Bipolarity of duodenal enterochromaffin cells in the rat

Summary Enterochromaffin cells of the rat duodenum have been studied immunocytochemically by use of a specific antiserum to serotonin. At the light-microscopic level serotonin immunoreactivity was observed in enterochromaffin cells located in the epithelium of the duodenal mucosa. Most of the serotonin-immunoreactive material was localized to the basal portion of the enterochromaffin cells, but small amounts of immunoreactive material were regularly observed in the apical portion. At the electron-microscopic level serotonin immunoreactivity in enterochromaffin cells was found to be concentrated over the dense cores of the cytoplasmic granules. The majority of these granules was located in the basal cytoplasm of the enterochromaffin cells, but serotonin-immunoreactive granules were also observed in the apical cytoplasm immediately beneath the microvilli. These observations indicate that duodenal enterochromaffin cells are bipolar and that they secrete serotonin both basally, to the circulation, and apically, to the gut lumen. Rat duodenal enterochromaffin cells thus appear to have an exocrine as well as an endocrine function.     

Ultrastructure of endocrine cells in the stomach of two teleost fish Perca fluviatilis L. and Ameiurus nebulosus L.

Summary In the gastric mucosa of two teleost species, the perch (Perca fluviatilis) and the catfish (Ameiurus nebulosus) three endocrine cell types were found, located predominantly between the mucoid cells of the gastric mucosa. A fourth cell type is present in the gastric glands of catfish. Each cell type was defined by its characteristic secretory granules. Type-I cells were predominant in both fish. These cells contained round or oval granules with a pleomorphic core. The average diameter of granules was 400 nm for the perch and 270 nm for the catfish. Type-II cells of both species displayed small, highly osmiophilic granules about 100 nm in diameter. The secretory granules of type-III cells (260 nm in the perch and 190 nm in the catfish) were round or slightly oval in shape and were filled with a finely particulate electron-dense material. Type-IV cells of the catfish were found in the gastric glands only. Their cytoplasm was filled with homogeneous, moderately electron-dense granules averaging 340 nm in diameter. The physiological significance of these different morphological types of gastric endocrine cells requires further investigation.     

On argyrophil reactions of endocrine cells in the human fetal pancreas

Summary The endocrine cells in the pancreas of five human fetuses with gestational ages of 18–20 weeks were examined by light and electron microscopy with special regard to argyrophil reactions. B-cells and typical A and D-cells were easily identified electron microscopically on the basis of their typical secretory granules. In the Grimelius argyrophil silver stain, a concentration of silver grains over the less electron dense peripheral mantle of the A-cell secretory granules was observed by electron microscopy. In the Hellerström and Hellman modification of the argyrophil Davenport alcoholic silver stain, silver grains were concentrated over the internal structures of the D-cell secretory granules. With this stain an accumulation of silver grains was also seen at the surface of the A-cell secretory granules. The argyrophil reaction of the A-granules was less pronounced than in the D-cells. In addition to B-cells and A- and D-cells, two other types of endocrine cell were observed by electron microscopy. These cells were argyrophil with the silver impregnation method of Grimelius. The electron microscopic findings at least partly explain the frequent overlapping between the two staining methods observed at the light microscope level.This study was supported by the Swedish Medical Research Council (Project No. 102)     

Quantitative electron microscopy of endocrine cells in oxyntic mucosa of normal human stomach

Summary An ultrastructural morphometric study of the endocrine cells of the oxyntic mucosa of the stomach in gastric biopsies collected from five male and five female healthy volunteers aged 19–31 was performed. No sex-related differences were disclosed. Endocrine cells accounted for 1.2±0.4% of the epithelial volume and 0.9±0.4% of the mucosal volume, i.e., including the lamina propria. After classification of the specific endocrine cell types according to the ultrastructural morphology of secretory granules, the volume densities of ECL, P and D cells (30±9%, 24±7%, and 22±4% of the entire endocrine cell mass, respectively) were higher than those of other endocrine cell types. In particular, EC cells contributed less than 10% and X cells represented a very low proportion of the total cells. Non-granulated profiles of cells which in all other respects appeared to be endocrine were also found with a volume density of 8±4%. D cells were distinguished by the high fraction of cytoplasm occupied by secretory granules (31±5%). Subdivision of the whole mucosa into four horizontal segments revealed the endocrine cells to be mostly distributed in the three lower, with virtually no endocrine cells in the superficial segment. The quantitative ultrastructural analysis of the endocrine cell population of the normal human oxyntic mucosa provided by this study may allow a better evaluation of physiological and pharmacological variations of the endocrine cell population.     

