with respect to c-fos induction. Also other residues of gastrin-17, notably glycine-2 for the rat CCK2 receptor and glutamic acid 8-10 and tyrosine-12 for the human receptor, were found to be important, although to a lesser extent. Alanine-substitution variants of each of the four carboxy-terminal amino acids of gastrin-17 showed strongly reduced receptor activation but did not act as competitive inhibitors of gastrin-17. Identification of the essential role of the carboxy-terminal tetrapeptide of gastrin-17 in CCK2 receptor-mediated c-fos induction indicates that gastrin inhibitory therapeutic strategies should mainly be targeted toward this region of gastrin.

Review article: human pepsins - their multiplicity, function and role in reflux disease.

Human gastric juice contains a multiplicity of proteinases. These are classified as aspartic proteinases because of enzymic activity dependent on two oppositely placed aspartic acids in the active site region. At least seven zones of activity can be visualized by agar gel electrophoresis and a similar number of separate proteins resolved by high performance ion exchange chromatography. The major enzyme secreted (up to 70% of the total) pepsin 3b is sensitive to the selective inhibitor pepstatin whereas gastricsin or pepsin 5 (20% of the total) is not. Minor enzymes including pepsin 1, which has an associated proteincarbohydrate complex attached is variable and can be < 5% in normal and up to 20% of the total as in peptic ulcer patients. The activity of these enzymes depends on the substrate and pH with significant digestion occurring up to pH 4.5. It has also been shown that these enzymes can bind to substrates like collagen up to pH 5.5. In gastric secretion studies of patients with reflux oesophagitis the amount of pepsin and the profile of the enzymes in basal secretions, and that after pentagastrin stimulation, was found to be not different from healthy non-refluxers. Thus the problem with reflux is that gastric juice appears in the oesophagus, an area without any natural protection from proteolytic damage. The ability to reduce gastric secretion is therefore important in effective treatment. However, being able also to inhibit enzymic activity or protect substrates from damage using alginates offers considerable scope for future therapies.

Effect of peripherally administered ghrelin on gastric emptying and acid secretion in the rat.

Ghrelin is a gut peptide that is secreted from the stomach and stimulates food intake. There are ghrelin receptors throughout the gut and intracerebroventricular ghrelin has been shown to increase gastric acid secretion. The aim of the present study was to examine the effects of peripherally administered ghrelin on gastric emptying of a non-nutrient and nutrient liquid, as well as, basal and pentagastrin-stimulated gastric acid secretion in awake rats. In addition, gastric contractility was studied in vitro. Rats equipped with a gastric fistula were subjected to an intravenous infusion of ghrelin (10-500 pmol kg(-1) min(-1)) during saline or pentagastrin (90 pmol kg(-1) min(-1)) infusion. After administration of polyethylene glycol (PEG) 4000 with 51Cr as radioactive marker, or a liquid nutrient with (51)Cr, gastric retention was measured after a 20-min infusion of ghrelin (500 pmol kg(-1) min(-1)). In vitro isometric contractions of segments of rat gastric fundus were studied (10(-9) to 10(-6) M). Ghrelin had no effect on basal acid secretion, but at 500 pmol kg(-1) min(-1) ghrelin significantly decreased pentagastrin-stimulated acid secretion. Ghrelin had no effect on gastric emptying of the nutrient liquid, but significantly increased gastric emptying of the non-nutrient liquid. Ghrelin contracted fundus muscle strips dose-dependently (pD2 of 6.93+/-0.7). Ghrelin IV decreased plasma orexin A concentrations and increased plasma somatostatin concentrations. Plasma gastrin concentrations were unchanged during ghrelin infusion. Thus, ghrelin seems to not only effect food intake but also gastric motor and secretory function indicating a multifunctional role for ghrelin in energy homeostasis.

Necessity of intracellular cyclic AMP in inducing gastric acid secretion via muscarinic M3 and cholecystokinin2 receptors on parietal cells in isolated mouse stomach.

