THE FINE STRUCTURE OF VON EBNER''S GLAND OF THE RAT

The fine structure of von Ebner's gland was studied in untreated rats and rats stimulated to secrete by fasting-refeeding or injection of pilocarpine. Cytological features were similar to those reported for pancreas and parotid gland. Abundant granular endoplasmic reticulum filled the basal portion of the cell, a well-developed Golgi complex was located in the vicinity of the nucleus, and the apical portion of the cell was filled with dense secretory granules. Dense heterogeneous bodies resembling lysosomes were closely associated with the Golgi complex. Coated vesicles were seen in the Golgi region and also in continuity with the cell membrane. Granule discharge occurred by fusion of the granule membrane with the cell membrane at the secretory surface. Successive fusion of adjacent granules to the previously fused granule formed a connected string of granules in the apical cytoplasm. Myoepithelial cells were present within the basement membrane, and nerve processes were seen adjacent to acinar and myoepithelial cells. Duct cells resembled the intercalated duct cells of the major salivary glands.     

Ultrastructure and complex carbohydrate ultracytochemistry of the cat parotid gland

The structure and glycoconjugate content of the cat parotid gland were analyzed at electron microscopic level by applying morphological techniques and three ultrastructural histochemical methods - HID-TCH-SP, LID-TCH-SP and PA-TCH-SP. This gland appeared as a typical salivary gland composed of acinar secretory cells, intercalated ducts, striated ducts and excretory ducts. The most common configuration of secretory granules consisted of a dense core surrounded by a variable electron-lucent halo. All ductal segments were characterized by the presence of different cell populations and small apical granules greatly different from those localized in the acinar cells. By using HID-TCH-SP we were able to demonstrate that in a few acinar cells there are sulphated sites, whereas PA-TCH-SP staining revealed the presence of vic-glycol radicals in all acinar cells preferentially located on the halo of secretory granules.     

Peculiar salivary glands in a silk‐producing mite Bakericheyla chanayi (Cheyletidae)

This is the first ultrastructural investigation of salivary glands in the family Cheyletidae. In both sexes of Bakericheyla chanayi, paired acinous salivary glands and tubular coxal glands were shown to be united into the common podocephalic system. The secretory portion of the salivary gland includes medial and lateral lobes composed of the five and two cells, respectively, with clearly distinct ultrastructure. The cytoplasm of the cells is occupied by the secretory granules containing fine fibrous material. The fine structure of both cell types suggest a proteinaceous nature of their secretions. A single central process extending from the apical face of each secretory cell passes through the common acinar cavity to enter the conducting duct. A pair of intercalary cells at the base of the conducting duct links it with the secretory portion of the gland. Extending towards the acinar cavity, protrusions of intercalary cells alternate the apical regions of the secretory cells and form with them highly‐specialized contacts characterized by the apical network of microtubules and microfilaments. Two possible ways of secretion are suggested: 1) exocytosis into the acinar cavity and 2) direct passage via the central processes. The detection of axon profiles in the gland body suggests a neural control for the glandular cell function. In tritonymphs, neither secretion nor large lateral lobe cells were observed up to the pharate stage when the lateral lobe undergoes rapid differentiation. The arrangement of the acinous gland is compared to that of other arthropods. Its composition appears to be close to the class three of insect glands. The involvement of the lateral lobe cells in silk production is discussed. J. Morphol. 276:772–786, 2015. © 2015 Wiley Periodicals, Inc.     

Immunofluorescence-microscopic demonstration of myosin and actin in salivary glands and exocrine pancreas of the rat

Summary Actin and myosin were localized in various salivary glands (parotid, submandibular, sublingual, lingual and Harderian gland) and the exocrine pancreas of rats by indirect immunofluorescence microscopy using specific rabbit antibodies against chicken gizzard myosin and actin. A bright immunofluorescent staining with both antibodies was observed at three main sites: (1) In myoepithelial cells of all salivary glands, (2) in secretory gland cells underneath the cell membrane bordering the acinar lumen (except Harderian and mucous lingual gland), and (3) in epithelial cells of the various secretory ducts (of all glands) in similar distribution as in acinar cells. The present immunohistochemical findings in acinar cells could lend further support to a concept suggesting that myosin and actin are involved in the process of transport and exocytosis of secretory granules.Supported by grants form Deutsche Forschungsgemeinschaft (Dr. 91/1, Ste. 105/19 and U. 34/4). We thank Mrs. Ursula König, Mrs. Christine Mahlmeister and Miss Renate Steffens for excellent technical assistance.     