Ultrastructural localization of radiolabelled L-dopa in the endocrine hypothalamus of the rat

Summary Light and electron microscopic autoradiography has been employed to define the neuroanatomical patterns of uptake and binding of radiolabelled L-dopa in the endocrine hypothalamus of the rat. A dorsomedial continuum of arcuate and periventricular neurons selectively sequester ³H L-dopa 20 min following its intraventricular infusion. By 40 and 60 min following the infusion labelling of neurons is minimal and supports the notion of rapid degradation. Other cell compartments such as tanycytes demonstrate uptake of ³H L-dopa. The ultrastructural localization and distribution of radiolabelled L-dopa (or its metabolites) in the rodent hypothalamus is discussed with respect to mechanisms and cell compartments involved in neuroendocrine regulatory processes.Supported by USPHS Program Project Grant NS-11642-04 (DES) and RR-05403Career Development Awardee RO4GM-70001     

Intestinal endocrine cells of chicks around the time of hatching

Summary The duodenum of 16-day Black Australorp chick embryos, and the duodenum, ileum, large intestine and caeca of 18-day embryos and of chicks within 30 h of hatching, have been studied by electron microscopy. Cells were found with secretory granules resembling those in mammalian EC, S, A-like, EG and D cells (terminology of Solcia et al., 1973), and were on this basis tentatively identified accordingly. The distribution and frequency of the chick cells in different parts of the tract correspond well to the situation in mammals.Supported by grants from the Senate Research Committee of the University of the Witwatersrand, JohannesburgThe author gratefully acknowledges the help of Professors E. Solcia and N. Ferreira     

Immunohistochemical localization of neurotensin in endocrine cells of the gut

Summary Endocrine cells displaying neurotensin immunoreactivity are found scattered in the jejuno-ileum of all mammals studied, including man. They are rather scarce in rat, guinea pig, rabbit and pig and fairly numerous in cat, dog and man. In most mammals the neurotensin cells predominate on the villi. Only in the dog are they more numerous in the crypts. In the chicken, neurotensin cells occur all along the intestinal tract. They are particularly numerous in the zone that joins the gizzard with the duodenum. The ontogeny of the neurotensin cells in the gut was studied in rats and chickens. In the rat, the cells are first observed in the jejuno-ileum immediately before birth. The adult frequency is reached 4–5 days later. In the chicken, neurotensin cells first appear in the colon in the 18 day old embryo and in the small intestine two days later (i.e. one or two days before hatching). A few days after hatching, the gut has achieved the adult number of neurotensin cells per unit area.     

An immunocytochemical and electron-microscopical study of endocrine cells in the gut and pancreas of a stomachless teleost fish,Barbus conchonius (Cyprinidae)

Summary Four immunoreactive endocrine cell types can be distinguished in the pancreatic islets of B. conchonius: insulin-producing B cells, somatostatin-producing A₁ (= D) cells, glucagon-producing A₂ cells and pancreatic poly-peptide-producing PP cells. The principal islet of this species contains only a few PP cells, while many PP cells are present in the smaller islets. Except for the B cell all pancreatic endocrine cell types are also present in the pancreatic duct.At least six enteroendocrine cell types are present in the gut of B. conchonius: 1. a cell type (I) with small secretory granules, present throughout the intestine, and possibly involved in the regulation of gut motility; 2. a C-terminal gastrin immunoreactive cell, probably producing a caerulein-like peptide; these cells are located at the upper parts of the folds, especially in the proximal part of the intestinal bulb; 3. a met-enkephalin-immunoreactive cell, present throughout the first segment; 4. a glucagon-immunoreactive cell, which is rare in the first segment; 5. a PP-immunoreactive cell, mainly present in the first half of the first segment; 6. an immunoreactive cell, which cannot at present be specified, located in the intestinal bulb. The latter four cell types are mostly located in the basal parts of the folds, although some PP-immunoreactive cells can also be found in the upper parts.Most if not all enteroendocrine cells are of the open type. The possible functions of all enteroendocrine cell types are discussed.Abbreviations BPP bovine pancreatic polypeptide - CCK cholecystokinin - GEP gastro-entero-pancreatic - GIP gastric inhibitory peptide or glucose-dependent insulin releasing peptide - PPP pig pancreatic polypeptide - VIP vasoactive intestinal polypeptide     

