An immunohistochemical study of gut endocrine cells in two species of insectivorous vespertilinid bats (Chiroptera: Pipistrellus abramus and Plecotus auritus sacrimontis)

11 endocrine cell types immunoreactive for either 5-hydroxytryptamine (5-HT), somatostatin, gastrin, cholecystokinin (CCK), gastric inhibitory peptide (GIP), motilin, secretin, neurotensin, pancreatic glucagon, enteroglucagon or bovine pancreatic polypeptide (BPP) were found in gastrointestinal tract of 2 species of insectivorous bats. 5 of these 11 types of endocrine cells were located in the stomach and all 11 types of endocrine cells were found in the intestine. However, the distribution and relative frequency of each immunoreactive endocrine cell varied among the cell types and between the 2 species of bats examined. In Brunner's glands, gastrin- and 5-HT-immunoreactive cells were detected very rarely in Pipistrellus and only occasionally in Plecotus. The present results obtained from the insectivorous bats were compared with those of the sanguivorous vampire bats.     

Ultrastructural identification of a new cell type — the N-cell as the source of neurotensin in the gut mucosa

The neurotensin-cell is identified immunohistochemically and ultrastructurally by differential counting of endocrine cells in the gut of a primate (Tupaia belangeri). Utilizing light microscopy, the EC-cells are identified by the Masson-Fontana silver stain; with the same method the neurotensin cells are not stained. The other endocrine cells have been quantified in the small intestine using the peroxidase-antiperoxidase stain with antisera against glucagon, somatostatin, cholecystokinin, gastrin, secretin, pancreatic polypeptide, gastric inhibitory peptide and neurotensin. In the ileal mucosa of Tupaia, the most frequent endocrine cell is the EC-cell followed by the glucagonoid cell, (L-cell). The immunoreactive neurotensin cell represents the third most frequent endocrine cell in this region. On the ultrastructural level, this third most frequent endocrine cell is a heretofore undescribed cell, the N-cell, containing electron dense secretory granules measuring 335 +/- 87 nm in diameter.     

An immunocytochemical investigation with monoclonal antibodies to somatostatin

Four monoclonal antibodies specific for somatostatin have been produced and characterized. These antibodies were used to assess the anatomical relationship of somatostatin-containing cells in the pancreas and gastrointestinal tract of man, baboon and rat with ten other peptide-containing endocrine cells. The peptides investigated were gastrin, cholecystokinin, motilin, secretin, neurotensin, gastric inhibitory polypeptide, gut-glucagon, pancreatic glucagon, pancreatic polypeptide and insulin. The only regions in which somatostatin cells were seen in close contact with another endocrine cell were in the pancreas and the gastric antrum. In the pancreas somatostatin cells were commonly seen in close contact with insulin, glucagon and pancreatic polypeptide cells and infrequent contact was demonstrable with the gastrin-immunoreactive cells in the antrum of both rat and man. In all other cases no evidence was obtained for a close anatomical relationship between somatostatin cells and the other enteroendocrine cells.     

Regulatory peptides in the gastrointestinal tract of Alligator mississipiensis

Summary The gastrointestinal tract of the alligator Alligator mississipiensis has been investigated for the presence of immunoreactivity to fourteen regulatory peptides all known to occur in the mammalian gut system.Mucosal endocrine cells reacting specifically with the antisera to neurotensin, C-terminal gastrin, somatostatin, bombesin, secretin, pancreatic glucagon and enteroglucagon were detectable, the distribution of these cells being, in general, similar to the mammalian pattern. Peripheral nerve cell bodies and nerve fibres were detected with the antisera to vasoactive intestinal polypeptide, substance P, bombesin and somatostatin again with a distribution similar to that seen in mammals.No immunoreactivity was observed with the available antisera to glicentin, motilin, gastric inhibitory polypeptide, gastrin 34, cholecystokinin 9–20 and met-enkephalin.     