Ultrastructure of human labial salivary glands. I. Acinar secretory cells

The structure of human labial salivary gland acini was studied by light and electron microscopy. Contrary to previous reports, these glands were pure mucous in nature; no serous elements were present. The acinar cells were found in all stages of maturation. Immature cells were characterized by an extensive and highly organized rough-surfaced endoplasmic reticulum. The Golgi complex was extremely prominent, consisting of stacks of flattened cisternae and swarms of small vesicles. Mucous droplets were almost completely absent. As secretory activity progressed, the endoplasmic reticulum involuted, while the Golgi cisternae became distended and formed many vacuoles. In mature mucous cells, the apical cytoplasm was filled with membrane-bounded mucous droplets, and the nucleus was displaced basally. The droplets frequently showed great variation in density from cell to cell, and even within the same cell they sometimes were quite heterogeneous. They were liberated from the acinar cells by an apocrine process, so that droplets with intact limiting membranes were often observed in the acinar lumen. These droplets soon lysed, their contents fusing into streams of mucus. Occasionally during apocrine secretion a mucous cell failed to reconstitute its apical surface, and its entire contents spilled into the acinar lumen. Unusual cytoplasmic inclusions were present in many of the acinar cells. These inclusions, which were surrounded by a single membrane, consisted of lipid droplets closely associated with bundles of fine filaments.     

An electron microscopic study of the acinar cells of the submaxillary salivary glands of white rats]

Numerous rows of rough endoplasmic reticulum, large Golgi apparatus with condensation vacuoles and other ultrastructures typical for secreting cells of the exocrine type (secretory granules, smooth and covered vesicles, lysosomes, microtubules, mitochondria) were found in glandular acinar cells of the submaxillary salivary glands of albino rats. The substantial features of the given object are: firstly, low electron density of the content of the secretory granules which seems to be connected with high content of polysaccharides in the secretion and secondly, the presence of special inclusions covered with a typical three-layer membrane. The latter together with the rest of the content of the granules may come into the lumen of the central duct of the acinus and intercellular secretory capillaries.     

An ultrastructural study of the prostate gland of megascolicid earthworm Lampito mauritii (Kinberg)

The ultrastructure of prostate gland of Lampito mauritii revealed two types of secretory cells. Type 1 cells with a broad basal region and a long apical region contain electron dense oval secretory granules with an increased density at the core region. Numerous electron lucent granules with fine filamentous and electron dense amorphous materials also occur at the basal region of these cells. Type 2 cells contain electron lucent mucous-like secretory granules. This cell type contains exceptionally large Golgi complexes having 20-23 stacked cisternae. Both cell types open into a common lumen and numerous microtubules are visible at the apical end. Junctional complexes, such as desmosomes and septate junctions, are observed in this glandular tissue.     

The expression of water channel proteins during human salivary gland development: a topographic study of aquaporins 1, 3 and 5

Some members of aquaporin family (AQP) plays crucial functions in salivary synthesis and secretion. These proteins expression has already been reported during salivary gland formation, however no previous studies in human developing glands have been performed. We evaluated AQP1, 3 and 5 expression through the stages of human salivary gland morphogenesis and discuss the possible role of AQP for glandular maturation. Human salivary glands derived from foetuses aged between 14 and 25 weeks were submitted to immunohistochemistry. At the bud stage, membrane expression of AQP1, 3 and 5 were observed within the epithelial bud cells presenting a similar apicolateral pattern, also found at the pseudoglandular stage, present within the terminal portions of future acini, while AQP5 was also particularly strong at the apical membrane of pre-acinar and pre-ductal cells. AQP5 was co-localised with Cytokeratin 7. Similar AQP1, 3 and 5 expression were observed at the following canalicular stage, where distinct and strongly luminal and acinar AQP5 expression is present. During the final terminal bud stage, AQP1 was only identified in serous acini, myoepithelial and endothelial cells, while differentiated mucous acinar cells and ducts were negative. AQP3 was detected at apicolateral membranes of both mucous and serous acini. AQP5 also showed a diffuse expression in mucous and serous acini, in addition to strong apical membrane expression within lumen of intercalated ductal cells. This topographic analysis of AQP1, 3 and 5 revealed differences in the expression pattern throughout salivary gland developmental stages, suggesting different roles for each protein in human glandular maturation.     

