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Background/aims: Type 2 diabetes mellitus (T2DM) is correlated with gallbladder diseases. This study aimed to investigate the expression of CCK and IP3 receptors in patients with gallbladder stones and T2DM and its correlation with the hypomotility of the gallbladder., Methodology: 26 patients with gallstones and T2DM (Group 1) and 24 gallstones patients without T2DM (Group 2) were enrolled in this study. The emptying function of the gallbladder was measured by ultrasonography. The activity of CCK-R was analyzed by radioligand method and the IP3-R antibody was used to detect the IP3-R from patients in both groups., Results: Gallbladder ejection volume (EV) ((11.6±5.1) ml3 vs (21.5±7.8) ml3) and gallbladder ejection fraction (GBEF2)(%)((17.2±11.3) ml3 vs (52.8±12.9) ml3) were significantly lower (P<0.01) in patients with gallstones and T2DM. The amount of CCK-R and the activity of CCK-R in Group 1 were significantly lower than that in Group 2 (P<0.01). And IP3-R in Group 1 was much lower than that in Group 2, as well (P<0.01)., Conclusion: The expression of CCK-R and IP3-R in gallstones patients with T2DM was much lower in such patients, leading to impaired gallbladder emptying function and the formation of gallstones.

Two structurally-related guanine nucleotide-binding protein-coupled receptors for two related peptides, cholecystokinin (CCK) and gastrin, have evolved to exhibit substantial diversity in specificity of ligand recognition, in their molecular basis of binding these ligands, and in their mechanisms of biochemical and cellular regulation. Consistent with this, the CCK1 and CCK2 receptors also play unique and distinct roles in physiology and pathophysiology. The paradigms for ligand recognition and receptor regulation and function are reviewed in this article, and should be broadly applicable to many members of this remarkable receptor superfamily. This degree of specialization is instructive and provides an encouraging basis for the diversity of potential drugs targeting these receptors and their actions that can be developed.

Background: The etiology of chronic acalculous gallbladder dysfunction (CAGD) is unknown. However, cholecystectomy is being performed as treatment, based on gallbladder (GB) ejection fraction studies. The aim of this study was to examine the pathology and immunohistology of GBs from children with CAGD., Methods: Children with a diagnosis of CAGD were identified. Control patients had their GB removed for nonbiliary indications. Immunoperoxidase staining was performed using rabbit antihuman cholecystokinin receptor (CCK-R) antibody. The pathologist was blinded to the study and controls., Results: Fifteen children were evaluated: 6 children with CAGD and 9 controls. All children with CAGD had abnormal cholecystokinin-stimulated nuclear imaging. Ejection fractions ranged from 8% to 30%. All patients reported resolution of symptoms on follow-up at 6 months. Histopathology of the GB was normal for both the controls and children with CAGD. Both control and CAGD GBs demonstrated positive staining for CCK-R in the vascular endothelium and smooth muscle. Mucosal epithelial staining was only observed in 5 of 6 of GBs of children with CAGD. In the sixth GB, the epithelium was too necrotic to assess., Conclusions: In this pilot study, expression of CCK-Rs in the epithelial cells is noted in children with CAGD compared with controls. The significance of this finding requires further investigation.

Background & Aims: The localization and functional role of cholecystokinin (CCK) receptor proteins in normal and diseased human pancreas, particularly in ductal pancreatic carcinomas, remain unclear., Methods: Tissue samples of normal human pancreas, chronic pancreatitis, and ductal pancreatic carcinomas were investigated under carefully controlled conditions for expression of CCK1 and CCK2 receptor messenger RNA (mRNA) and proteins using in situ hybridization and in vitro CCK receptor autoradiography by means of subtype-selective analogues. Synaptophysin immunohistochemistry was used concomitantly for optimal identification of islets, nerves, and tumor areas with neuroendocrine features., Results: CCK2 receptor mRNA and proteins were found abundantly in human pancreatic islets in normal pancreas and chronic pancreatitis. CCK1 receptor proteins were found occasionally in small-sized pancreatic nerves, whereas acini expressed a low density of CCK2 receptors in a few cases of chronic pancreatitis. Ductal pancreatic carcinomas rarely expressed CCK receptors; a few receptor-positive tumors, often characterized by neuroendocrine differentiation, expressed the CCK2 receptor at the mRNA or protein level. However, the main source of CCK receptors in the pancreatic tumor samples consisted of CCK2-expressing islets and/or CCK1-expressing nerves rather than neoplastic tissue., Conclusions: These data indicate that the presence of CCK receptors in human ductal pancreatic tumor samples is mainly due to CCK2 expression in residual pancreatic islets and CCK1 in pancreatic nerves. Pancreatic acini and ductal pancreatic tumor cells very rarely express CCK2 receptors. These observations suggest that CCK analogues may not be of clinical use to target most of these cancers.

Objective: To assess the effect of cholesterol in bile on cholecystokinin receptor (CCK-R) in the gallbladder., Methods: One hundred Guinea pigs were randomly divided into four groups, 25 animals for each. The control group was fed a standard diet, and the cholesterol group fed a diet containing 2% cholesterol. After taking the 2% cholesterol diet for two weeks, the natural group persisted on the standard diet, and the treated group was perfused by traditional Chinese medicine. Serum cholecystokinin (CCK) level in the portal vein and maximal binding capacity (B(max)) and Kd of CCK-R in the gallbladder were measured in the four groups by RIA and RBA, and the concentrations of cholesterol in bile were also observed., Results: Compared with the control group, after high-cholesterol feeding for two weeks, the gallbladder emptying rate [(65.83 +/- 7.32)% approximately (47.22 +/- 5.24)%] and B(max) of CCK-R [(60 +/- 27) approximately (32 +/- 13) fmol/mg protein] and in decreased fasting gallbladder volume (FV) [(0.89 +/- 0.26) approximately (1.34 +/- 0.61) cm(3)] and concentration of cholesterol [(0.44 +/- 0.11) approximately (0.60 +/- 0.13) mmol/L] in bile increased, but no change was in the serum CCK level and Kd of CCK-R in the cholesterol group. Compared with the natural group, after two-week in take of herb decoction of qingre lidan and liqi huoxue, FV [(1.27 +/- 0.60) approximately (0.90 +/- 0.27) cm(3)], RV [(0.85 +/- 0.45) approximately (0.32 +/- 0.12) cm(3)], FB [(0.92 +/- 0.35) approximately (0.73 +/- 0.21) cm(3)], RB [(0.76 +/- 0.34) approximately (0.29 +/- 0.08) cm(3)] in the treated group decreased significantly; but gallbladder emptying rate [(43.06 +/- 4.27)% approximately (67.01 +/- 6.82)%] increased significantly. The concentration of cholesterol in bile was lower in the treated group than in the natural group [(0.59 +/- 0.14) approximately (0.43 +/- 0.10) mmol/L], but no change was found in the serum CCK level. Bmax of CCK-R in the treated group increased significantly [(39 +/- 19) approximately (59 +/- 11) fmol/mg protein], Kd of CCK-R showed no significant changes between the treated group and natural group., Conclusion: High cholesterol in gallbladder bile causes defective muscle contraction by down-regulating CCK-R in the gallbladder, so the reduction of cholesterol concentration of bile may contribute to gallbladder contraction.

Malignant astrocytic tumors are characterized by the pronounced and diffuse migration of tumor astrocytes into the brain parenchyma. The present study shows that gastrin is a brain neuropeptide that is able to significantly modulate astrocytic tumor migration at both invasion and motility levels. In the matter of invasion, gastrin severely reduces the in vitro invasive abilities of C6 rat glioma, 9L rat gliosarcoma, and U373 human glioma cells in a collagen matrix. In vitro, gastrin also markedly modifies the motility features in both C6 and U373 cells, at least partly through a decrease in the expression of the RhoA small GTPase, and so brings about some dramatic modifications to the organization in the actin cytoskeleton. The in vitro preincubation of C6 tumor cells with gastrin significantly increases the life spans of rats stereotactically implanted with these cells as compared with the survival periods of rats implanted with gastrin-untreated C6 cells. As suggested by our in vitro experiments, these effects, observed in vivo cannot relate to only the gastrin-induced decrease in tumor astrocyte migratory abilities. Indeed, gastrin also induces immunomodulatory effects, because we observed a marked gastrin-induced recruitment of lymphocytes into C6 gliomas and 9L gliosarcomas. These data all suggest that gastrin can act as an endogenous modulator of glioma progression.