An immunocytochemical investigation with monoclonal antibodies to somatostatin

Summary Four monoclonal antibodies specific for somatostatin have been produced and characterized. These antibodies were used to assess the anatomical relationship of somatostatin-containing cells in the pancreas and gastrointestinal tract of man, baboon and rat with ten other peptide-containing endocrine cells. The peptides investigated were gastrin, cholecystokinin, motilin, secretin, neurotensin, gastric inhibitory polypeptide, gut-glucagon, pancreatic glucagon, pancreatic polypeptide and insulin.The only regions in which somatostatin cells were seen in close contact with another endocrine cell were in the pancreas and the gastric antrum. In the pancreas somatostatin cells were commonly seen in close contact with insulin, glucagon and pancreatic polypeptide cells and infrequent contact was demonstrable with the gastrin-immunoreactive cells in the antrum of both rat and man. In all other cases no evidence was obtained for a close anatomical relationship between somatostatin cells and the other enteroendocrine cells.     

The distribution of endocrine cells in the mucosa of the gastrointestinal tract of the house musk shrew,Suncus murinus (Insectivora)

Summary The distribution of endocrine cells in the gastrointestinal tract of the house musk shrew, Suncus murinus (Family Soricidae, Order Insectivora) was studied immunohistochemically. The hormones investigated were gastrin, cholecystokinin (CCK), somatostatin, secretin, glucagon, gastric inhibitory polypeptide (GIP), motilin and neurotensin. In the gastric mucosa, gastrin and somatostatin cells were only found in the pyloric regions, and no other hormonal cell-types were observed. In the intestinal mucosa, the largest number of endocrine cells belonged to the gastrin and glucagon/glicentin cell-types, whereas CCK-33/39 and secretin cells were the least numerous. Numbers of other cell-types were intermediate between these two groups. The gastrin and GIP cells were mostly localized in the proximal portion of the intestine, decreasing in number towards the distal portion. The motilin and CCK-33/39 cells were restricted to the proximal half. The glucagon/glicentin and neurotensin cells were most abundant in the middle portion. The somatostatin and secretin cells, although only present in small numbers, were randomly distributed throughout the intestine. This characteristic distribution of gastrointestinal endocrine cells is discussed in comparison with the distribution patterns of other mammals.Dr. Munemitsu Hoshino, who was Professor of the Department of Pathology and directed this study, passed away on May 23rd 1988     

Gastrin and ACTH-like immunoreactivity occurs in two ultrastructurally distinct cell types of rat antropyloric mucosa

Summary The properties of endocrine cells of rat antropyloric mucosa, which simultaneously store both gastrin and ACTH-like immunoreactivity have been examined. In freely fed animals all or nearly all antral gastrin cells contain also large quantities of ACTH-like immunoreactivity. Following three days of fasting the gastrin cell content of ACTH-like peptides is drastically reduced, but increases rapidly upon refeeding of the starved animals for 30 min. At the electron microscopical level, the vast majority of cells storing both gastrin and ACTH-like peptides are identified as G cells but, in addition, a few, previously unrecognized, endocrine cells have also been found to store both types of peptides. The latter new cell type has tentatively been labelled the G_a cell. In normal freely fed animals the G cell is characterized by the occurrence of both electron-dense and electron-lucent granules. Correlative immunocytochemical and ultrastructural studies indicate that gastrin and the ACTH-like peptides are both stored in the cytoplasmic granules. Our results indicate that the gastrin cells release their content of ACTH-like peptides in response to fasting and that this release is blocked by refeeding. The differential release of two hormone-like substances from the same endocrine cell type is of great interest for analysis of mechanisms of peptide hormone release.     

Distribution of gastrin and CCK cells in the rat gastrointestinal tract. Evidence for the occurrence of three distinct cell types storing COOH-terminal gastrin immunoreactivity.

Gastrin and cholecystokinin (CCH) cells of the rat gastrointestinal tract have been studied by immunocytochemistry and radioimmunoanalysis. With antisera directed against the COOH-terminal tetrapeptide sequence, which is common to gastrin and CCK, three distinct endocrine cell types are detected. One of the cell types predominates in the antrum, is scarce in the rest of the gut and corresponds to the gastrin cell. The second cell type is virtually confined to the duodenum and jejunum and corresponds to the CCK cell. The third cell type occurs disseminated in the small intestines, predominates in the ileum, and reacts with COOH-terminus-specific antisera only following diethylpyrocarbonate and not following formaldehyde fixation. It is possible that the third cell type stores a third member of the gastrin-CCK family of gut hormones.     