Ontogeny and distribution of certain endocrine cells in the human fetal large intestine

Summary In 9 fetuses, 9 to 24 weeks-old, the occurrence and relative distribution of argentaffin cells, as well as of cells immunoreactive to somatostatin (SRIF), glucagon-like polypeptide (GLI), pancreatic polypeptide (PP) and substance P (SP) were studied in five segments of the colon (appendix, cecum, ascending colon, descending colon, and rectosigmoid). For each colonic segment, data concerned with the occurrence of endocrine cells were expressed either as mean absolute numbers of specific cells per entire mucosal section, or as cell densities per mm³ of mucosa after calculation of the mucosal volume of the sections. Argentaffin, GLI, SRIF and PP immunoreactive cells are all present in relatively large numbers, scattered along the entire length of the colonic mucosa as early as the 9th–10th week of gestation, whereas substance P-containing cells occur sporadically and first appear during the 14th–17th week. Until the 20th week, with progressing embryonic development, an increase was determined in absolute numbers per section of all types of endocrine cells in all segments of the colon. This observation is clearly related to the general growth of the colonic mucosa, since cell densities per mm³ of mucosa do not greatly change or even decrease during gestation. However, it is possible that densities of argentaffin, GLI and BPP cells increase in the appendix around the 14th–17th week of gestation. Between the 20th and 24th week, absolute numbers of cells per section remain stable or slightly increase, while cell densities tend rather to decrease in all segments. These data demonstrate that some endocrine cells are present very early in the human fetal colon, but their functional significance remains to be elucidated.This work was supported by the Institut National de la Santé et de la Recherche Médicale (INSERM)     

Ultrastructural studies on the ventricular surface of the frog cerebellum

Summary Ultrastructural studies of the ventricular surface of the frog cerebellum showed regional differences. In the midline region of the adult cerebellum was found a band of profusely ciliated squamous ependymal cells. In the rest of the cerebellum the ependymal cells were columnar and each had a single cilium. In the cerebellum of the premetamorphic tadpole, the squamous ependymal cells of the midline region also were monociliated. During metamorphosis they gradually became multiciliated. Additionally, supraependymal cells and synaptic elements were present on the ventricular surface of the cerebellum of adult frogs as well as in late metamorphic tadpoles. In contrast, supraependymal cells were rarely observed in premetamorphic tadpoles, and it was concluded that the supraependymal system develops during metamorphosis. It is postulated that the band of cilia may be associated with the circulation of cerebrospinal fluid, and supraependymal synaptic elements function in neuroendocrine regulation.     

Ultrastructural studies on the secretory process in the epithelioid cells of the juxtaglomerular apparatus

The epithelioid cells of the juxtaglomerular apparatus have been studied with respect to the release mechanism of the secretory granules. Invaginations of the plasma membrane into the interior of the epithelioid cells are interpreted as stages before or after an exocytotic process. Granules are sometimes observed in close contact with the plasma membrane, and material with electron density similar to that of the granules can also be observed in the invaginations. These morphological features suggest that the granular material of the epithelioid cells is extruded into the texture of the basal lamina. Furthermore, a dense network of microtubules and microfilaments is described and the functional role of this system in exocytosis is discussed.     

Gut hormones in Salamandra salamandra

Summary Histological, cytochemical and immunocytochemical methods were used in light and electron microscopical studies to demonstrate the presence of a neuroendocrine system in the gut of the urodele, Salamandra salamandra.Cytochemical stains capable of detecting peptide-producing endocrine cells demonstrate cells reacting with Masson's silver (argentaffin) method, Grimelius' argyrophil silver method, masked metachromasia method and the lead haematoxylin stain.Using antisera raised to a variety of mammalian gut peptides, cells containing bombesin-, gastrin-, somatostatin-, substance P- and glucagon-like immunoreactivity were identified; vasoactive intestinal polypeptide- and substance P-like immunoreactivities were found in nerve fibres in the submucous and myenteric plexus. No immunoreactivity was detected for motilin, gastric inhibitory polypeptide, cholecystokinin or secretin.The ultrastructure of the immunoreactive cells and nerves was revealed by the semithin/thin method. All the cells identified contained numerous electrondense secretory granules, which varied in their chracteristic morphological structure from one cell type to another.The evidence collected in this study indicates that a complex neuroendocrine system regulating gut function is present in this amphibian and may have developed prior to the emergence of the phylum.     