The high sensitivity of the pentagastrin stimulation test in detecting primary or metastatic medullary thyroid cancer (MTC) suggests a widespread expression of the corresponding receptor type on human MTC. Indeed, autoradiographic studies demonstrated cholecystokinin (CCK)-B/gastrin receptors not only in more than 90% of MTCs, but also in a high percentage of small-cell lung cancers, stromal ovarian tumors, and potentially a variety of other tumors, including gastrointestinal adenocarcinomas, neuroendocrine tumors, and malignant glioma. The aim of our work was to develop and systematically optimize suitable radioligands for targeting CCK-B receptors in vivo and to investigate their role in the staging and therapy of MTC and other CCK-B receptor expressing malignancies. For this purpose, a variety of CCK/gastrin-related peptides, all having in common the C-terminal CCK-receptor binding tetrapeptide sequence-Trp-Met-Asp-PheNH(2) or derivatives thereof, were investigated. They were members of the gastrin or cholecystokinin families or possessed characteristics of both, which differ by the intramolecular position of a tyrosyl moiety. Their stability and affinity were studied and optimized in vitro and in vivo; their biodistribution and therapeutic efficacy were tested in preclinical models. Best tumor uptake and tumor to nontumor ratios were obtained with members of the gastrin family, because of their superior selectivity and affinity for the CCK-B receptor subtype. Radiometal-labeled derivates of minigastrin showed excellent targeting of CCK-B receptor expressing tissues in animals and healthy human volunteers. Preclinical therapy experiments in MTC-bearing animals showed significant antitumor efficacy. In a subsequent clinical study, 45 MTC patients with metastatic MTC were investigated; 23 had known and 22 had occult disease. CCK-B receptor scintigraphy was performed with (111)In-diethylenetriamine pentaacetic acid-d-Glu(1)-minigastrin. The normal organ uptake was essentially confined to the stomach (and, to a lesser extent, to the gallbladder and, in premenopausal women, to normal breast tissue) as a result of CCK-B receptor specific binding and to the kidneys, as excretory organs. All tumor manifestations known from conventional imaging were visualized as early as 1 hour postinjection, with increasing tumor to background ratios over time; at least 1 lesion was detected in 20 of 22 patients with occult disease (patient-based sensitivity, 91%). Among them were local recurrences and lymph node, pulmonary, hepatic, splenic, and bone (marrow) metastases. Eight patients with advanced metastatic disease were injected in a dose-escalation study with potentially therapeutic activities of a (90)Y-labeled minigastrin derivative at 4 to 6-week intervals (30-50 mCi/m(2) per injection for a maximum of 4 injections). Hematologic and renal toxicities were identified as the dose-limiting toxicities at the 40 and 50 mCi/m(2) levels. Two patients experienced partial remissions, and 4 experienced stabilization of their previously rapidly progressing disease. These data suggest that CCK-B receptor ligands may be a useful new class of receptor-binding peptides for diagnosis and therapy of a variety of (CCK-B receptor expressing) tumor types. They allow for sensitive and reliable staging of patients with metastatic MTC. Initial therapeutic results are promising, but nephrotoxicity is a major concern to be solved., (Copyright 2002, Elsevier Science.)

Regulatory peptides and their analogs are being extensively investigated as radiopharmaceuticals for cancer imaging. In particular, cholecystokinin (CCK) receptors of the subtype B (CCK-BR) have been shown to be overexpressed in certain neuroendocrine tumors including medullary thyroid cancer. Our recent work has focused on new methods to radiolabel the CCK8 peptide with (111)In or (99m)Tc for the purpose of developing radiopharmaceuticals for in vivo CCK-B receptor imaging. Labeling of CCK8 with (111)In was achieved at the N-terminus of the peptide by adding, in solid phase, a glutamate coupled diethylenetriaminepentaacetic acid (DTPA) moiety through a glycine linker, yielding DTPA-Glu-G-CCK8. For labeling with (99m)Tc, the CCK8 peptide was modified at its N-terminus by introducing, in the following order--cysteine, glycine, and a diphenylphosphinopropionyl moiety--giving a 10-residue peptide derivative, Phos-GC-CCK8. A cell culture model was developed for the purpose of evaluating the binding properties of these two ligands. The human epidermoid carcinoma cell line, A431, was transfected with a plasmid containing the full coding sequence of the human CCK-BR under a strong viral promoter, obtaining a number of receptors in the range of 2-5 x 10(6) per cell. Control cells were transfected with vector alone. An animal tumor model utilizing these two cell lines was developed to evaluate the specificity of interaction with the CCK-BR and biodistribution properties of the compounds. CCK-BR positive and control cells were subcutaneously injected in opposite flanks of CD1 female nude mice in order to obtain xenografts differing only in their ability to express CCK-B receptors. High performance liquid chromatography (HPLC) and other chromatographic methods were utilized to assess stability of the radiolabeled compounds after injection. Both (111)In-DTPA-Glu-G-CCK8 and (99m)Tc-Phos-GC-CCK8 showed similar binding affinities for cultured CCK-BR expressing cells, with dissociation constants in the range of 20-40 nM. With the two xenograft approach, we were able to demonstrate specific interaction with the receptor of both CCK analogs in our animal model. The data obtained shows rapid specific localization of both compounds on the CCK-BR overexpressing xenografts. Both tracers show rapid plasma clearance of unbound peptide. Clearance of (111)In-DTPA-Glu-G-CCK8 appears to be preferentially through the kidneys, whereas (99m)Tc-Phos-GC-CCK8 clearance occurs both through kidneys and the hepatobiliary system. Both our labeling approaches appear adequate for clinical use of peptide based radiopharmaceuticals, although (99m)Tc-Phos-GC-CCK8 shows elevated accumulation in the gastrointestinal tract, which causes high background activity., (Copyright 2003 Wiley Periodicals, Inc.)

Gastrin17gly acts as a growth factor for the colonic mucosa. Studies of the receptor involved have generally been restricted to its binding properties, and no investigation of the structure of gastrin17gly receptors on human colorectal carcinoma cell lines has yet been reported. The aim of this study was to optimise the conditions for binding of gastrin17gly to the human colorectal carcinoma cell line DLD-1, and to investigate the structure of the receptor responsible. Binding of 125I[Met15]gastrin17gly to DLD-1 cells was measured in competition experiments with increasing concentrations of either gastrin17gly or gastrin17, or with single concentrations of gastrin receptor antagonists. The molecular weights of the gastrin17gly binding proteins were determined by gel electrophoresis and autoradiography after covalent cross-linking of 125I[Nle15]gastrin2,17gly to cells or membranes with disuccinimidyl suberate. The IC50 value for binding of gastrin17gly to DLD-1 cells was 2.1+/-0.4 microM. Binding was inhibited by the non-selective gastrin/cholecystokinin receptor antagonists proglumide and benzotript, but not by the cholecystokinin-A receptor antagonist L364,718, or the gastrin/cholecystokinin-B receptor antagonist L365,260. The molecular weight of the major gastrin binding protein on DLD-1 cells or membranes was 70,000. We conclude that the major gastrin17gly binding site on the human colorectal carcinoma cell line DLD-1 is clearly distinct from the cholecystokinin-A and gastrin/cholecystokinin-B receptors, but is similar in some respects to the gastrin/cholecystokinin-C receptor.

Background: The interrelationship between histamine and gastrin in the physiological regulation of gastric acid secretion is still a matter of dispute. CCK-2 receptors are located on enterochromaffin-like (ECL) cells in corpus mucosa and gastrin stimulates acid production by releasing histamine from the ECL cells, which in turn stimulates the parietal cells. Whether parietal cells also possess gastrin receptors of physiological significance is unclear. The aim of the present study was to localize the CCK-2 receptor cellularly and concomitantly demonstrate a gastrin receptor response (histamine release)., Methods: Fluorescein labelled cholecystokinin-8 (Fluo-CCK-8) was added to the arterial infusion to totally isolated, vascularly perfused rat stomachs to a final concentration of 130 pmol L(-1) for 1 min, either alone or along with 520 nmol(-1) CCK-8 after 10-min pre-perfusion with CCK-8. Immediately after the Fluo-CCK-8 had reached the oxyntic mucosa, biopsies were taken and the binding sites were localized by double immunohistochemistry combined with the tyramide signal amplification (TSA) technique. Venous histamine was measured before and during stimulation., Results: Fluo-CCK-8 (130 pM) evoked histamine release, and binding sites were found in the basal part of corpus mucosa, co-localized with histidine decarbocylase (HDC) immunoreactive ECL cells. No binding of Fluo-CCK was found in the mid-glandular region of corpus, dominated by parietal cells. Binding of Fluo-CCK-8 was abolished by concomitant perfusion with excess CCK-8., Conclusion: Fluo-CCK-8 given to isolated rat stomachs in a physiological concentration binds to CCK-2 receptors on ECL cells and causes histamine release, whereas no binding of Fluo-CCK-8 to parietal cells was found.

The objective of this study was to examine the effects of 10% food restriction on body weight, plasma cholecystokinin (CCK) levels, and exocrine pancreatic function in male Sprague-Dawley rats. A matched group of rats with unrestricted access to food served as controls. After ingesting the diets for 32 da, the rats were killed and blood obtained for plasma cholecystokinin, glucose, and insulin determinations. To evaluate pancreatic function, the pancreases were removed, weighed, and digested with collagenase to isolate pancreatic acini, which were incubated with maximal stimulating dose of CCK. The fraction of amylase that was released into the medium was measured. To explore the role of membrane receptors in exocrine pancreatic secretion, CCK receptor affinity and CCK receptor capacity were determined by radioligand binding assays in isolated, purified membranes from pancreatic acini. Compared to the control group, rats with 10% food restriction showed (a) reduced body weight gain, (b) increased pancreatic weight, (c) increased plasma CCK level, and (d) no significant changes in plasma glucose or insulin levels. The food-restricted group showed a reduction of pancreatic function, assessed by measuring amylase release in response to maximal CCK stimulation; the amylase release was diminished by 35% in the food-restricted group. In isolated acinar cell membranes from food-restricted rats, CCK receptor affinity and capacity were reduced by 23% and 16%, respectively, compared to controls. These results indicate that consumption of less food than normal affects pancreatic function by a mechanism that evidently involves CCK release and downregulation of CCK receptors. The data suggest that CCK plays an important physiological role in the adaptation to eating less food, and thereby to the lowering of body weight in rats and, possibly, in other animals.