Distribution of gastrin and CCK cells in the rat gastrointestinal tract

Summary Gastrin and cholecystokinin (CCK) cells of the rat gastrointestinal tract have been studied by immunocytochemistry and radioimmunoanalysis. With antisera directed against the COOH-terminal tetrapeptide sequence, which is common to gastrin and CCK, three distinct endocrine cell types are detected. One of the cell types predominates in the antrum, is scarce in the rest of the gut and corresponds to the gastrin cell. The second cell type is virtually confined to the duodenum and jejunum and corresponds to the CCK cell. The third cell type occurs disseminated in the small intestines, predominates in the ileum, and reacts with COOH-terminus-specific antisera only following diethylpyrocarbonate and not following formaldehyde fixation. It is possible that the third cell type stores a third member of the gastrin-CCK family of gut hormones.     

Targeted ablation of secretin-producing cells in transgenic mice reveals a common differentiation pathway with multiple enteroendocrine cell lineages in the small intestine.

The four cell types of gut epithelium, enteroendocrine cells, enterocytes, Paneth cells and goblet cells, arise from a common totipotent stem cell located in the mid portion of the intestinal gland. The secretin-producing (S) cell is one of at least ten cell types belonging to the diffuse neuroendocrine system of the gut. We have examined the developmental relationship between secretin cells and other enteroendocrine cell types by conditional ablation of secretin cells in transgenic mice expressing herpes simplex virus 1 thymidine kinase (HSVTK). Ganciclovir-treated mice showed markedly increased numbers of apoptotic cells at the crypt-villus junction. Unexpectedly, ganciclovir treatment induced nearly complete ablation of enteroendocrine cells expressing cholecystokinin and peptide YY/glucagon (L cells) as well as secretin cells, suggesting a close developmental relationship between these three cell types. In addition, ganciclovir reduced the number of enteroendocrine cells producing gastric inhibitory polypeptide, substance-P, somatostatin and serotonin. During recovery from ganciclovir treatment, the enteroendocrine cells repopulated the intestine in normal numbers, suggesting that a common early endocrine progenitor was spared. Expression of BETA2, a basic helix-loop-helix protein essential for differentiation of secretin and cholecystokinin cells was examined in the proximal small intestine. BETA2 expression was seen in all enteroendocrine cells and not seen in nonendocrine cells. These results suggest that most small intestinal endocrine cells are developmentally related and that a close developmental relationship exists between secretin-producing S cells and cholecystokinin-producing and L type enteroendocrine cells. In addition, our work shows the existence of a multipotent endocrine-committed cell type and locates this hybrid multipotent cell type to a region of the intestine populated by relatively immature cells.     

Signaling pathways mediating gastrin's growth-promoting effects.

In addition to its fundamental role in stimulating gastric acid secretion, the peptide hormone gastrin induces growth-promoting effects on diversity of target cells. Various mechanisms, including endocrine, paracrine, and autocrine, have been proposed for gastrin's growth-promoting actions. The mitogenic effects of gastrin are mediated by specific cell surface receptors activated after gastrin binding. The functionally defined receptors for gastrin include cholecystokinin A (CCKA) receptor, which is discriminating for sulfated CCK8; cholecystokinin B (CCKB)/gastrin receptor, which binds gastrin17 sulfated, and nonsulfated CCK8 with nearly equal affinities; cholecystokinin C (CCKC), which is a low-affinity gastrin binding protein; and novel, high-affinity receptors selective for amidated gastrin, processing intermediates of gastrin, or both. The signaling pathways mediating gastrin's stimulation of the CCKB/gastrin receptor have been progressively outlined, and the pathways mediating other receptors have been slowly emerging. Engagement of the gastrin receptor initiates various biochemical and molecular events, including recruitment and activation of tyrosine kinases, activation of the phospholipase C signaling pathway leading to phosphoinositide breakdown, intracellular calcium mobilization and protein kinase C stimulation, activation of the mitogen-activated protein kinase pathway, and induction of early response genes. Current emphasis is on understanding the functional significance of processing intermediate forms of gastrin, and the receptor subtypes and pathways that promote the trophic/mitogenic effects of the different molecular forms of gastrin.     

Evolutionary relationships of the gut hormones.