An electron microscopic study of the salivary gland of Rhynchosciara angelae

Summary The structure of the salivary gland of the dipteran insect Rhynchosciara angelae in a defined stage of the larval development, characterized by the synthesis and storage of secretion product, is described. Observations were made with both Nomarski optics and electron microscopy. Filiform projections extending into the lumen of the gland were observed in the apical portion of the cells. At the basal region junctions, characterized as hemidesmosomes, were observed between the membrane of the cell and the basal lamina. The plasma membrane presents numerous infoldings into the cell increasing considerably the surface area at this region. Throughout the cytoplasm of the gland cells numerous mitochondria, Golgi complexes, microtubules, profiles of endoplasmic reticulum, secretion granules and glycogen granules were observed. Carbohydrates were detected on ultrathin sections by using the periodic acid-silver methenamine and the periodic acid-thiosemicarbazide-silver proteinate techniques.     

The amylase-secreting cells of the stomach of the scallop, Pecten maximus: ultrastructural, immunohistochemical and immunocytochemical characterizations

(1) alpha-amylase was extracted and purified from the stomach/digestive gland complex of the scallop Pecten maximus and an anti-serum was induced against the purified amylase by rabbit immunization. (2) The anti scallop amylase was used to localize the amylase-secreting cells in the stomach of Pecten maximus by immunofluorescence and immunogold labelling. The amylase-secreting cells are glandular cells particularly numerous in the main sorting area of the stomach. Their secretory granules were found strongly positive for anti-amylase. Three types of glandular cells were observed, actually corresponding to the three stages of the glandular-cell activity, synthesis, secretion and excretion. (3) The synthesizing cell shows the characteristic features of a protein-synthesizing cell: a conspicuous nucleolus and abundant granular endoplasmic reticulum. In the secretory cell, the secretory granules are formed by the Golgi apparatus and accumulate in the apical part of the cell. The secretory cell is filled with two types of secretory granules which are released in the stomach lumen by apocrine excretion. (4) The present study brings the first demonstration of the synthesis and extracellular release of amylase by glandular cells of the stomach epithelium of a bivalve.     

Postnatal development of the Mongolian gerbil uterine glands

The development of gerbil uterine glands has been described. Gland formation starts about day 6 postnatally (p.n.). At day 10 p.n. the glandular lumen contains already some fluid, and the glandular cells are in an activated state. The apical shape of the glandular cells depends mainly on the width of the lumen. Secretory vesicles in early developmental stages are electron-lucent. As sexual maturity is reached, secretory granules become electron-dense. This change in secretory granular structure indicates a change in the quality of uterine gland fluid.     

Ultrastructure of human labial salivary glands. 3. Myoepithelium and ducts

Myoepithelial cells were present between the basal lamina and the acinar secretory cells of human labial salivary glands. In form and disposition, they resembled myoepithelial cells in the major salivary glands. Many of these cells possessed single cilia on their upper surfaces. Such cilia occasionally extended into invaginations of the overlying secretory cell. The intercalated ducts were variable in occurrence. Their epithelium ranged from columnar to squamous, and showed few signs of secretory activity. Few intralobular ducts possessed basal striations. While mitochondria were abundant in non-striated cells, they were randomly disposed in both basal and apical cytoplasm, and the basal plasmalemma showed only occasional infoldings. The paucity of true striated ducts in labial salivary glands may be responsible for the high concentration of sodium and chloride in unstimulated labial gland salivary secretions.     