One of the physiological functions of circulating cholecystokinin (CCK) is in the control of the pyloric sphincter and the subsequent delivery of nutrients to the small intestine. In order to identify the site(s) of action of CCK in the gastropyloric region, we performed immunohistochemistry using an antibody directed to the C-terminal region of the cholecystokinin A receptor (CCKAR). In the rat, cells that display strong CCKAR immunoreactivity and fit the morphological description of interstitial cells of Cajal (ICC) were found in the distal sphincter muscle and in the circular muscle of the proximal duodenum. Double labeling showed that these cells coexpressed vimentin, but that not all vimentin-positive cells expressed CCKAR. Confirmation that the CCKAR-expressing cells were ICC also came from kit double-labeling experiments in mice. In addition to ICC, circular smooth muscle cells at the tip of the comma-shaped sphincter muscle, but not elsewhere, also exhibited strong, membrane-bound CCKAR immunoreactivity. With higher antibody concentrations, the entire circular muscle displayed moderate CCKAR immunoreactivity, suggesting that circular smooth muscle cells express low levels of CCKAR. Select neurons in the myenteric ganglia near the sphincter muscle proper, the distal antrum, and proximal duodenum, as well as a few single neurons in the submucosa, also expressed strong CCKAR immunoreactivity. Finally, CCKAR-immunoreactive ICC and neurons were not specifically related to vagal afferent intramuscular and intraganglionic endings, and vagal afferents themselves did not exhibit any CCKAR immunoreactivity. These results suggest a role for ICC and enteric neurons in the mediation of CCK effects on pyloric sphincter pressure in addition to direct effects of the hormone on circular smooth muscle.

Background: It has been assumed that gastrin stimulates the growth of pancreatic cancer in an autocrine way through co-expression of gastrin and the cholecystokinin-B receptor (CCK-BR). However, pancreatic cancer cell lines established directly from patients have revealed a great heterogeneity in cell proliferation when exposed to CCK, gastrin and their receptor antagonists. The aim of this study was therefore to examine co-expression of CCK-A and CCK-B receptor (CCK-AR and CCK-BR), and gastrin mRNA as well as the secretion of CCK and gastrin peptides in these cell lines., Methods: Fourteen cell lines were established from primary pancreatic cancers or their metastases. Total RNA was isolated from the cell lines and reverse-transcribed into single-stranded cDNA. A PCR technique based on Taq polymerase-antibody interaction and CCK-AR, CCK-BR and gastrin-specific primers, followed by Southern blot analysis, were the methods used. The incubation mediums were analysed for the presence of secreted CCK/proCCK and gastrin/progastrin peptides by specific radioimmunoassays (RIA)., Results: By means of nested Reverse-Transcribed Polymerase Chain Reaction (nested RT-PCR), combined with Southem blot analysis of the PCR amplified products, CCK-AR and gastrin mRNA co-expression was detected in cell lines LPC-6p and LPC-10m, whereas CCK-BR and gastrin mRNA could be detected in cell lines LPC-8p and LPC-12m. A low level of secreted CCK peptides was detected in cell line LPC-6p, which also expressed CCK-AR mRNA. In no other cases were CCK or gastrin peptides detected in the cell culture mediums., Conclusion: The lack of CCK-BR and gastrin mRNA co-expression, and not detectable levels of secreted CCK and gastrin in culture media, does not lend support to the hypothesis that concomitant gene-expression of CCK receptors and gastrin or CCK are essential to maintaining pancreatic cancer cell proliferation.

Regulatory peptides are small, readily diffusable and potent natural substances with a wide spectrum of receptor-mediated actions in humans. High affinity receptors for these peptides are (over-) expressed in many neoplasms, and these receptors may represent, therefore, new molecular targets for cancer diagnosis and therapy. This review aims to give an overview of the peptide-based radiopharmaceuticals which are presently already commercially available or which are in advanced stages of their clinical testing so that their broader availability is anticipated soon. Physiologically, these peptides bind to and act through G protein-coupled receptors in the cell membrane. Historically, somatostatin analogs are the first class of receptor binding peptides having gained clinical application. 111In-DTPA-[D-Phe1]-octreotide is the first and only radiopeptide which has obtained regulatory approval in Europe and the United States to date. Extensive clinical studies involving several thousands of patients have shown that the major clinical application of somatostatin receptor scintigraphy is the detection and the staging of gastroenteropancreatic neuroendocrine tumors (carcinoids). In these tumors, octreotide scintigraphy is superior to any other staging method. However, its sensitivity and accuracy in other, more frequent neoplasms is limited. Radiolabeled vasoactive intestinal peptide (VIP) has been shown to visualize the majority of gastrointestinal adenocarcinomas, as well as some neuroendocrine tumors, including insulinomas (the latter being often missed by somatostatin receptor scintigraphy). Due to the outstanding diagnostic accuracy of the pentagastrin test in detecting the presence, persistence, or recurrence of medullary thyroid cancer (MTC), we postulated the expression of the corresponding (ie. cholecystokinin [CCK-] -B) receptor type in human MTC. This receptor is also widely expressed on human small-cell lung cancer. Indeed, 111In-labeled DTPA derivatives of gastrin showed excellent targeting of CCK-B receptor expressing tissues in animals and patients. A variety of further peptide-based radioligands, e.g. among many others, gastrin-releasing peptide/bombesin, neurotensin, substance-P, pan-somatostatin (somatostatin derivatives which bind to all five receptor subtypes) or glucagon-like peptide-1 (glp-1) analogs (the latter for the specific detection of insulinomas), is currently under development. Summarizing, radiolabeled regulatory peptides have opened new horizons in nuclear oncology for diagnosis (and potential internal radionuclide therapy). Future work will probably reveal a multitude of novel potentially clinically useful peptide-based radioligands.

In vitro cholecystokinin (CCK) contracts the human lower oesophageal sphincter by stimulating muscular receptors. The aim of this study was to characterize the muscular CCK receptor subtypes in the human lower oesophageal sphincter. Twenty-five circular strips from six patients were studied. RNA was extracted, reverse transcribed, and cDNAs were amplified with primers for human CCK-A and B receptors. The potency of the contraction induced by CCK-8, desulphated CCK-8, and gastrin-I, and the effect of the CCK-A (loxiglumide and SR 27897) and the CCK-B (YM022 and L-365 260) specific receptor antagonists were compared. Both CCK-A and CCK-B receptor mRNAs were found in functional lower oesophageal sphincter strips. The potency of the CCK-8 concentration-dependent contraction was two and three orders of magnitude higher than that of desulphated CCK-8 and gastrin-I, respectively. The CCK-8-induced contraction was blocked by the CCK-A receptor antagonists loxiglumide (IC50 11 micromol L-1) and SR 27897 (IC50 74 nmol L-1) but not by CCK-B receptor antagonists (1 micromol L-1). Our data suggest that, although the human lower oesophageal sphincter expresses both CCK-A and CCK-B receptors, the contractile effect of CCK-8 on the circular muscle is mainly due to the activation of CCK-A receptors.

Unlabelled: Cholecystokinin (CCK)-B receptors have been demonstrated on a high percentage of medullary thyroid carcinomas (MTC) in vitro. After encouraging results both in vitro and in animal studies, we studied the efficacy of an octapeptide [111In-DTPA]-CCK analogue in seven patients with MTC. In four of five patients in whom serum calcitonin levels were monitored, a significant rise was found following the injection, indicating retained biological activity of the radiopeptide. In all patients there was visualization of the CCK-B receptor-positive stomach. In one of two patients with known MTC lesions, some of the lesions were visualized; in addition some lesions were visualized in one of the five other patients who had elevated serum tumour markers but negative localizing studies. Radioactivity in the presumed tumour sites was still present at 48 h p.i. The uptake in the presumed tumour sites and stomach was low. Background radioactivity dropped rapidly owing to urinary excretion. After 1 h, breakdown products of the labelled analogue predominated both in urine and in serum, and virtually no intact peptide was present., In Conclusion: (1) the CCK-B receptor-positive gastric mucosa and presumed MTC lesions could be visualized in patients using an octapeptide [111In-DTPA]-CCK analogue that is probably internalized, proving the feasibility of CCK-B receptor imaging in vivo; (2) there was a relatively low uptake of the CCK analogue in the strongly CCK receptor positive stomach, and rapid degradation of the peptide in serum.

Background: To evaluate the function of cholecystokinin B (CCKB)/gastrin receptors in the rat kidney, we identified the receptors by Northern blot and localized the receptors by immunohistochemistry. The functional effects of gastrin were studied under standardized in vitro conditions using the isolated perfused kidney., Methods: Rat kidneys were mounted in an organ bath by attaching the renal artery to a perfusion system. A catheter was inserted into the renal vein and the ureter to collect samples that were analyzed for the concentrations of electrolytes. After a preperfusion period, gastrin-17-I was given via the renal artery (10-8 to 10-6 mol/L). Subsequently, hemodynamic parameters (for example, perfusate flow) and changes in sodium and potassium absorption were determined. All data were subjected to a nonparametric analysis of variance and, in case of significant results, to subsequent paired comparisons by the a posteriori Wilcoxon test., Results: Northern blot analysis detected CCKB receptor transcripts in total RNA isolated from kidneys. Immunohistochemistry localized CCKB receptors on tubules and collecting duct cells. Compared with controls, gastrin (10-6 mol/L) caused a decrease in the fractional sodium reabsorption (basal 80%, 10 minutes after application of gastrin 71%, after 20 minutes 62%, P < 0.05). This effect was inhibited by the CCKB receptor antagonist L-365,260. Gastrin decreased urinary potassium excretion at 10-8 and 10-6 mol/L [maximal decrease at 10-6 mol/L from baseline values (100%) to 49% after 10 minutes and to 69% after 20 minutes, P < 0.05, N = 6]. This effect was also abolished by the CCKB receptor antagonist L-365,260. Gastrin (10-6 mol/L) reduced perfusate flow by 31% (P < 0.05)., Conclusions: CCKB receptors are expressed in the rat kidney on tubules and collecting ducts. These receptors mediate changes in renal potassium and sodium absorption. In addition, gastrin causes a decrease in perfusate flow, indicating that CCKB receptors might also modulate vascular resistance in the kidney.