Peptides identical or related to mammalian gut hormones occur widely, not just in gut endocrine cells but also in central or peripheral nerves, amphibian skin glands, and a variety of invertebrate tissues. The dual distribution in brain and gut was probably already established early in the vertebrate line; representatives of the oldest vertebrate group, the cyclostomes, have cholecystokinin-like factors in gut endocrine cells and in brain. The related sequences of certain gut peptides, notably gastrin and cholecystokinin (CCK), and secretin, glucagon, vasoactive intestinal polypeptide (VIP), and gastric inhibitory peptide (GIP), indicate evolution from common ancestral molecules by gene duplication and divergence. Functionally important residues are conserved. Thus the COOH-terminal pentapeptide common to gastrin and CCK also contains their minimal active fragment. There are also evolutionary changes at the level of the target organ receptor mechanisms: these are also evolutionary changes at the level of the target organ receptor mechanisms; these are illustrated by evidence suggesting that secretin regulates the flow of pancreatic juice in mammals whereas the structurally related peptide VIP has a similar role in birds.     

Morphological variation of immunoreactive cells positive to cholecystokinin 33 (10–20) and gastrin 34 (1–15) in human duodenum

Summary Human duodenal endocrine cells reactive with antibodies to cholecystokinin (CCK) 33 (10–20) and/or gastrin 34 (1–15) were studied by a combination of immunohistochemical and electron-microscopic methods. By immunohistochemistry, three types of endocrine cells were distinguished in human duodenal mucosa, i.e., those only positive for only CCK, those positive for both CCK and gastrin and those only positive for only gastrin. Ultrastructurally, the first cell type is characterized by many secretory granules with an eccentric dense core (mean diameter; 271+-74 nm). The second cell type, which was less frequent than the other two, has ultrastructural features that resemble type-I cells. The last cell type was composed of two types of cells containing small secretory granules identical to those of IG cells (mean diameter; 171+-31 nm) or large secretory granules indistinguishable from those of I cells (mean diameter; 286+-50 nm).     

Ultrastructural localization of cholecystokinin in endocrine cells of the dog duodenum by the immunogold technique

Cholecystokinin (CCK) has been localized by the immunogold technique in a type of endocrine cell of the dog duodenum characterized by small (166 +/- 38 nm) secretory granules with fairly dense, homogeneous core separated from its enveloping membrane by a thin clear space. The CCK cell is immunocytochemically distinct and cytologically different from other types of endocrine cells, as the secretin, GIP and motilin cells, already identified in the dog duodenum.     

An immunohistochemical survey of endocrine cells and nerves in the proximal small intestine of the platypus,Ornithorhynchus anatinus

The relative frequencies of endocrine cells and peptidergic nerve elements in the proximal small intestine of the adult platypus were studied by immunohistochemistry. Six kinds of endocrine cells - serotonin (5-HT)-, somatostatin-, gastrin-, motilin-, cholecystokinin (CCK)- and bovine pancreatic polypeptide (BPP)-immunoreactive cells - were identified in this study. These endocrine cells were found most frequently in the intestinal glands, in moderate numbers in the tubular ducts and were infrequent in the surface folds. 5-HT-immunoreactive cells were most numerous, somatostatin-, gastrin-, motilin- and BPP-immunoreactive cells were moderately numerous, whereas CCK-immunoreactive cells were rare. Five kinds of neuropeptides: substance P, vasoactive intestinal polypeptide (VIP), gastrin releasing peptide (GRP), somatostatin and leu-enkephalin, were detected in the intramural nerve elements. Substance P-, VIP- and GRP-immunoreactive nerve fibers were found most frequently in the lamina propria mucosae of the surface folds. The relationships between the possible functions of the peptides and amine detected in this study as well as the characteristic structure of the digestive tract of the adult platypus are discussed.     

Ultrastructural localization of cholecystokinin in endocrine cells of the dog duodenum by the immunogold technique

Summary Cholecystokinin (CCK) has been localized by the immunogold technique in a type of endocrine cell of the dog duodenum characterized by small (166±38 nm) secretory granules with fairly dense, homogeneous core separated from its enveloping membrane by a thin clear space. The CCK cell is immunocytochemically distinct and cytologically different from other types of endocrine cells, as the secretin, GIP and motilin cells, already identified in the dog duodenum.     