Monoclonal antibodies as probes for plasma membrane domains in the exocrine pancreas

Monoclonal antibodies (mAb) were generated as probes for the plasma membrane domains of pancreatic acinar cells. Primary monolayer cultures of mouse pancreatic acinar cells, which have an expanded apical surface relative to normal pancreas, were used to immunize rats. With conventional immunization and fusion protocols, 3% of the hybridomas were positive against the acinar lumen by indirect immunofluorescence of mouse pancreas cryosections. Culturing of spleen cells from an immunized rat on the apical surface of acinar cell monolayer cultures before fusion with the myeloma (an in vitro boost) doubled the percentage of hybridomas producing apical membrane-specific mAb. Monoclonal antibodies were characterized by immunofluorescence, ultrastructural immunoperoxidase cytochemistry, immunoprecipitation, and immunoblotting. One antibody, acinar-1 (IgG2a), labeled the apical membranes of pancreatic acinar cells, hepatocytes, salivary and lacrimal gland acinar cells, and the brush border of small intestine enterocytes. This mAb precipitated and blotted a protein of 94 KD. Acinar-2 (IgM) also labeled pancreatic acinar cell apical membranes but did not label other tissues and did not precipitate or blot. Acinar-3 labeled pancreatic acinar cell lateral membranes. Duct-1 (IgM) labeled pancreatic duct apical membrane and ducts in liver and salivary glands but did not precipitate or blot. These domain-specific mAb demonstrate that common antigenic determinants occur in the apical surfaces of several exocrine epithelia and may be important in secretion.     

Exocytosis in living salivary glands: direct visualization by video-enhanced microscopy and confocal laser microscopy

Although exocytosis is widely believed to involve granule movement, membrane fusion and the emptying of granule content, direct study of these processes has been difficult in living cells because of the limited resolution of conventional light microscopy. Using video-enhanced microscopy and confocal laser microscopy, we have now studied these processes in living rat parotid and submandibular gland acinar cells. Under a differential interference contrast (DIC) microscope equipped with a CCD camera and a high speed image processor, secretory granules were in general stationary even after secretory stimulation with isoproterenol (IPR). Following IPR stimulation, however, there were abrupt changes in light intensity of secretory granules, and many granules disappeared. Confocal microscopy was then performed to confirm whether the observed changes in granules were related to membrane fusion and content release. For this, cells were perfused with the fluid-phase tracer Lucifer Yellow; confocal images thus obtained clearly demonstrated the appearance of fluorescence in omega-shaped invaginations of the apical plasma membrane which corresponded to the sites at which changes were observed in DIC images. The time sequence analyses of confocal images showed that there was a repetitive appearance and disappearance of omega-shaped fluorescent foci at the apical plasma membrane until most of the granules were depleted. During this time, there did not appear to be any significant expansion of the apical plasma membrane and if endocytic uptake of the tracer occurred, it was below the limit of detection. These observations provide new insights into the exocytotic process in salivary glands and are at variance in some respects with previous interpretations made from electron microscopy.     

Spatiotemporal analysis of exocytosis in mouse parotid acinar cells

Exocrine cells of the digestive system are specialized to secrete protein and fluid in response to neuronal and/or hormonal input. Although morphologically similar, parotid and pancreatic acinar cells exhibit important functional divergence in Ca²⁺ signaling properties. To address whether there are fundamental differences in exocytotic release of digestive enzyme from exocrine cells of salivary gland versus pancreas, we applied electrophysiological and optical methods to investigate spatial and temporal characteristics of zymogen-containing secretory granule fusion at the single-acinar cell level by direct or agonist-induced Ca²⁺ and cAMP elevation. Temporally resolved membrane capacitance measurements revealed that two apparent phases of exocytosis were induced by Ca²⁺ elevation: a rapidly activated initial phase that could not be resolved as individual fusion events and a second phase that was activated after a delay, increased in a staircaselike fashion, was augmented by cAMP elevation, and likely reflected both sequential compound and multivesicular fusion of zymogen-containing granules. Optical measurements of exocytosis with time-differential imaging analysis revealed that zymogen granule fusion was induced after a minimum delay of 200 ms, occurred initially at apical and basolateral borders of acinar cells, and under strong stimulation proceeded from apical pole to deeper regions of the cell interior. Zymogen granule fusions appeared to coordinate subsequent fusions and produced persistent structures that generally lasted several minutes. In addition, parotid gland slices were used to assess secretory dynamics in a more physiological context. Parotid acinar cells were shown to exhibit both similar and divergent properties compared with the better-studied pancreatic acinar cell regarding spatial organization and kinetics of exocytotic fusion of zymogen granules. membrane capacitance; differential imaging; zymogen; gland slice; exocrine cells     