The existence of a direct action of acetylcholine and gastrin on muscarinic M3 and cholecystokinin2 (CCK2) receptors on gastric parietal cells has not yet been convincingly established because these stimulated acid secretions are remarkably inhibited by histamine H2 receptor antagonists. In the present study, we investigated the necessity of intracellular cyclic AMP in inducing gastric acid secretion via muscarinic M3 and CCK2 receptors on parietal cells using an isolated mouse stomach preparation. Bethanechol (10-300 microM) produced a marked increase in acid output and this increase was completely blocked by famotidine (10 microM). In the presence of famotidine, bethanechol (1-30 microM) augmented the acid secretory response to dibutyryl AMP (200 microM) in a concentration-dependent manner. The augmentation was blocked by atropine (1 microM), 4-DAMP (0.1 microM), a muscarinic M3-selective antagonist, and by Ca2+ exclusion from the serosal nutrient solution. Pentagastrin (0.3-3 microM) also concentration-dependently stimulated gastric acid secretion, but the effect was completely inhibited by famotidine. In the presence of famotidine, pentagastrin (0.1-0.3 microM) elicited a definite potentiation of the acid secretory response to dibutyryl cyclic AMP (200 microM). This potentiation was inhibited by YM022 (1 microM), a CCK2 receptor antagonist, and by exclusion of Ca2+ from the serosal nutrient solution. The present results suggest that gastric acid secretion via the activation of muscarinic M3 and CCK2 receptors on the parietal cells is induced by activation of the cyclic AMP-dependent secretory pathway.

G protein-coupled receptor desensitization as a measure of signaling: modeling of arrestin recruitment to activated CCK-B receptors.

Gastrin is one of the principle hormonal mediators of gastric acid secretion, and its cognate receptor (CCK-B) is a member of the superfamily of GPCRs. Patients with hypergastrinemia may present with a variety of symptoms, including gastric ulcers or malignant tumors. Thus, the molecular mechanisms that terminate CCK-B receptor signaling, as well as an ability to measure gastrin bioactivity in a timely manner, have important clinical implications. In order to assess CCK-B receptor regulation, we have constructed a single cell biosensor containing the CCK-B receptor and an arrestin/GFP chimera. The gastrin biosensor responded to both immunologically detectable gastrin-17 and undetectable pentagastrin, and was able to determine the gastrin bioactivity of serum from a patient with clinical hypergastrinemia. We determined that the CCK-B receptor binds arrestin with a pharmacology mirroring CCK-B receptor signaling through inositol phosphate, and that the rate of arrestin dissociation from internalized receptor mirrors receptor recycling to the plasma membrane. Moreover, the CCK-B recycling rate is intermediate between that of Class A GPCRs such as the beta2-adrenergic receptor and Class B GPCRs such as the vasopressin type 2 receptor. Mathematical modeling of these results indicates that a common receptor conformation may underlie both CCK-B signaling and desensitization. In addition to its use in drug screening, this methodology should generalize to other receptors for use in diagnosis and monitoring of bioactive ligands involved in GPCR-based disease.

A defined human gastrin sequence stimulates the growth of Helicobacter pylori.

This study describes the interaction between gastrin and Helicobacter pylori. Human gastrin amino acids 4-17 were found to be the minimal growth-stimulating sequence. Gastrin from other mammals did not stimulate bacterial growth. When human serum was used to stimulate bacterial growth in brucella broth, gastrin was shown to be a necessary and sufficient growth-stimulating factor. Competition for the gastrin effect by pentagastrin and cholecystokinin (CCK-8) resulted in inhibition of bacterial growth. This effect was mediated by the four C-terminal amino acids which are shared by gastrin, CCK-8 and pentagastrin. In conclusion, the interaction between gastrin and H. pylori was shown to be specific, essential, and dependent on a defined gastrin sequence.

Very early detection of RET proto-oncogene mutation is crucial for preventive thyroidectomy in multiple endocrine neoplasia type 2 children: presence of C-cell malignant disease in asymptomatic carriers.