In rodents, cholecystokinin (CCK) induces pancreatic enzyme secretion and pancreas growth through its CCK(A) receptors. It is unknown whether occupation of the CCK(B) receptors present in pig and human pancreas can cause the same effects. This study evaluates CCK(B) receptor expression in rat, mouse, pig, and fetal human pancreata using Northern blot, Western blot, and immunofluorescence techniques. The reported 2.7-kb CCK(B) receptor mRNA transcript in the rat brain and gastric fundus is absent in pancreas; the message was, however, detected by RT-PCR and by a CCK(B) receptor antibody as an 80-kDa protein present uniquely in islet delta-cells. Proteins of 50 and 80 kDa appear in mouse pancreas, and proteins of 50 and 115 kDa appear in pig and human pancreas, respectively, all localized in islet delta-cells. Gastrin mRNAs are strongly present in fetal rat pancreas, and the hormone is localized in islets; both are repressed 10 days after birth. In conclusion, the CCK(B) receptors are present in pancreas of four species with exclusive location in islet delta-cells. In such a location, they could be indirectly involved in the control of enzyme secretion.

The CCK-A (cholecystokinin-A) receptor is selectively expressed by human pancreatic adenocarcinomas, suggesting a possible role in pancreatic tumorigenesis. In animals, pancreatic CCK receptor expression varies during ontogeny and neoplastic transformation. This study examined the temporal expression of CCK receptors in human fetal, postnatal, and adult pancreas to determine whether the appearance of CCK-A receptors in pancreatic adenocarcinomas reflected oncofetal antigen or pancreatic neoantigen expression. Messenger ribonucleic acid (mRNA) was isolated from six paraffin-embedded normal pancreatic autopsy specimens ranging in age from 17 weeks postfertilization through 26 days following full-term delivery, and samples of adult human tissues, including pancreas and pancreatic adenocarcinoma. Using reverse transcription-polymerase chain reactions, CCK-B receptor mRNA was expressed in all specimens of normal fetal and postnatal human pancreas, adult pancreas, and pancreatic adenocarcinomas. CCK-A receptor mRNA was selectively expressed only in pancreatic adenocarcinomas. These data suggest that selective CCK-A receptor expression in pancreatic adenocarcinomas reflects neoantigen expression in humans.

Gastrin stimulates gastric acid secretion by acting on the cholecystokinin B/gastrin receptor (CCK-BR). The localization of this receptor at the cellular level showed conflicting results in animal studies and has not been described in man by immunohistochemistry. The aim of the present study is to characterize the precise cellular location of the CCK-BR in the human stomach. Polyclonal antisera were raised against different epitopes of the CCK-BR molecule and used for immunohistochemical investigations. CCK-BR mRNA was detected in paraffin tissue sections by the highly sensitive method of in situ reverse transcriptase-polymerase chain reaction (RT-PCR). Using immunohistochemistry, CCK-BR could successfully be localized in gastric parietal cells. In the majority of parietal cells, CCK-BR immunoreactivity was present a he basolateral cell membrane domain. In some parietal cells, a granular pattern of immunoreactivity was exclusively confined to the cytoplasm of the cells. CCK-BR mRNA was found in parietal cells and in enterochromaffin-like (ECL) cells by means of in situ RT-PCR. No expression of CCK-BR was found in the gastric antral mucosa. Our data support the concept that gastrin stimulates gastric acid secretion directly via CCK-B receptors on parietal cells and indirectly by inducing histamine release from histamine-containing ECL cells, which contributes to acid secretion by parietal cells.

Unlabelled: The presence of cholecystokinin (CCK)-B (gastrin) receptors has been shown in more than 90% of medullary thyroid cancers (MTCs) and in a high percentage of small cell lung cancers, stromal ovarium cancers and several other tumor types., Methods: The aim of this study was to evaluate in vitro and in vivo whether 111In-labeled CCK-B receptor-specific CCK8 analog [D-Asp26,Nle28,31]CCK26-33 (D-Asp-Tyr-Nle-Gly-Trp-Nle-Asp-Phe-NH2) is suitable for CCK-B receptor scintigraphy based on the finding that unlabeled nonsulfated diethylenetriamine pentaacidic acid [DTPA0]CCK8 and tetraazacyclododecanetetraacetic acid [DOTA0]CCK8 analogs show high and specific binding for CCK-B receptors in human tumors. Fifty percent inhibitory concentrations were in the low nanomolar range., Results: In vitro, [111In-DOTA0]CCK8 showed specific internalization in CCK-B receptor-positive rat pancreatic tumor cells AR42J. Internalization of the analog appeared to be time and temperature dependent and receptor specific. From the data obtained with [111In-DOTA0]CCK8 and (125I)I-gastrin, the latter being a specific ligand for the CCK-B receptor, the rat pancreatic cell line CA20948 also appeared to be CCK-B receptor positive. This provides an in vitro and in vivo rat tumor model because this cell line can be grown to solid tumors in Lewis rats. In vivo biodistribution experiments in CA20948 tumor-bearing Lewis rats showed rapid clearance of [111In-DOTA0]CCK8, and specific uptake was found in the CCK-B receptor-expressing stomach and tumor. Furthermore, comparing [111In-DOTA0]CCK8 with the radioiodinated nonsulfated CCK10 analog (D-Tyr-Gly-Asp-Tyr-Nle-Gly-Trp-Nle-Asp-Phe-NH2), both ligands having high affinity for the CCK-B receptor, tumor-to-blood ratios were significantly higher for [111In-DOTA0]CCK8 than for 125I-CCK10, analogous to the findings with radioiodinated and 111In-labeled octreotide. The study in humans with [111In-DTPA0]CCK8 showed receptor-specific uptake in the CCK-B receptor-positive stomach and in metastases in the neck region up to 48 h after injection., Conclusion: [111In-DOTA0]CCK8 is most promising for scintigraphy and, after coupling to therapeutic radionuclides, for radionuclide therapy of human CCK-B receptor-positive tumors such as MTC and small cell lung cancer.

Background: Overexpression of regulatory peptide receptors in selected human tumours is of diagnostic and therapeutic relevance., Aims: To evaluate the expression of somatostatin, vasoactive intestinal peptide (VIP), substance P, cholecystokinin (CCK) A and B, and neurotensin receptors in hepatocellular carcinoma (HCC)., Methods: In vitro receptor autoradiography for the various peptide receptors using selective iodinated radioligands on tissue sections in 59 cases of HCC., Results: 41% of HCC expressed somatostatin receptors; 47% expressed VIP receptors. VIP receptors were always identified in non-neoplastic liver tissue. Substance P receptors were only identified in 5% of HCC but in the majority of their peritumorous and intratumorous vessels. CCK-A and -B and neurotensin receptors were not detected in HCC. The somatostatin receptors showed high affinity for somatostatin and octreotide. The VIP receptors had high affinity for VIP, pituitary adenylate cyclase activating peptide (PACAP) 27, and a VIP1 selective analogue, suggesting the presence of VIP1/PACAP II type receptors. PACAP I receptors were identified in two cases. Substance P receptors were all of the NK1 subtype. The density of somatostatin receptors in HCC was low compared with the density found in liver metastases of neuroendocrine tumours. The VIP receptor density was always lower in HCC than in adjacent liver tissue., Conclusions: Somatostatin, VIP, and substance P may have a receptor mediated role in HCC. Substance P receptors may be involved in regulation of tumour associated blood flow; somatostatin receptors and VIP receptors may mediate tumour growth. Diagnostic and therapeutic evaluation of somatostatin and VIP analogues may be of interest in receptor positive HCC.

The high sensitivity of pentagastrin stimulation in detecting primary or metastatic medullary thyroid cancer (MTC) suggests widespread expression of the corresponding receptor type on human MTC. Indeed, autoradiographic studies demonstrated cholecystokinin (CCK)-B/gastrin receptors not only in >90% of MTCs but in a high percentage of small cell lung cancers and potentially a variety of gastrointestinal adenocarcinomas. In a pilot study, we have demonstrated the feasibility of radiolabeled gastrin-I to target CCK-B receptor-expressing tissues in vivo in animals and patients (T. M. Behr et al., Eur. J. Nucl. Med., 25: 424-430, 1998). The aim of the present study was to systematically optimize, in a preclinical model, suitable radioligands for targeting CCK-B receptors in vivo. For this purpose, a variety of CCK/gastrin-related peptides, all having in common the COOH-terminal CCK-receptor binding tetrapeptide sequence Trp-Met-Asp-PheNH2 or derivatives thereof, were studied. They were radioiodinated by the Iodogen or Bolton-Hunter procedures. The peptides tested were members of the gastrin- or cholecystokinin families or possessed characteristics of both, which differ by the intramolecular position of a tyrosyl moiety (occurring in native or sulfated form). Their stability and affinity were studied in vitro and in vivo; their biodistribution and therapeutic efficacy were tested in nude mice bearing s.c. human MTC xenografts. Diethylene-triamine-pentaacetate derivatives of suitable peptides were synthesized, evaluated, and labeled with (111)In. All members of the CCK or gastrin family were stable in serum (with t(1/2)s of several hours at 37 degrees C); nevertheless, the stability of those peptides was highest that bore the NH2-terminal pGlu residues (e.g., big gastrin, gastrin-I, caerulein, and others) or D-amino acids. In accordance to their comparably low affinity, nonsulfated members of the CCK family showed fairly low uptake in the tumor and other CCK-B receptor-expressing tissues (e.g., the stomach). Sulfated CCK derivatives performed significantly better but additionally displayed a high uptake in normal, CCK-A receptor-expressing tissues (such as the liver/gallbladder, pancreas, and bowel). Best tumor uptake and tumor:nontumor ratios were obtained with members of the gastrin family, probably because of their selectivity and affinity for the CCK-B receptor subtype. Pilot therapy experiments in MTC bearing animals showed significant antitumor efficacy as compared with untreated controls. (111)In-Labeled diethylene-triamine-pentaacetate derivatives of minigastrin showed excellent targeting of CCK-B receptor-expressing tissues in animals and a normal human volunteer. These data suggest that CCK/gastrin analogues may be a useful new class of receptor binding peptides for diagnosis and therapy of CCK-B receptor-expressing tumors, such as MTC or small cell lung cancer. Nonsulfated gastrin derivatives may be preferable because of their CCK-B receptor selectivity, and hence, lower accretion in normal CCK-A receptor-expressing organs. Further preclinical as well as clinical studies are ongoing.

