Release Of Secretory Products Research Articles

The neuro-exocrine secretion: A new type of gland in tapeworms?

The phenomenon of exocrine secretion via nervous cells into the host tissue has been discovered in cestodes. In five cestode species of different orders specialized “cup-shaped” free nerve endings located in the tegument have been found. Their ultrastructure is characterized by the presence of a septate junction, a thin support ring and neurosecretory vesicles 90–110 nm in diameter, which are secreted onto the surface of the tegument through a thin pore. The phenomenon is referred to in this article as the neuro-exocrine secretion. We observed a direct relationship between neurosecretory processes in the deep subtegument and free endings in a series of ultrathin sections in two species. The peripheral neurosecretory neurons of species studied are characterized by similar ultrastructural features: size and location; diameter of neurosecretory granules; absence of microtubules and mitochondria in the neurites. The size of neurosecretory granules has been found to decrease from perikaryon towards neurosecretory terminals that lead to the tegument. In two species, we examined the neurosecretion during incubation in the host's blood serum. Depending on the time of incubation we have shown the changes a) in the diameter of the cup-shaped endings, b) in the number of secretory vesicles in the endings; c) changes in number and diameter of neurosecretory vesicles in the processes of neurosecretory neurons in the subtegument. The detected changes differ in D.dendriticus and L.interrupta and, taken together, indirectly confirm the secretory specialization of the cup-shaped endings. Supposed targets for the neurosecretory neurons in the studied cestodes are the following: (a) eccrine frontal gland ducts, especially their terminal regions involved in the release of secretory products; (b) longitudinal and circular muscles in the subtegument region; (c) the basal membrane of the tegument. Besides the discovered secretion vesicles through the cup-shaped terminals, we observed vacuoles derived from the basal membrane of the tegument containing extracellular substances released into the host tissue. Their possible role in the release of neurosecretory substances is discussed. Considering the data acquired via immunocytochemical methods, an assumption about involvement of FMRFamide-like related peptides (FaRPs) in the neuro-exocrine secretion is proposed. Possible functions of the neuro-exocrine secretion are discussed in the context of host-parasite interactions.

First evidence of holocrine secretory mechanism in granular ducts inmouse mandibular gland

The mouse mandibular gland contains granular ducts, which possess a particular structure. They are termed granular ducts because they contain numerous granules in the cells lining them, having a secretory activity. We aimed to assess the secretory mechanism in granular ducts. Consequently, we harvested the mandibular glands from five male mice and processed them for histological investigations. A large number of cells appeared intact, without obvious distortions or disruptions in the cell membrane. Based on the morphological aspect, these cells either transferred the secretory product through exocytosis or were in "standby" mode. There were cells that presented prominent bulges of the apical pole, thus suggesting an apocrine mechanism. Some cells presented discontinuities in the apical pole and a tendency of granule transfer towards the lumen. According to the morphological aspects, these cells eliminated the secretory product through a holocrine mechanism. The morphological aspects noticed in this study revealed that the release of secretory products in some cells lining the granular ducts in the mouse mandibular gland can also be achieved through a holocrine mechanism.

Ultrastructural studies on the neurosecretory cells in the pars intercerebralis of larvae of tasar silkworm, Antheraea mylitta (D) eco-race Bhandara (Lepidoptera: Saturniidae)

The cerebral neurosecretory cells (NSC) constitute four paired groups, medial (MNC), lateral (LNC-I, LNC-II) and posterior (PNC) in the brain of larvae of tasar silkworm Antheraea mylitta (D) Eco-race Bhandara. The MNC is the largest groups of peptidergic neurosecretory cells and are located in the pars intercerebralis region. The transmission electron microscopic (TEM), ultrastructure of the NSC confirmed the presence of mitochondria, endoplasmic reticulum, Golgi bodies, lysosomes and neurosecretory granules. The median neurosecretory cells shows secretory activity and release of secretory products, the neurosecretory granules (NSG) in the axons of NSC as well as the blood sinus.

Ultrastructural Characteristics of Molecular Release of Secretory Products from Thyroid Mast Cells Induced by Lipopolysaccharide

Ultrastructural characteristics of molecular release of secretory products from thyroid mast cells during different periods after LPS injection were studied in rats. The main morphological manifestation of this mechanism was progressing hydration of the hyaloplasm around secretory granules and dissolution and release of osmiophilic contents from the granules to the cytoplasm with the formation of a hydropic vacuole at the site of the granule.

Cardiac Natriuretic Peptides and Development of Hereditary Hypertension in Rats

Ultrastructure of the right atrial cardiomyocytes of suckling ISIAH rats was studied to clarify the role of cardiac natriuretic peptides in hypertension development during the period when blood pressure is not yet elevated. Cardiomyocytes diameter was significantly greater, Golgi complex was more developed, and granules in the sarcoplasm were more abundant in ISIAH rats as soon as on postnatal day 12 in comparison with age-matched normotensive animals. The smaller diameter of granules and their qualitative composition (ratio of forming, mature, and dissolving forms) attest to active synthesis and release of secretory product. In 21-day-old ISIAH rats, granule size and qualitative composition reflected increased accumulation of hormones in the cells. Thus, morphological features of increased production of natriuretic peptides in the right atrial myocytes were revealed in rats during the first postnatal month before manifestation of hereditary hypertension.