BACKGROUND: Multiple endocrine neoplasia type 2 (MEN 2) is an inherited disease caused by germline mutations in the RET proto-oncogene, and is responsible for the development of endocrine neoplasia. Its prognosis is dependent on the appearance and spread of medullary thyroid carcinoma (MTC). Relatives at risk can be identified before clinical or biochemical signs of the disease become evident. METHODS: Twenty-one families with MEN 2 (16 families with MEN 2A and 5 families with MEN 2B) were studied. Peripheral blood DNA was amplified by polymerase chain reaction. DNA sequence or restriction enzyme analysis was performed to detect mutations of RET proto-oncogene exons 10, 11, and 16. Molecular analysis was carried out in all index patients as well as in 98 relatives of MEN 2A patients (60 juveniles, ages 6 months to 21 years, and 38 adults, ages 22 to 81 years) and in 13 relatives (6 juveniles ages 10 to 21 years, and 7 adults ages 41 to 66 years) from MEN 2B families. RESULTS: Molecular studies showed a mutation at codon 634, exon 11 in all MEN 2A patients. All MEN 2B patients showed an ATG to ACG (Met918Thr) mutation. In MEN 2A families, 42 out of 98 relatives were affected. Total thyroidectomy was performed in 18 juvenile carriers ages 17 months to 21 years. Histopathologic studies of the glands revealed parafollicular cell (C-cell) hyperplasia in all of these carriers, medullary thyroid carcinoma in 15 carriers, and only one carrier with lymph node metastases. CONCLUSIONS: The consistent finding of C-cell disease in all the juvenile carriers who underwent preventive thyroidectomy emphasizes the relevance of early screening in children at risk of developing MTC. The presence of MTC in the specimen of prophylactic thyroidectomy from a 17 month old girl highlights the importance of thyroidectomy as soon as the molecular diagnosis is confirmed.

Effects of orally administered enteric-coated omeprazole on gastric acid secretion in horses.

OBJECTIVES: To determine the effects of orally administered omeprazole, as enteric-coated capsules, on baseline and stimulated gastric acid secretion in horses. ANIMALS: 5 healthy 8-year-old mixed-breed horses fitted with gastric cannulas. PROCEDURE: Enteric-coated granules of omeprazole were mixed with corn syrup and administered orally once daily for 5 consecutive days. On days 1 and 5 beginning 5 hours after omeprazole administration, 4 gastric fluid samples were collected, each for 15 minutes, via the gastric cannula (baseline samples). Pentagastrin was administered IV as a constant infusion for the subsequent 2 hours, and 15-minute gastric fluid samples were again collected (stimulated samples). Fluid volume, acidity (mmol H-/L), and pH and gastric acid production (mmol H+) were determined for all baseline samples and for stimulated samples collected during the second hour of pentagastrin infusion. Control experiments were done in a similar manner after giving corn syrup alone to the same horses. RESULTS: Compared with values obtained during control experiments, baseline and stimulated gastric fluid acidity and gastric acid production significantly decreased, and the mean pH of gastric fluid samples significantly increased, after horses were given 5 daily doses of omeprazole. CONCLUSIONS AND CLINICAL RELEVANCE: Enteric-coated omeprazole (1.0 mg/kg of body weight; PO) administered once daily for 5 days significantly inhibited unstimulated and pentagastrin-stimulated gastric acid secretion in horses. This commercially available formulation of omeprazole may be efficacious in the treatment of gastroduodenal ulcers in horses.

Full and partial agonist activity of C-terminal cholecystokinin peptides at the cloned human CCK-A receptor expressed in Chinese hamster ovary cells.