We have evaluated the presence and localization of the CCK(A) receptor in rat, mouse, pig and human fetal pancreas by Northern, Western blots and immunofluorescence techniques. In the rat, parallelism exists between development of the CCK(A) receptor mRNA and protein with maximal peaks of expression during the suckling period. In the course of pancreatitis induction, CCK(A) receptor mRNA were maximally expressed and sustained during the gland's regeneration. In the rat and mouse pancreas, the CCK(A) receptor protein is localized around the acinar cells and beta cells of the islets of Langerhans. In the adult pig and fetal human pancreas, the CCK(A) receptor proteins were detected by Western blot. By immunofluorescence, its detection was possible only in the islet of Langerhans of the pig pancreas. These new findings support the views that CCK plays important and various roles in specific physiological systems of the pancreas of different species., (Copyright 1999 Academic Press.)

Objective: To characterise the cholecystokinin (CCK) receptor subtypes in medullary thyroid cancer by measuring the expression of CCK-A and CCK-B/gastrin receptor mRNA., Design: Open study., Setting: Teaching hospital, Sweden., Subjects: 6 patients with medullary thyroid cancer., Intervention: Pentagastrin stimulation test and measurement of calcitonin concentration. Biopsy specimens were analysed using reverse transcription polymerase chain reaction (RT-PCR)., Main Outcome Measure: Presence of CCK-A and CCK-B/gastrin receptors., Results: All 6 patients with medullary thyroid cancer had positive pentagastrin tests preoperatively. CCK-B/gastrin receptors but not CCK-A receptors were detected by RT-PCR in all six biopsy specimens. By contrast, no CCK receptors were found in normal thyroid tissues or in other thyroid tumours (follicular adenoma, papillary carcinoma, or anaplastic carcinoma)., Conclusion: The presence of CCK-B/gastrin receptors in medullary thyroid tumours may have important clinical implications.

Unlabelled: The high sensitivity of pentagastrin stimulation in detecting primary or metastatic medullary thyroid cancer (MTC) suggests widespread expression of the corresponding receptor type on human MTC. Indeed, autoradiographic studies have demonstrated cholecystokinin (CCK)-B/gastrin receptors not only in more than 90% of MTCs but also in a high percentage of small cell lung cancers, some ovarian cancers, astrocytomas and potentially a variety of adenocarcinomas. The aim of this study was to systematically screen and optimize, in a preclinical model and a pilot clinical study, suitable radioligands for targeting CCK-B receptors in vivo., Methods: A variety of CCK/gastrin-related peptides, all bearing the C-terminal CCK receptor-binding tetrapeptide sequence Trp-Met-Asp-PheNH2 or derivatives thereof, were studied. They were radioiodinated by the lodogen or Bolton-Hunter procedures. The peptides were members of the gastrin or CCK families, which differ by the intramolecular position of a tyrosyl moiety. Their stability and affinity were studied in vitro and in vivo; their biodistribution and therapeutic efficacy were tested in nude mice bearing subcutaneous human MTC xenografts. Diethylenetriamine pentaacetic acid (DTPA) derivatives of suitable peptides were synthesized successfully, and their preclinical and initial clinical evaluations were performed, labeled with 111In., Results: All members of the CCK or gastrin families were stable in serum (with half-lives of several hours at 37 degrees C); nevertheless, the stability of those peptides bearing N-terminal pGlu residues or D-amino acids was significantly higher. In accordance with their comparably low affinity, nonsulfated members of the CCK family showed fairly low uptake in the tumor and other CCK-B receptor-expressing tissues. Sulfated CCK derivatives performed significantly better but also displayed a comparably high uptake in normal CCK-A receptor-expressing tissues. This effect was probably due to their similar affinity for both CCK-A and CCK-B receptors. Best tumor uptake and tumor-to-nontumor ratios were obtained with members of the gastrin family because of their selectivity and affinity for the CCK-B receptor subtype. Pilot therapy experiments in MTC-bearing animals showed significant antitumor efficacy compared with untreated controls. DTPA derivatives of minigastrin were successfully developed. In a pilot clinical study, radioiodinated and 111In-labeled derivatives showed excellent targeting of physiological CCK-B receptor-expressing organs, as well as all known tumor sites., Conclusion: CCK/gastrin analogs may be a useful new class of receptor-binding peptides for diagnosis and therapy of CCK-B receptor-expressing tumors, such as MTC or small cell lung cancer. Nonsulfated gastrin derivatives may be preferable because of their CCK-B receptor selectivity, hence lower accretion in normal CCK-A receptor-expressing organs.

Receptors for cholecystokinin (CCK), gastrin, neurotensin, somatostatin and vasoactive intestinal peptide (VIP) are over-expressed in several human tumors, where they have diagnostic and therapeutic implications. Since reports on the expression of these peptide receptors in primary gastric and colonic adenocarcinomas are either non-existent or conflicting, a detailed evaluation with particular emphasis on the tissue localization was undertaken. CCK-A, CCK-B, neurotensin, somatostatin and VIP receptors were localized by in vitro receptor autoradiography with iodinated radioligands on histological sections of surgical samples of 27 gastric and 25 colonic adenocarcinomas. CCK-A, CCK-B and neurotensin-1 receptors were found in a minority of both tumor types. Somatostatin receptors were found in 18/27 gastric and 2/25 colonic cancers. VIP receptors were found in 14/26 gastric and 23/25 colonic cancers; subtype characterization suggests VIP1 receptors. In addition, resected tumor samples contained non-malignant tissues (mucosa, smooth muscle, nerves or vessels) with high amounts of the various peptide receptors. Therefore, regulatory peptide receptors are expressed differentially in gastric and colonic cancers but also very frequently in "contaminating" non-malignant tissues. Since results using morphological techniques are superior to those using homogenates, we recommend that localization of these receptors to the tissues should always be attempted, to minimize receptor over-estimation in tumors and to prevent spurious results.

Gastrin and cholecystokinin (CCK) are two major regulatory peptides synthesized by human gut and brain tissues as well as by selected tumors, in particular gastrin-producing neuroendocrine tumors. In the present study we have evaluated gastrin and CCK gene expression in a group of primary human tumors, including neuronal, renal, and myogenic stem cell tumors, using in situ hybridization techniques. In addition, CCK-A and CCK-B receptors were evaluated in the same group of tumors with receptor autoradiography. Most tumors had gastrin messenger ribonucleic acid (mRNA): 10 of 11 medulloblastomas, 5 of 5 central primitive neuroectodermal tumors, 11 of 11 Ewing sarcomas, 8 of 10 neuroblastomas, 4 of 4 Wilms' tumors, 5 of 5 rhabdomyosarcomas, and 10 of 10 leiomyosarcomas. CCK mRNA was restricted predominantly to Ewing sarcomas (9 of 11) and leiomyosarcomas (5 of 10). CCK-A and CCK-B receptors were not frequently found in these tumors, except for leiomyosarcomas. These data suggest that gastrin and CCK may play a previously unrecognized role in this group of human stem cell tumors. If the increased gastrin mRNA indeed translates into increased gastrin production, measurement of gastrinemia may have a diagnostic significance in the early detection of these tumors. As these two hormones have been reported to act as potent growth factors, they may be of pathophysiological relevance for patients with such stem cell tumors.

Exocrine pancreatic cancer is significantly more common in younger men than in younger women. The male-to-female sex ratio is, in most countries, between 1.25 and 1.75 to 1, but decreases with increasing age. Moreover, prior oophorectomy appeared in one study to be significantly more common in women with pancreatic cancer than in controls. This has raised interest in sex hormones in the development in pancreatic cancer. It has been questioned if there are estrogen receptors in ductal pancreatic cancer, but there are no doubt estrogen receptors and estrogen-binding protein in human healthy pancreas. It is also well proven that it is possible to influence experimental pancreatic cancer with estrogens. However, in clinical studies tamoxifen has repeatedly been shown to be without significant effects. On the other hand, there are also androgen receptors in pancreatic cancer and testosterone has been shown to strongly promote growth in experimental pancreatic cancers. It is therefore of considerable interest that an antiandrogen recently was shown to significantly prolong life in patients with unresectable pancreatic carcinoma. However, in patients with advanced pancreatic carcinoma the S-testosterone is low, far lower than what could be expected due to weight-loss and malnourishment alone. Pancreatic cancer has etiologically been connected to diet, for example the intake of fat. Cholecystokinin receptors have been found on human pancreatic cancer, possible to stimulate in vitro by cholecystokinin (CCK). Studies with CCK-receptor binding, hybridization with radiolabeled complementary DNA (cDNA) probes, or reverse-transcription polymerase chain reaction have shown that CCK-A receptors also are present in rat pancreatic putative preneoplastic lesions and cancer tissue, rat pancreatic-cancer cell lines, pancreatic carcinomas in transgenic mice, hamster pancreatic cancer, and human pancreatic cancer cell lines and tumors. Also, CCK-B receptors have been found in some human pancreatic cancers. There are a vast number of experiments done on CCK-stimulation of pancreatic cancer. They indicate that CCK may have a promotional effect on exocrine pancreatic cancer, but it is not probable that hyperstimulation with CCK alone induce pancreatic cancer. At present, however, despite a lot of evidence for a hormone-dependence of pancreatic cancer there are no data confirming a role for estrogens, androgens, CCK or their antagonists in clinical treatment of exocrine pancreatic cancer.