Development of female accessory glands of Chrysomya putoria (Wiedemann) (Diptera: Calliphoridae) during oogenesis

Females of Chrysomya putoria (Diptera: Calliphoridae) have two sexual accessory glands, which are tubular and more dilated at the distal extremity. The glands open independently into the common oviduct. Two morpho-physiological regions were distinguished in the longitudinal semi-thin sections of the glands. The secretory region is constituted by three layers: a cuticular intima, lining the lumen, followed by a layer of small cells, and then a layer of very large secretory cells. The ductal region of the gland presents only two layers: the cuticular intima and a cellular layer. In both regions a basement membrane is present. Each secretory cell has in its apical region a reservoir, which enlarges throughout oogenesis; in its basal region there is a large nucleus. The ductal cells are cylindrical and smaller than the secretory cells. The glandular secretion is synthesized in the cytoplasm of the secretory cells, stored and/or modified in the reservoir, then drained to the lumen through an end apparatus seen in the apical region of the secretory cell. Histochemical tests indicate that this secretion is a glycoprotein. Measurements of the glands from females at different physiological conditions and fed on different diets correlate with the results obtained for changes in the ovary during oogenesis. Cell number averaged 561.2 ± 77.54 per gland. There was no increase in cell number during oogenesis.     

Reinternalization of secretory proteins during membrane recycling in rat pancreatic acinar cells

Membrane recycling in pancreatic acinar cells involves endocytic vesicle formation at the apical cell surface and rapid membrane traffic to the Golgi complex. During this process a small amount of extracellular content is taken up from the acinar lumen. In order to determine whether secretory proteins already released into the pancreatic acinar lumen are reinternalized during membrane retrieval, 3H-labeled amylase or 125I-labeled secretory proteins were reinfused through the pancreatic duct until the lumina were reached. Tissue samples from various time points were prepared for light and electron microscope autoradiography. The observations showed that [3H]amylase and, to a lesser extent, the 125I-labeled secretory proteins were internalized at the apical cell surface and rapidly (within 2-5 min) transferred to the Golgi cisternae and the condensing vacuoles; only a minor proportion of silver grains was observed over lysosomes. In addition, at later time points, mature secretion granules close to the Golgi complex became labeled. The results indicate that exocytosis in the rat exocrine pancreas does not operate at 100% efficiency; part of the exported amylase and part of the total secretion product are reinternalized concomitantly with the endocytic removal of plasma membrane and are copackaged together with newly synthesized secretory proteins.     

Rab27b regulates exocytosis of secretory vesicles in acinar epithelial cells from the lacrimal gland

Tear proteins are supplied by the regulated fusion of secretory vesicles at the apical surface of lacrimal gland acinar cells, utilizing trafficking mechanisms largely yet uncharacterized. We investigated the role of Rab27b in the terminal release of these secretory vesicles. Confocal fluorescence microscopy analysis of primary cultured rabbit lacrimal gland acinar cells revealed that Rab27b was enriched on the membrane of large subapical vesicles that were significantly colocalized with Rab3D and Myosin 5C. Stimulation of cultured acinar cells with the secretagogue carbachol resulted in apical fusion of these secretory vesicles with the plasma membrane. Evaluation of morphological changes by transmission electron microscopy of lacrimal glands from Rab27b(-/-) and Rab27(ash/ash)/Rab27b(-/-) mice, but not ashen mice deficient in Rab27a, showed changes in abundance and organization of secretory vesicles, further confirming a role for this protein in secretory vesicle exocytosis. Glands lacking Rab27b also showed increased lysosomes, damaged mitochondria, and autophagosome-like organelles. In vitro, expression of constitutively active Rab27b increased the average size but retained the subapical distribution of Rab27b-enriched secretory vesicles, whereas dominant-negative Rab27b redistributed this protein from membrane to the cytoplasm. Functional studies measuring release of a cotransduced secretory protein, syncollin-GFP, showed that constitutively active Rab27b enhanced, whereas dominant-negative Rab27b suppressed, stimulated release. Disruption of actin filaments inhibited vesicle fusion to the apical membrane but did not disrupt homotypic fusion. These data show that Rab27b participates in aspects of lacrimal gland acinar cell secretory vesicle formation and release.     