The agonist activities of the C-terminal cholecystokinin peptides sulfated cholecystokinin octapeptide (CCK-8S), non-sulfated cholecystokinin octapeptide (CCK-8NS), pentagastrin and CCK-4 at the cloned human CCK-A receptor expressed in Chinese hamster ovary cells were evaluated in two functional assays of receptor activation. [125I]-CCK-8S displacement studies employing membranes derived from these cells revealed the expected rank order of affinity for a number of CCK receptor ligands. CCK-8S was a potent agonist in (i) stimulating the mobilization of intracellular free Ca2+, measured with the Ca2+ sensitive fluorescent indicator FURA-2, and (ii) stimulating increases in extracellular acidification rates, measured by microphysiometry. Consistent with their lower affinities for CCK-A receptors, CCK-8NS, pentagastrin and CCK-4 were weaker agonists in both functional assays. In addition, these peptides exhibited partial agonist activity relative to the maximum response observed with CCK-8S in both assays. These results demonstrate that CCK-8S represents the minimum ligand requirement for both high affinity and full agonist activity at the human CCK-A receptor subtype.

Gastrin-CCK-B type receptors on human T lymphoblastoid Jurkat cells.

The presence of specific receptors for gastrointestinal hormones on T cells and their involvement in the immune response are still matters of debate. We reported the effects of gastrin-cholecystokinin (CCK)-related peptides on J.RT3-T3.5 Jurkat cells. A single class of high-affinity binding sites (dissociation constant approximately 0.1 nM) for gastrin and CCK-8 was evident on these cells. These peptides dose-dependently induced a transient increase in intracellular Ca2+ concentration ([Ca2+]i), which was independent of extracellular Ca2-. L-365,260 was 150- to 300-fold more potent than L-364,718 to inhibit radiolabeled ligand binding or peptide-stimulated [Ca2+]i increase, confirming the gastrin-CCK-B nature of the receptor. Gastrin caused a rise in inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] level within 5 s of stimulation. Finally, gastrin increased interleukin (IL)-2 secretion in J.RT3-T3.5 cells. We conclude that 1) J.RT3-T3.5 cells possess "gastrin-CCK-B type" receptors coupled to phospholipase C activation, Ins(1,4,5)P3 formation, and Ca2+ release from intracellular Ca2+ pools, and 2) these receptors could be involved in the regulation of IL-2 production.

Hormonal regulation of adenylate cyclase activity in circulating lymphocytes and its interrelationship with hormone sensitivity of tumor tissue in colorectal cancer patients.

The purpose of this study was to investigate the peculiarities of hormonal regulation of adenylate cyclase (AC) of blood lymphocytes in colorectal cancer patients and to compare these peculiarities with hormone sensitivity of AC of colorectal tumors and normal colonic mucosa. Basal and stimulated lymphocyte AC activity was studied in 51 healthy persons and 52 cancer patients (14 with colon cancer, 21 with rectal cancer and 17 with stomach cancer) aged 20-75 years. In 31 of 35 patients with colorectal cancer the AC activity was studied simultaneously in lymphocytes, tumor tissue and normal colonic mucosa. To evaluate basal and stimulated AC activity the measurement of c-AMP (Amersham kits) formed in the presence of ATP regenerating system was used. Basal and by VIP, pentagastrin and sodium fluoride stimulated AC activity in lymphocytes of gastrointestinal cancer patients was lower than in lymphocytes of healthy subjects of similar age. Stage dependence of the parameters under study was not found. There was a tendency for higher basal and stimulated lymphocyte AC activity in colon cancer patients as compared to stomach and rectal cancer patients. In colorectal cancer patients the peculiarities of lymphocyte AC reactions to stimulation were closer to those in tumor tissue but not to those in normal colonic mucosa. The reaction of lymphocyte AC to VIP and glucagon coincided more frequently with tumor AC reactions to the same hormones in case of hormone nonsensitive tumors. Thus, basal and stimulated lymphocyte AC activity in colorectal cancer patients was modified to some degree by tumor factors. Lymphocyte AC reactions to VIP and glucagon may be considered as indirect markers of hormone sensitivity of colonic tumors. Moreover, the probability of discovery of hormone nonsensitive tumors by this way is more reliable than hormone sensitive ones.

[The observance of Ramadan and its repercussion on gastric secretion]. / L'observance du Ramadan et son retentissement sur la sécrétion gastrique.