Anti-serum raised against the human cholecystokinin B (CCKB)/gastrin receptor was used in Western blotting to differentiate the cellular locations of receptor isoforms expressed by human gastro-intestinal (GI) tumour cell lines. Using anti-serum directed against the amino-terminal extracellular tail of the CCKB/gastrin receptor, 8/9 cell lines were shown to express immunoreactive proteins of either m.w. 70 or 40 kDa, or both. Both isoforms were found to be associated with intracellular, non-nuclear membranes, whereas only the 70 kDa protein was expressed in the plasma membrane. Receptor expression was related to gastrin production and secretion. Both progastrin and glycine-extended gastrin-17 were produced and secreted by the tumour cell lines; however, carboxy amidated gastrin was not detected by radioimmunoassay. A CCKB/gastrin receptor transfectant NIH3T3 cell line did not produce detectable gastrin and showed exclusive expression of the 70 kDa receptor on the plasma membrane. One cell line had <50 pg/ml cell-associated progastrin and no detectable receptor form. Cell lines expressing 50-150 pg/ml had both 40 and 70 kDa receptor forms. Those expressing >150 pg/ml progastrin had only the 40 kDa isoform, which was shown to be exclusively expressed on intracellular, non-nuclear membranes, in one of the cell lines. Of the 4 cell lines exclusively expressing the lower m.w. receptor, 3 had gastrin present within the cell, which was not secreted. Thus, if cell-associated gastrin induces a proliferative effect, it may be by an intracrine pathway. Our study has identified the presence of CCKB/gastrin receptor isoforms in different cellular locations and may help toward understanding the complex autocrine and intracrine pathways mediated by gastrin peptides.

Background: Gastrin/cholecystokinin type B receptors (CCKBRs) can be found on parietal cells and smooth muscle cells and are the predominant brain CCK receptors. Recent cloning studies indicate that this is receptor type might also be expressed in the kidney., Materials and Methods: We used Northern blot analysis in guinea pig. kidney and reverse transcriptase polymerase chain reaction (RT-PCR) in several murine kidney cells lines to evaluate this organ for the expression of the CCKBRs. The receptor was pharmacologically characterized by displacement experiments using [125I]-BH-CCKs and various agonists and antagonists. Polyclonal antibodies vs. the CCKBRs were raised in chicken, and immunohistochemistry on tissue sections was used to localize the receptor within the organ. The effect of gastrin on renal cell growth was measured using proximal tubulus (MCT) cells, which were cultured with gastrin (10-9 M) for 24-72 h. Cell counts and [3H]-thymidine incorporation experiments were performed., Results: CCKBR transcripts can be detected in kidney RNA (tubules > glomeruli > interstitium). RT-PCR revealed CCKBR transcripts in proximal tubules (MCT cells) and in mesangium (MMC). The medullary thick ascending limb of Henle's loop and several control tissues such as liver and muscle were negative. Displacement experiments using [125I]-BH-CCK and various agonists and antagonists identified binding sites with typical CCKBR pharmacology. CCKBRs were localized in the proximal tubulus, distal collecting ducts and mesangium cells. Treatment of rested MCT cells with gastrin 17-1 induced cell proliferation and [3H]-thymidine incorporation by at least 40% compared with normal growth (P < 0.05)., Conclusion: These results show for the first time that CCKBRs are present in selected areas of the kidney, and strongly confirm our previous observation that this organ expresses binding sites for [125I]-gastrin. Furthermore, gastrin might act as a growth factor in the kidney.

Receptors for regulatory peptides such as somatostatin or vasoactive intestinal polypeptide are expressed by a number of human neoplasms and can be visualized in vivo with peptide receptor scintigraphy. Recently, the CCK-B receptor, which binds both gastrin and cholecystokinin with high affinity, was shown using in vitro methods to be overexpressed in a number of human tumor tissues, including medullary thyroid carcinomas, small cell lung cancers, astrocytomas, gastrointestinal tumors, and stromal ovarian cancers. In the present study, we have designed novel, unsulfated CCK octapeptide analogs linked to the metal chelating DTPA and DOTA, and have tested them for their binding affinity to CCK-B receptor-positive tissue from human tumors: The most potent compounds assayed were DTPA-[Nle28, 31]-CCK(26-33) (MP2286) and DTPA-[d-Asp26,Nle28,31]-CCK(26-33) (MP2288) with an IC50 of 1.5 nM. For comparison, analogs with C-terminal DTPA, such as [Nle28,31,Aphe33(p-NH-DTPA)]-CCK(26-33) and CCK-(26-33)-NH(CH2)2 NH-DTPA, had an IC50 of >100 nM. DOTA-[D-Asp26, Nle28,31]-CCK(26-33) had an IC50 of 3.9 nM. The compounds were selective for CCK-B receptors as they did not bind with high affinity to CCK-A receptors expressed in human tumors (meningiomas or gastroenteropancreatic tumors). In vivo rat biodistribution studies with indium-111 labeled MP2286 and MP2288 showed that the primary mode of clearance was renal, and the primary sites of uptake (% ID/g 24 h p.i.) were kidneys (0.270 and 0.262, respectively) and the gastrointestinal tract. The CCK-B receptor-expressing gastric mucosa showed specific in vivo accumulation of 111In-labeled MP2288 which could be blocked in the presence of excess unlabeled MP2288. 111In-labeled MP2286 and MP2288 were also found to be stable in human plasma whereas both compounds were degraded in urine (>40% after 3 h at 37 degrees C). The affinity, specificity, biodistribution, and stability of these two DTPA-CCK analogs indicate that these compounds have substantial promise for use in the in vivo visualization of CCK-B receptor-expressing tumors.

In an earlier experimental study, intracellular recording suggested that cholecystokinin (CCK) suppresses a K+ conductance in thalamic reticular (RE) neurons, yet the reversal potential of the CCK response, revealed using voltage clamp, was hyperpolarized significantly relative to the K+ equilibrium potential. Here, biophysical models of RE neurons were developed and used to test whether suppression of the K+ conductance, gK, can account for the CCK response observed in vitro and also to determine the likely site of CCK receptors on RE neurons. Suppression of gK in model RE neurons can reproduce the relatively hyperpolarized reversal potential of CCK responses found using voltage clamp if the voltage clamp becomes less effective at hyperpolarized potentials. Three factors would reduce voltage-clamp effectiveness in this model: the nonnegligible series resistance of the voltage-clamp electrode, a hyperpolarization-activated mixed cation current (Ih) in RE neurons, and the dendritic location of CCK-sensitive K+ channels. Although suppression of gK in the dendritic compartments of model RE neurons simulates both the magnitude and reversal potential of the CCK response, suppression of gK in just the somatic compartment of model RE neurons fails to do so. Thus the model predicts that CCK should effectively suppress K+ conductance RE neuron dendrites and thereby regulate burst firing in RE neurons. This may explain the potent effects of CCK on intrathalamic oscillations in vitro.

The outstanding sensitivity of pentagastrin in detecting the presence of primary, recurrent or metastatic medullary thyroid cancer (MTC) suggests widespread expression of the corresponding receptor type in human MTC. Indeed, recent autoradiographic studies have demonstrated the presence of cholecystokinin (CCK)-B (= gastrin) receptors not only in more than 90% of MTCs but also in a high percentage of small cell lung cancers, stromal ovarian cancers, astrocytomas and several other tumour types. The aim of this study was to evaluate whether radiolabelled gastrin may be suitable for targeting CCK-B receptor-expressing tumours in vivo. For this purpose, the biodistribution of the radioiodinated human heptadecapeptide gastrin-I was studied in nude mice bearing subcutaneous xenografts of the human MTC cell line, TT. Initial therapy experiments were undertaken. Finally, the biodistribution of iodine-131- labelled gastrin-I was studied in a patient with metastatic MTC. At a peptide amount of approximately 1 microg, maximum tumour uptake (8.9+/-2.9%ID/g) was observed in animals at 1 h post injection, with tumour-to-blood ratios as high as 6.3+/-1.9. Physiological CCK-B receptors in the stomach, gallbladder and pancreas of the mice were targeted as well. The major route of excretion was renal, but strong evidence for a biliary excretion pathway also exists. Pilot therapy studies with 131I-labelled gastrin showed significant anti-tumour efficacy as compared with untreated controls. In accordance with the preclinical data, good receptor targeting was observed in the tumour sites, stomach, gallbladder and pancreas of a patient with metastatic MTC. These data suggest that gastrin and its analogues may represent a useful new class of receptor binding peptides for diagnosis and therapy of a variety of tumour types, including MTC and small cell lung cancer. Future preclinical and clinical studies will address in more detail the molecular features that render CCK-B receptor binding agents potentially useful candidates for in vivo scintigraphy and radionuclide therapy.