Ultrastructural characteristics of vasopressin-containing neurons in the paraventricular nucleus of the hypothalamus

Summary Vasopressin-containing neurons, identified by immunocytochemistry, are located predominantly in the posterior magnocellular division of the paraventricular nucleus of the rat hypothalamus. By electron microscopy, the immunoreaction product is seen within the cell bodies and neuronal processes. In the perikarya and dendritic processes, the immunoreactive material is associated primarily with neurosecretory granules. Axonal processes, identified by their content of microtubules and accumulation of neurosecretory granules, show the immunoreaction product in association with both of these organelles. Afferent axo-dendritic, axo-somatic and putative axo-axonic synapses with immunostained vasopressinergic neurons can be identified. The presynaptic profiles do not contain immunoreactive material. This study contributes to the ultrastructural characterization of vasopressinergic neurons in the paraventricular nucleus and of their afferent synaptic input.Supported by NIH Grants HD-12956 and 2SO7RR05403     

Immunocytochemical study of the lung of domestic fowl and pigeon: endocrine cells and nerves

the presence of endocrine cells and nerves in the lung of 2 avian species (Gallus gallus and Columba livia domestica) has been studied by peroxidase-antiper-oxidase (PAP) and avidin-biotin complex (ABC) immunocytochemical methods at the light-microscopic level. Two immunoreactive cell-types have been identified in the epithelium of the primary and secondary bronchi of chick lung: serotonin- and bombesin-immunoreactive cells; and 3 cell-types, namely, serotonin-, bombesin- and CGRP-(calcitonin gene related peptide) immunoreactive cells, have been located in the bronchial epithelium of pigeon lung. Co-localization of 2 different immunoreactivities within the same cell has not been detected. VIP-immunoreactive nerves have been observed in different locations in chick lung.     

A cytochemical and immunofluorescence study of endocrine cells in the gut of the ascidian Styela clava

Summary Strong secretin-like immunofluorescence has been demonstrated in endocrine-like cells from the gastric epithelium of Styela. These cells also stain with lead haematoxylin and exhibit a brilliant formaldehyde-induced fluorescence, but do not show any other cytochemical features characteristic of the mammalian APUD series. Tests with antisera to glucagon, gastrin and somatostatin all proved negative. In the oesophagus tests with all four antisera proved negative. The significance of these results is discussed in relation to the phylogeny of vertebrate gastro-intestinal hormones.     

Ultrastructural immunocytochemical localization of neurophysin and vasopressin in the median eminence and posterior pituitary of the guinea pig

Summary With the use of tissue prepared by freeze-substitution and the unlabelled antibody enzyme technique, neurophysin and vasopressin were localized at the ultrastructural level in the posterior pituitary and median eminence of the guinea pig. In the posterior pituitary neurophysin was found in the large neurosecretory granules (1300–1500 Å) of axons, Herring bodies, and nerve terminals. In some of these axons immunoreactive neurophysin was found outside of granules in the axoplasm. By light microscopy neurophysin was found in both the zona interna and zona externa of the median eminence; this was confirmed by electron microscopy. In the zona interna as in the posterior pituitary, neurophysin was localized both inside and outside the large neurosecretory granules. In the zona externa, immunoreactive deposit was primarily located in granules with a diameter of 900–1100 Å in nerve terminals abutting on the primary portal plexus. The distribution of vasopressin paralleled that of neurophysin except that the hormone was rarely extragranular. These results demonstrate for the first time that both neurophysin and vasopressin are present in granules of axons that are in contact with the hypophysial portal vasculature.The authors wish to thank Dr. Alan Robinson for the gifts of antiserum to bovine neurophysin I and for purified bovine neurophysin I; Dr. Ludwig Sternberger for the peroxidase-anti-peroxidase complex; and Dr. Robert Utiger for antiserum to lysine vasopressinSupported in part by U.S. Public Health Service grant RR-00167 to the Wisconsin Regional Primate Research Center from the National Institutes of Health. Primate Center publication No. 14-017.Recipient of NIH, NINDS Teacher-Investigator Award NS-1108.     