Ultrastructure and cytochemistry of salivary glands of the predator Podisus nigrispinus (Hemiptera: Pentatomidae)

Podisus nigrispinus Dallas (Hemiptera: Pentatomidae) is a zoophytophagous insect with a potential for use as a biological control agent in agriculture because nymphs and adults actively prey on various insects by inserting mouthparts and regurgitating the contents of the salivary glands inside the prey, causing rapid paralysis and death. However, the substances found in saliva of P. nigrispinus that causes the death of the prey are unknown. As a first step to identify the component of the saliva of P. nigrispinus, this study evaluated the ultrastructure and cytochemistry of the salivary glands of P. nigrispinus. The salivary system of P. nigrispinus has a pair of principal salivary glands, which are bilobed with a short anterior lobe and a long posterior lobe, and a pair of tubular accessory glands. The principal gland epithelium is composed of a single layer of cells enclosing a large lumen. Epithelial cells of the principal salivary gland vary from cubic to columnar shape, with one or two spherical and well-developed nuclei. Cells of the anterior lobe of the principal salivary gland have an apical surface with narrow, short, and irregular plasma membrane foldings; apical and perinuclear cytoplasm rich in rough endoplasmic reticulum; and mitochondria with tubular cristae. The basal portion of the secretory cells has mitochondria associated with many basal plasma membrane infoldings that are short but form large extracellular canals. Secretory granules with electron-dense core and electron-transparent peripheral are dispersed throughout the cytoplasm. Cells of the posterior lobe of the principal salivary gland are similar to those of the anterior lobe, except for the presence of mitochondria with transverse cristae. The accessory salivary gland cells are columnar with apical microvilli, have well-developed nucleus and cytoplasm rich in rough endoplasmic reticulum, and have secretory granules. Cytochemical tests showed positive reactions for carbohydrate, protein, and acid phosphatase in different regions of the glandular system. The principal salivary glands of P. nigrispinus do not have muscle cells attached to its wall, suggesting that saliva-releasing mechanism may occurs with the participation of some thorax muscles. The cytochemical and ultrastructural features suggest that the principal and accessory salivary glands play a role in protein synthesis of the saliva.     

Natural killer cells regulate murine cytomegalovirus-induced sialadenitis and salivary gland disease

The transmission of herpesviruses depends on viral shedding at mucosal surfaces. The salivary gland represents a major site of persistent viral replication for many viruses, including cytomegalovirus. We established a mouse model of salivary gland dysfunction after acute viral infection and investigated the cellular requirements for the loss of secretion. Murine cytomegalovirus (MCMV) infection severely impaired saliva secretion independently of salivary gland virus levels. Lymphocytes or circulating monocytes/macrophages were not required for secretory dysfunction. Dysfunction occurred before glandular inflammation, suggesting that a soluble mediator initiated the disruption of acinar cell function. Despite genetic differences in innate resistance to MCMV, NK cells protected the host against acinar atrophy and the loss of secretions under conditions of an exceedingly low virus inoculum. NK cells also modulated the type of glandular inflammation after infection, as they prevented an influx of Siglec-F(+) polymorphonuclear leukocytes (PMNs). Therefore, beyond their recognized role in controlling MCMV replication, NK cells preserve organ integrity and function and regulate the innate inflammatory response within the gland.