Gastrin plays an important role in regulating gastric acid secretion and gastrointestinal mucosal growth but its cellular sites of action in man have not been determined. Using cryostat sections of gastric mucosal tissue we have identified (125I-gastrin binding followed by fixation-wet emulsion autoradiography) and characterized (125I-gastrin binding followed by counting) a gastrin receptor binding site in the human stomach. This site displayed binding characteristics similar to those observed in isolated cell systems: specifically, 125I-gastrin binding was rapid (t1/2 approximately 10 min at 37 degrees C), temperature-dependent (3.5 fold more radioligand bound at 22 degrees C than at 4 degrees C) and saturable. The binding of the radioligand was also tissue specific and was five-fold greater in the gastric body than in the gastric antrum and duodenum. In the autoradiographs, silver grains were localized only to parietal cells and not to other epithelial cell types. In the presence of 40 nM gastrin grains were no longer present over parietal cells demonstrating that these sites were both saturable and of high affinity. These data provide the first demonstration of gastrin binding sites (putative receptors) on parietal cells in the human stomach and suggest that gastrin acts directly on these cells to help regulate gastric acid secretion and/or mucosal growth.

The processes underlying the development of neuronal tolerance to and dependence upon opiates are not yet fully understood. To evaluate a possible role for cholecystokinin (CCK) in these processes, quantitative receptor autoradiography and in situ hybridisation histochemistry were used to study both the density and distribution of sulphated CCK octapeptide (CCK8S) binding sites and preproCCK peptide mRNA levels within the dorsal (oxytocin neurone-rich) supraoptic nuclei of rats given an intracerebroventricular (i.c.v.) infusion of morphine over 5 days, which is known to induce tolerance and dependence in mechanisms regulating oxytocin neurones. Specific CCK8S binding was significantly increased in the supraoptic nuclei of both morphine-dependent and salt-loaded (2% sodium chloride to drink for 48 h) rats compared to their respective controls (P < 0.05). In situ hybridisation histochemistry revealed no difference in preproCCK mRNA levels within supraoptic neurones of (i.c.v.) morphine-treated compared with either i.c.v. vehicle-treated or untreated control animals. These results suggest that CCK receptor mechanisms involved in the control of magnocellular oxytocin neurone activation are upregulated during chronic morphine treatment, and this may favour increased sensitivity to CCK, thereby offsetting the inhibitory actions of morphine, contributing to tolerance and perhaps to the withdrawal excitation characteristic of dependence.

Background & Aims: The hypothesis that progastrin-derived peptides act as autocrine growth factors for colorectal carcinomas has generated considerable interest. However, the influence of autocrine gastrins on nontumorigenic colonic cells has not been investigated. This study tested the above hypothesis in the nontumorigenic, conditionally immortalized mouse colon cell line YAMC., Methods: The effects of expression of antisense or sense gastrin messenger RNA, treatment with antibodies against progastrin-derived peptides, or treatment with gastrin receptor antagonists on YAMC cell proliferation were measured., Results: YAMC clones expressing antisense gastrin messenger RNA had reduced levels of immunoreactive progastrin-derived peptides and a reduced rate of proliferation, relative to vector only-transfected cells. Glycine-extended gastrin17, but not amidated gastrin17, reversed the antisense-induced inhibition of proliferation and stimulated the proliferation of sense- or vector only-transfected cells. YAMC cells bound 125I-glycine-extended gastrin17 (Kd, 0.36 nmol/L, 1810 sites/cell), but not 125I-amidated gastrin17, and binding was unaffected by gastrin receptor antagonists including benzotript. Proliferation of all YAMC clones was partially inhibited either by an antibody selective for glycine-extended gastrin or by preincubation with benzotript, and the inhibitory effects were additive., Conclusions: YAMC cells use nonamidated progastrin-derived peptides as autocrine growth factors, partly through binding to an extracellular receptor selective for glycine-extended gastrin, and partly through an intracellular mechanism.

The enterochromaffin-like cell (ECL) cells of the stomach are principally regulated by gastrin via a gastrin/CCK(B) receptor (G[R]) which modulates both histamine secretion and cell proliferation. In the African rodent (mastomys) hypergastrinemia generated by the histamine-2 receptor antagonist (loxtidine) results in ECL cell hyperplasia and neoplasia at 8 and 16 weeks respectively. The expression, structure and function of the G(R) during transformation is however unknown. We utilized a pure (approximately 90%) preparation of ECL cells to evaluate alterations in the G(R) utilizing immunocytochemistry, Western blot analysis, reverse transcription polymerase chain reaction (RT-PCR), 5-bromo-2-deoxyuridine uptake and phosphorylation site analysis. Although the expression of ECL cell G(R) was upregulated at both mRNA (PT-PCR) and protein (Western analysis) level, its affinity to gastrin was decreased in the hyperplastic phase and lost during transformation. The coding sequence of the G(R) of mastomys tumor ECL cells was identical to that of normal ECL cells, parietal cells and the brain. However, the mRNA sequence of the third introcytoplasmic loop of the G(R) was significantly different to other species. In addition, the G(R) exhibited phosphorylation site on serine residue(s). We have thus noted a direct correlation between hypergastrinemia and G(R) alteration and function during ECL cell transformation. It is possible that the unique mastomys gastrin receptor mediated ECL cell transformation involves the novel phosphorylation sites and a divergence in the introcytoplasmic domain.

The expression of cholecystokinin (CCK) and gastrin (G) receptors in human gastrointestinal cancers remains poorly documented and is still of a controversial nature. We have measured the levels of mRNA for CCK-A and CCK-B/gastrin receptors using quantitative reverse transcription-polymerase chain reaction (RT-PCR) in primary digestive cancers and hepatic metastases. CCK-A-receptor mRNA was detected in 5 out of 8 esophageal cancers (0.1-1 fg/microg), in 5 out of 8 gastric cancers (0.05-4.2 fg/microg) and in 5 out of 12 colon cancers (0.1-1 fg/microg RNA). CCK-B/gastrin mRNA was not detected in esophageal cancers but was detected in 7 out of 8 gastric cancers (0.05-5.2 fg/microg), and in only 2 out of 12 colon adenocarcinomas (0.05-1 fg/microg RNA). The expression of the CCK-A receptor in esophageal, gastric and colon cancers and of the CCK-B/gastrin receptor in the majority of gastric adenocarcinomas screened may be an important indicator of the influence of CCK and gastrin of local or systemic origin on the growth of these cancers.

Receptor autoradiography was used to investigate the distribution of brainstem binding sites for cholecystokinin, dopamine and N-methyl-D-aspartate with particular reference to the nucleus of the solitary tract of the rat, an area involved in the control of ingestive behavior. Binding sites for the A and B subtypes of the cholecystokinin receptor, labeled with [(125)I]cholecystokinin octapeptide sulfate in the presence or absence of antagonists for the devazepide (A) or L-365,260 (B) receptor, were present throughout the caudal rostral extent of the nucleus of the solitary tract, the A type predominating in the commissural, medial and gelatinous part and the B type in the lateral part. In the most rostral part of the medial nucleus of the solitary tract, both A and B receptors were present. Dopamine D2 receptors, labeled with [(125)I]NCQ-298, were found in all parts of the nucleus of the solitary tract. No binding to the dopamine D1 receptor, labeled with [(125)I]SCH-23982, was found in the brainstem. N-Methyl-D-aspartate receptors, labeled with [(3)H]dizocilpine maleate, were also present in the entire caudorostral extent of the nucleus of the solitary tract. Binding to cholecystokinin A receptors was co-distributed with [(125)I]NCQ-298 and [(3)H]dizocilpine maleate binding in the caudal and rostral parts of the nucleus of the solitary tract, and binding to cholecystokinin B receptors overlapped with [(125)I]NCQ-298 and [(3)H]dizocilpine maleate binding in the rostral nucleus of the solitary tract. These results are consistent with the hypothesis that cholecystokinin, dopamine and glutamate interact in the nucleus of the solitary tract in the control of ingestive behavior.

Gastrin/CCK-B receptors are involved in the regulation of several types of cells of the gastric mucosa, including the parietal cells, the ECL cells and the D cells. In this study, we aimed at localizing such receptors in the gastric mucosa. For this purpose, we prepared monospecific antibodies against two sequences of the canine gastrin/CCK-B receptor. Sections of formalin-fixed, paraffin-embedded corpus and antrum from dog and guinea-pig were immunostained with these antibodies. In parallel, sections were stained with antibodies against somatostatin. Staining with gastrin/CCK-B receptor antibodies was observed in a few, small epithelial cells in the bottom part of the corpus mucosa. Immunoreactive cells of the antral mucosa were structurally similar, but more frequent. The same cells also stained with somatostatin antibodies. In addition one of the gastrin/CCK-B antibodies reacted with canine submucosal smooth muscle cells. No staining was observed in sections exposed to antibodies that were pre-absorbed with the corresponding antigen. We conclude that gastrin/CCK-B receptors are present in D cells of the gastric mucosa and in submucosal smooth muscle cells.