Ultrastructural and cytochemical studies on nucleolus-like bodies in early postimplantation rat embryos

Summary This study investigates the role of the developing diencephalic floor or mesenchymal tissue in the differentiation of ACTH-producing cells.The adenohypophysial primordia of fetal rats on days 12.5 and 13.5 of gestation were treated with collagenase; some primordia were allowed to retain an association with the brain and mesenchyme, but in others the brain and/or mesenchyme were removed. These different combinations of tissues were cultured and examined by immunohistochemical techniques using antisera against pACTH and synthetic -MSH. Removal of mesenchyme alone had little effect on the development of ACTH cells as compared to primordia maintained with brain and mesenchyme. In contrast, removal of the brain with or without mesenchyme on day 12.5 resulted in a marked decrease of ACTH cells accompanied by a mal-growth of adenohypophysial tissue. Such changes were slight when the brain was separated from day 13.5 primordia. Immunoreactive -MSH cells were sparse or absent in all cases.These results suggest that in fetal rats the developing diencephalic floor is essential for differentiation of ACTH cells before day 13.5 of gestation whereas mesenchyme has no apparent effect.     

Immunocytochemical and ultrastructural characterization of endocrine cells in chicken proventriculus

Summary The endocrine cells of the chicken proventriculus were investigated immunocytochemically, using the peroxidase-antiperoxidase technique on paraffin and semithin sections for light microscopy, and immunogold staining in osmium-fixed material for electron microscopy. The fixation procedure also allowed a detailed ultrastructural investigation. Twenty-three antisera were tested and 7 immunoreactive cell-types were identified: D-cells containing somatostatin-like peptide; EG-cells immunoreactive to anti-glucagon, anti-GLP1 and antineurotensin; NT-cells labelled only with anti-neurotensin; BN-cells containing bombesin-like material; ENK-cells showing met-enkephalin immunoreactivity; EC-cells reactive to anti-serotonin; and APP-cells positive to anti-avian pancreatic polypeptide. In addition, enterochromaffin-like (ECL) cells, were also detected by electron microscopy. The presence of ENK-cells and the ultrastructure of these and NT-cells are described for the first time in chicken proventriculus, and glucagon, GLP1 and neurotensin are shown to be colocalized in the EG-cells.     

Ultrastructural and cytochemical investigations on the functional role of proteinaceous nuclear inclusions (PNIs) in chlorenchyma plant cells

Summary— Detailed investigations on the fine ultrastructural organization of different forms of proteinaceous nuclear inclusions (PNls) in chlorenchyma plant cells suggest they consist of the same elementary subunits. Previous high magnification of TEM micro-graphs had shown that the amorphous type of inclusion (A) was mainly composed of elementary fibrils measuring 3.0–3.5 nm in diameter, with no orderly spatial arrangement. New computer image treatments of electron micrographs allowed us to establish that the 8.0–13.0 nm thick filaments — forming the fibrillar (F), crystalline (C) and lamellar (L) inclusions — consist of two elementary fibrils which are coiled in a helix with variable pitch, depending on the type of inclusion. A further secondary coiling of two filaments, about 8.0–9.0 nm in diameter, gives the 20.0–25.0 nm thick tubules which form the characteristic tubular inclusion (T). Correlating the distributive data of PNIs with observations on their ultrastructural morphology and with micrographs of partial aggregation or disgregation patterns of the inclusions, led to the hypothesis that the different forms are not different classes of proteins, but simply different stages of structural complexity of the same protein. To determine whether the intranuclear inclusion protein is nucleolar or nucleolus-associated, cytochemical and immunocytochemical tests were performed on ultrathin sections or leaf lamina tissue in block. These techniques proved that PNIs do not belong to the class of argyrophilic proteins (AgNOR-proteins), and particularly not nucleolin and fibrillarin, two of the major nucleolar proteins. Structural similarities to other plant inclusions, especially P-proteins, and to animal and plant intermediate cytoskeletal filaments (IFs) are discussed with regard to the functional role of PNIs.