The influence of the Ramadan on the gastric secretion has been assessed in 13 volunteers. Their basal and pentagastrin-stimulated secretion has been collected before, during and one month after the Ramadan. Gastric activity, pepsin activity, sialic acid bound to glycoprotein, choline and gastrinaemia have been measured. During Ramadan, acid secretion was increased (+ 159%; P = 0.02) and it recovered the pre-Ramadan level, one month later. Pepsin secretion was also increased during the Ramadan (+ 133%; P = 0.05) and it was significantly reduced after Ramadan. The secretion before the Ramadan was related to vagal hypertony, decreased during Ramadan and was substituted by a gastrinic stimulation after the Ramadan. Mucolysis and gastrinaemia were not modified and no duodenogastric reflux was observed during and after the Ramadan. The Ramadan induces an increase of acid and pepsin secretion. This increase was reversed when Ramadan stopped. These gastric secretion modifications are likely involved in the increase of dyspeptic symptoms observed during the Ramadan.

Combined dose-ratio analysis of cholecystokinin receptor antagonists, devazepide, lorglumide and loxiglumide in the guinea-pig gall bladder.

1. Interactions between cholecystokinin octapeptide (CCK-8) and CCKA-receptor antagonists derived from benzodiazepines (devazepide) and glutamic acid (lorglumide and loxiglumide) have been examined in an improved bioassay using the guinea-pig, isolated, gall bladder preparation. 2. The presence of CCKB-receptors in the assay was provisionally-ruled out on the basis of the low potency of pentagastrin in the assay. By applying analyses of both agonism and antagonism, pentagastrin was shown to behave as a partial agonist at the CCKA-receptor. 3. Devazepide, lorglumide and loxiglumide behaved as simple competitive antagonists of CCKA-receptors and pKB values of 9.98, 7.59 and 7.07 were estimated, respectively. 4. Application of a combined dose-ratio analysis to the interactions between CCK-8 and combinations of devazepide/lorglumide and devazepide/loxiglumide indicated that these molecules behave as syntopic, competitive, antagonists at the CCKA-receptor. 5. We conclude that the guinea-pig gall bladder assay contains a homogeneous population of CCKA-receptors and offer an explanation for the differences between our results and those obtained recently by Maubach et al. (1991) which were taken as preliminary evidence for CCKA-receptor heterogeneity.

Evaluation of sensitive PDN-21 (katacalcin) determination as tumor marker in medullary thyroid carcinoma.

PDN-21 (katacalcin), a peptide from the calcitonin (CT) gene, was measured in plasma from healthy persons and patients with medullary thyroid carcinoma (MTC). PDN-21 was detectable (greater than or equal to 10 ng/l) in 73% of normal persons (n = 40). In 17 normal persons with undetectable basal plasma levels, PDN-21 became detectable (greater than or equal to 10 ng/l) by stimulation with iv pentagastrin in 7 cases. Basal levels were more often detectable in men than in women. In 65 patients with MTC, PDN-21 levels were highly correlated with CT levels as determined by an "in house" RIA (r = 0.99); the mean ratio of CT/PDN-21, on a molar base, was 0.96 +/- 0.33 over the entire range. In iv stimulation tests with pentagastrin, PDN-21 and CT showed good parallelism (mean ratio of CT/PDN-21: 1.1 +/- 0.62); in MTC patients with normal basal levels, however, peak to basal ratios during iv pentagastrin testing were higher for PDN-21 than for CT, due to the more sensitive PDN-21 assay. In a selective venous catheter study of a patient with MTC, the mean CT/PDN-21 ratio for all samples was 1.04 +/- 0.12, but the peak to peripheral levels were higher for PDN-21 (4.1-fold) than for CT (2.8-fold). In conclusion, determination of PDN-21 by RIA is equivalent to determination of CT in diagnosing MTC patients. In few patients, it might be even slightly more sensitive. PDN-21 should be determined in all cases with borderline CT results.

[The participation of CCK-8-ergic mechanisms in the regulation of emotional behavior in rodents]. / Uchastie KhTsK-8-ergicheskikh mekhanizmov v reguliatsii émotsional'nogo povedeniia u gryzunov.