Secretin-cells of the mammalian intestine contain serotonin

Summary Various endocrine cells contain biogenic amines in addition to their peptide hormones. In the digestive tract, one of these amines is serotonin that is regularly present in enterochromaffin (EC-) cells. Previously, it has been assumed that other entero-endocrine cell types also contain this amine. Moreover, it was presumed that chromogranin A, an acidic glycoprotein, is involved in storage mechanisms for biogenic amines in endocrine cells. Using immunohistochemical techniques, we now exemplarily investigated cholecystokinin (CCK-) and secretin (S-) cells of five adult mammalian species for their content of serotonin and of chromogranin A. In all mammalian species, CCK-cells were devoid of serotonin but contained chromogranin A immunoreactivity of varying densities. In contrast, S-cells of all mammals were immunoreactive for serotonin; however, immunoreactivities for this biogenic monoamine were heterogeneous and varied from dense to faint or lacking immunostainings. Likewise, immunoreactivities for chromogranin A in S-cells showed inter-species and inter-cellular heterogeneities. S-cells containing serotonin were simultaneously immunoreactive for chromogranin A and the density of immunoreactivities for both were correlated in given S-cells. Based on mutual relationships of chromogranin A and serotonin immunoreactivities, we assume that chromogranin A is virtually a prerequisite for the S-cells' content of serotonin and that this protein participates in storage mechanisms for biogenic amines in endocrine cells.S-cells have now to be added to the family of amine-storing endocrine cells. Basically, serotonin-storing endocrine cells in the digestive tract cannot be simply regarded as enterochromaffin (EC-) cells any longer; the current nomenclature and classification of entero-endocrine cells should be reviewed in this respect.This work was supported by grants of the Deutsche Forschungs-gemeinschaft (EN 65/15-2)     

Changes in gastric endocrine cells in Balb/c mice bearing CT-26 carcinoma cells: an immunohistochemical study

The distribution and density of gastric endocrine cells in Balb/c mice bearing CT-26 carcinoma cells were studied immunohistochemically employing specific antisera against serotonin, somatostatin, glucagon, gastrin, cholecystokinin (CCK)-8 and human pancreatic polypeptide (hPP). The animals were divided into two groups, a non-implanted sham group and a CT-26 carcinoma cell-implanted group. Samples were collected from two regions of the stomach (fundus and pylorus) at 28 days after implantation of the medium or the CT-26 cells (1x10(5) cells/mouse). Five of the 6 types of immunoreactive (IR) cells were identified, with only the hPP IR cells not being detected. The regional distribution of the gastric endocrine cells in the CT-26 implanted group was similar to that of the non-implanted sham group. However, the endocrine cells were significantly decreased in the CT-26-implanted group as compared to those of the non-implanted sham group. Serotonin- and somatostatin-IR cells in the fundus and pylorus , and gastrin- and CCK-8-IR cells in the pylorus of the CT-26 implanted groups were significantly decreased compared to those of the sham group. In addition, glucagon-IR cells were restricted only to the fundus of the sham animals. hPP-IR cells were not detected in either the T-26 implanted- or the non-implanted group. Since endocrine cells are the anatomical units responsible for the production of gut hormones, a change in their density may reflect a change in their capacity to produce such hormones. Implantation of the tumor cell mass induced severe quantitative changes in gastric endocrine cell density, an abnormality which may contribute to the development of gastrointestinal symptoms, such as anorexia and indigestion, frequently encountered in cancer patients.     

Immunocytochemical study of gastro-entero-pancreatic (GEP) endocrine cells in the vampire bat (Desmodos rotundus)

The distribution and frequency of gastro-entero-pancreatic (GEP) endocrine cells were studied in vampire bats by immunocytochemistry. Moderate numbers of somatostatin- and a few 5-hydroxytryptamine (5-HT)- and glucagon-immunoreactive cells were seen in the fundic cecum of the stomach. Numerous gastrin- and moderate numbers of somatostatin- and 5-HT-immunoreactive cells were found in the pyloric region. Moderate numbers of 5-HT-, somatostatin-, and gastrin-immunoreactive cells also were found in BRUNNER's glands. In addition to the above-mentioned 4 immunoreactive cell types, cells immunoreactive for glicentin, secretin, cholecystokinin (CCK), gastric inhibitory peptide (GIP), and neurotensin were found in the intestine. Numerous insulin-, moderate numbers of somatostatin- and glucagon-, and a few 5-HT-immunoreactive cells were detected in the pancreatic islets with lesser numbers scattered within the exocrine pancreas. Motilin- and pancreatic polypeptide-immunoreactive cells were not observed in this study.