Background & Aims: Cholecystokinin (CCK) acting via CCK(A) receptors and gastrin acting via CCK(B) receptors exert trophic effects on a variety of nontransformed tissues. However, their role as hormonal regulators of pancreatic cancer is controversial. The aim of this study was to determine the effects of activation of CCK(A) and CCK(B) receptors on the growth of human pancreatic cancer cells in vitro., Methods: Two human pancreatic cell lines MiaPaca-2 and Panc-1 were transfected stably with both CCK receptor subtypes. Effects of CCK on various growth parameters including DNA synthesis, nuclear labeling, and colony formation were evaluated., Results: Cells expressing either receptor subtype, but not untransfected cells, bound ligand and mobilized Ca2+ in response to CCK. CCK treatment caused a sustained pronounced inhibition of anchorage-independent growth. Similarly, CCK treatment inhibited anchorage-dependent growth. Receptor activation caused a concentration and time-dependent reduction in [3H]thymidine incorporation and nuclear labeling in cells cultured anchored to a plastic substrate. However, these effects on anchorage-dependent growth were transient, suggesting cellular desensitization., Conclusions: These data indicate that both CCK receptor subtypes can mediate growth inhibitory responses in pancreatic cancer cell lines and raise the possibility that CCK exerts a predominant growth inhibitory action on human pancreatic cancer cells.

The effects of three C-terminal fragments of cholecystokinin (CCK) (CCK-8-sulphated form [SF], CCK-8-non-sulphated form [NSF] and CCK-4) on insulin and glucagon secretion were examined in sheep in vivo. Each CCK fragment was injected intravenously at a wide range of doses (1 pmol to 3 x 10(5) pmol kg-1). CCK-8(SF) had the lowest threshold dose (10 pmol kg-1) and a maximal response dose of 10(3) pmol kg-1 for increasing plasma insulin concentration; the respective threshold doses of CCK-8(NSF) and CCK-8 for increasing plasma insulin were 30 and 100 times greater than that of CCK-8(SF). A maximal insulin response was not obtained at the highest doses of CCK-8(NSF) or CCK-4 tested (3 x 10(3) and 3 x 10(5) pmol kg-1, respectively). These results indicate that CCK-A type receptors rather than CCK-B receptors may be involved in CCK-induced insulin secretion in sheep. None of the CCK fragments affected plasma glucagon concentration. The lack of a glucagon response to exogenous CCK-fragments may be one of the characteristics of the endocrine pancreatic responses of ruminant species.

The clinical reliability of measuring cholecystokinin (CCK) peptides in the cerebrospinal fluid (CSF) has not been fully elucidated. Therefore, we have assayed CCK-8S and CCK-4 in CSF obtained from 14 healthy male subjects, lumbar-punctured at the L4-5 level following a strictly standardised procedure. CSF concentrations of free CCK-8S and free CCK-4 were used as dependent variables while age, height, body weight, atmospheric pressure and some other factors served as independent variables. It was shown that the CCK-8S ratio between the second (7-12 ml) and first (0-6 ml) CSF fractions, correlated significantly with the atmosphere pressure at the time of puncture. Neither CCK-8S nor CCK-4 displayed concentration gradients in CSF. The CCK-4 levels, expressed as pmol l-1 in the total amount of CSF were found to be positively correlated with the neuraxis distance in the lying position and negatively with the neuraxis distance in the sitting position. Furthermore, CCK-4, expressed as pmol l-1 per min of tapping-time (pmol l-1 min-1), showed a negative correlation with storage time, presumably mirroring a proteolytic process. CCK-8S and CCK-4 intercorrelated positively independently of whether expressed as pmol l-1 or pmol l-1 min-1. In conclusion, the results of this exploratory study indicate that the neuraxis distance (in the sitting and lying positions) and storage-time have to be accounted for when interpreting data on CSF levels of CCK-4. Attention has to be paid to the potential influence of atmospheric pressure on the concentration ratio of CCK-8S.

Background & Aims: Cholecystokinin (CCK) receptors mediate pancreatic secretion and gallbladder contraction. Hitherto, little information on characteristics of CCK receptors in the human pancreas was available. This study identifies CCK receptors in the human pancreas and compares their characteristics with the CCK receptors in the human gallbladder., Methods: Visualization and quantification of 125I-Bolton-Hunter sulfated CCK octapeptide (125I-BH-CCK-8) binding to tissue sections of the human pancreas and gallbladder were performed by storage phosphor autoradiography., Results: Specific bindings for CCK were visualized in pancreatic tissue and the smooth muscle layer of the gallbladder. Binding of 125I-BH-CCK-8 to the pancreas was inhibited by agonists with the affinities (dissociation constant) of CCK (0.11 nmol/L) approximately gastrin (0.15 nmol/L) and by antagonists with the affinities of CCK-B receptor antagonist (L365,260, 0.18 nmol/L) > CCK-A receptor antagonist (lorglumide, 8.1 nmol/L). In contrast to the pancreas, binding of 125I-BH-CCK-8 to the gallbladder muscle was inhibited with high affinity by CCK-8 and lorglumide but was replaced to a small degree by gastrin and L365,260., Conclusions: The sub-types of receptors for CCK in the human pancreas and gallbladder are different. The human pancreas predominantly expresses CCK-B receptors, whereas only CCK-A receptors were localized in the human gallbladder muscle.

In view of recent evidence for a role for the B subtype of cholecystokinin (CCKB) receptor in panic and anxiety, the distribution of CCKB receptors in the forebrain of a Rhesus macaca monkey was examined by receptor autoradiography employing [125I]D-Tyr25(Nleu28,31)-CCK25-33S. CCKB receptors were widely and topographically distributed in cortex. Other structures with notable labelling included the basal ganglia, presubiculum, amygdala, mamillary bodies, cerebellar cortex and pineal gland. The distribution of CCKB receptors further supports roles for this peptide in behavioural processes.

The peptide hormone cholecystokinin (CCK) has been shown to stimulate the growth of azaserine-induced preneoplastic nodules in the rat pancreas. Previously, our labortory demonstrated by classical binding studies that CCK receptors are overexpressed in azaserine-induced rat pancreatic neoplasms. In the present study, we utilized autoradiography to determine the temporal course of this increased receptor binding. Male Lewis rats were given azaserine or saline injections and sacrificed at 2, 4, 8, 12, and 18 months of age. Pancreatic tissue was harvested and autoradiography using 125l-labeled. CCK-8 was performed. Densitometry measurements of azaserine-induced pancreatic nodules, internodular pancreas, and normal pancreatic tissue (from saline-treated controls) of each age group were taken with an image analyzer. There was no statistically significant difference in CCK binding to internodular pancreas and normal pancreas at any age. At 2 months of age, there was no significant increase in CCK binding to azaserine-induced pancreatic nodules. However, at 4, 8, 12, and 18 months of age there was significantly greater CCK binding to azaserine-induced pancreatic nodules than to both internodular pancreas and normal pancreas (p < 0.001 for all groups). At 18 months of age, one azaserine-treated animal developed a pancreatic acinar cell carcinoma, which likewise exhibited significantly greater CCK binding than internodular pancreas or normal pancreas (p < 0.001 for both). These findings demonstrate increased CCK binding in azaserine-induced preneoplastic pancreatic nodules and pancreatic acinar cell carcinoma, compatible with our previous demonstration of receptor overexpression in these tissues. Increased CCK binding first becomes apparent by 4 months following exposure to azaserine. These result suggest that overexpression of CCK receptors, located specifically on preneoplastic and neoplastic pancreatic lesions, results in increased CCK binding and is involved in the mediation of CCK-stimulated growth during azaserine-induced pancreatic carcinogenesis.

Studies on cholecystokinin (CCK) receptors in rat pancreas are usually performed on homogenates. Using storage phosphor autoradiography, a new imaging technique with a high sensitivity and large linear dynamic range, we visualized and characterized CCK receptors in tissue sections of normal rat pancreas. The density of CCK receptors in pancreatic tissue sections from 10 normal rats appeared to be unevenly distributed and variable in serial sections. The binding of labeled CCK-8 was markedly inhibited by CCK-8 and CCK-A receptor antagonists, but it was only weakly affected by gastrin and CCK-B receptor antagonists. At room temperature the CCK-8 dose-inhibition curve was fitted by a two-site model: one with a high-affinity but low-capacity site and another with a low-affinity but high-capacity site. The CCK-8 dose-inhibition curve showed that the inhibition of the variable high-density receptors took place at a low concentration of CCK-8, while the diffuse low-density receptors were inhibited at the high concentration of 1 microM CCK-8. Binding of labeled CCK-8 at 37 degrees C was homogeneous with a low affinity and comprised only 4% of that found at room temperature. In summary, an uneven density of CCK receptors in the rat exocrine pancreas was observed and attributed to the variable expression of high-affinity CCK receptors in pancreatic acini.

The 2 gastrointestinal peptides cholecystokinin (CCK) and gastrin, which act through CCK-A receptors (having high affinity for CCK) or CCK-B/gastrin receptors (having high affinity for CCK and gastrin), are considered to be important tumor growth factors. We have evaluated CCK-A and CCK-B/gastrin receptors in 34 human thyroid cancers using in vitro receptor autoradiography with 2 different radioligands. We demonstrate high-affinity CCK-B/gastrin receptors in medullary thyroid carcinomas, present at very high incidence (92%) but the absence of these receptors in non-medullary thyroid carcinomas or in normal thyroid glands. CCK-B/gastrin receptors are therefore likely to be the molecular substrate for the pentagastrin-stimulation test, widely used in medullary thyroid carcinomas; moreover, they represent the targets for physiologically secreted gastrin or CCK which, as growth factors, may stimulate the growth of medullary thyroid carcinomas. Furthermore, these results have diagnostic as well as therapeutic implications: radiolabeled gastrin and CCK analogs may be used for scintigraphic tumor localization in vivo, whereas CCK-B-selective antagonists may be of therapeutic value.