Effects of CCK-8 receptor agonists caerulein and pentagastrin and CCK-8 receptor antagonist proglumide on exploratory and locomotor activity of mice and rats were studied. Systemic administration of caerulein (500 ng/kg 1 mcg/kg) decreased significantly the exploratory activity of mice in elevated plus-maze. This anxiogenic-like action of caerulein was attenuated by acute pretreatment with proglumide (1 and 15 mg/kg) but not with diazepam (up to 0.75 mg/kg). Proglumide slightly increased the exploratory activity of rats in plus-maze; on the other hand, caerulein and pentagastrin potently decreased the measures of exploration in this test. Caerulein (10-100 mcg/kg) and proglumide (1 and 15 mg/kg) inhibited 3H-pentagastrin binding in mice brain in in vivo experiments. The data obtained indicate that CCK-8-ergic mechanisms in brain play an important role in the generation of anxiety states in rodents.

A new potent and selective non-peptide gastrin antagonist and brain cholecystokinin receptor (CCK-B) ligand: L-365,260.

L-365,260 (3R(+)-N-(2,3-dihydro-1-methyl-2-oxo-5-phenyl-1H-1,4- benzodiazepin-3-yl)-N'-(3-methylphenyl)urea), interacted in a stereoselective and competitive manner with guinea pig stomach gastrin and brain cholecystokinin (CCK) receptors. The affinity of L-365,260 for both gastrin (Ki = 1.9 nM) and brain CCK-B (Ki = 2.0 nM) receptors was greater than 2 orders of magnitude higher than its affinity for peripheral pancreatic CCK-A receptors or various other receptors. L-365,260 exhibited a similar high affinity for brain CCK-B receptors of rats, mice and man. A somewhat lower affinity for gastrin and brain CCK-B (IC50 = 20-40 nM) receptors was observed in dog tissues. In vivo, oral administration of L-365,260 antagonized gastrin-stimulated acid secretion in mice (ED50 = 0.03 mg/kg), rats (ED50 = 0.9 mg/kg) and guinea pigs (ED50 = 5.1 mg/kg). L-365,260 did not affect basal acid secretion and did not antagonize histamine- or carbachol-stimulated acid secretion in mice. L-365,260 represents a potentially powerful new tool for investigating gastrin and brain CCK-B receptors, and possibly their role in physiology and disease.

Plasma somatostatin activity in medullary cancer of the thyroid.

In a group consisting of 50 adults and 5 children with diagnosed or suspected medullary cancer of the thyroid, plasma somatostatin levels were measured in 77 samples with a sensitive radioimmunoassay. Only 2 patients had clearly enhanced plasma somatostatin immunoreactivity, both with highly active aggressive tumors. Other patients had plasma somatostatin levels of under 20 ng/l or undetectable ones. No evidence of response of somatostatin levels to pentagastrin stimulation was found.

Role of peptide structure in lipid-peptide interactions: nuclear magnetic resonance study of the interaction of pentagastrin and [Arg4]pentagastrin with dimyristoylphosphatidylcholine.

Complexes formed between dimyristoylphosphatidylcholine (DMPC) and the peptide pentagastrin or [Arg4]pentagastrin were examined by 31P- and 2H-NMR. The cationic [Arg4]pentagastrin produces larger changes in the lipid NMR spectra than does the anionic pentagastrin. 31P-NMR spectra of DMPC with [Arg4]pentagastrin below the phase transition exhibits two components one of which is motionally restricted compared with the pure lipid. The exchange between these two lipid domains is slow on the millisecond time scale. The interactions between this peptide and phospholipid are diminished above the melting temperature of the complex. The 2H-NMR spectra of DMPC which had been labelled in a choline methylene group is also affected more by the [Arg4]pentagastrin than by pentagastrin. In the presence of [Arg4]pentagastrin, even above the lipid phase transition, an additional doublet with a smaller quadrupole splitting is observed. These results clearly demonstrate the importance of peptide charge in determining the effects of peptides on lipid bilayers.