Adipocytes decrease Runx2 expression in osteoblastic cells: Roles of PPARγ and adiponectin

The mechanisms through which bone marrow adipocytes might influence differentiation and function of osteoblasts are not completely understood. To investigate the direct effects of bone marrow fat cells on osteoblast function, an ex vivo co-culture system was utilized comprising either primary fat cells or differentiated 3T3-L1 adipocytes and osteoblastic cells on transwells. In co-culture, both adipocytes and osteoblastic cells were differentiated into adipocytes or osteoblasts, respectively, before culturing on transwells. Co-culture with either primary fat cells or fully differentiated 3T3-L1 adipocytes significantly decreased mRNA and protein expression of runt-related transcription factor 2 (Runx2) in osteoblastic cells. An increase in mRNA and protein expression of peroxisome proliferator-activated receptor γ (PPARγ) occurred concomitantly with the reduction of Runx2 expression. Adiponectin concentration was increased in the media by co-culture. In addition, co-culture with conditioned media from fat cells increased PPARγ promoter activity and decreased Runx2 promoter activity. Knockdown of PPARγ or adiponectin receptor 1 in osteoblastic cells by siRNA prevented the down-regulation of mRNA expression of Runx2 in osteoblastic cells cultured with fully differentiated 3T3-L1 cells. Furthermore, co-transfection with PPARγ decreased Runx2 promoter activity. A marker of osteogenesis, alkaline phosphatase activity in osteoblastic cells was significantly decreased by co-culture. Annexin V/propidium iodide staining showed that co-culture did not induce apoptosis in osteoblastic cells. Thus, we conclude that adipocytes modulate key metabolic functions of osteoblasts through the release of secretory products. PPARγ plays a key role in mediating the effects of adipocytes on osteoblasts.

Serotonin-Producing Cells in Human Gastric Mucosa - Immunohistochemical and Electron Microscopic Study

The great many hormones released by the endocrine cells of the glands and lining epithelium of gastric mucosa determine its significance for the processes in the gastrointestinal tract. One of these hormones, serotonin, plays an important role in the regulation of the motility, secretion and sensation in the gastrointestinal tract. The aim of the present study was to conduct immunohistochemical and electron microscopic studies of serotonin-producing EC cell of gastric mucosa. Gastric mucosa biopsies were obtained and studied immunihistochemically for serotonin expression in the mucosa endocrine cells. Electron microscopic study was performed to specify the processes of synthesis, accumulation and release of secretory product by those cells. The immunohistochemical study revealed a considerable number of serotonin-containing EC cells scattered in the lining epithelium and between the glands in the corpus and pyloric region of the stomach. The electron microscopic study followed the stages of formation of the secretory granules from the initial accumulation of granular substance, its membrane packing and formation of mature granules to their disintegration in the secretory process. Serotonin as a neurotransmitter and gastrointestinal hormone appears to be a key to understanding a number of symptoms of gastrointestinal disorders like nausea, vomiting, pain, diarrhea and constipation. A detailed study of serotonin functions in the gastrointestinal tract realised through different types of receptors, and of the development of specific antagonists and agonists to these receptors would open up new opportunities for a more efficient treatment of gastrointestinal disorders.

Thyroid hormone affects secretory activity and uncoupling protein-3 expression in rat harderian gland.

The effects of T(3) administration on the rat Harderian gland were examined at morphological, biochemical, and molecular levels. T(3) induced hypertrophy of the two cell types (A and B) present in the glandular epithelium. In type A cells, the hypertrophy was mainly due to an increase in the size of the lipid compartment. The acinar lumina were filled with lipoproteic substances, and the cells often showed an olocrine secretory pattern. In type B cells, the hypertrophy largely consisted of a marked proliferation of mitochondria endowed with tightly packed cristae, the mitochondrial number being nearly doubled (from 62 to 101/100 microm(2)). Although the average area of individual mitochondria decreased by about 50%, the total area of the mitochondrial compartment increased by about 80% (from 11 to 19/100 microm(2)). This could be ascribed to T(3)-induced mitochondrial proliferation. The morphological and morphometric data correlated well with our biochemical results, which indicated that mitochondrial respiratory activity is increased in hyperthyroid rats. T(3), by influencing the metabolic function of the mitochondrial compartment, induces lipogenesis and the release of secretory product by type A cells. Mitochondrial uncoupling proteins 2 and 3 were expressed at both mRNA and protein levels in the euthyroid rat Harderian gland. T(3) treatment increased the mRNA levels of both uncoupling protein 2 (UCP2) and UCP3, but the protein level only of UCP3. A possible role for these proteins in the Harderian gland is discussed.

Structure and Composition of the Fusion Pore

Earlier studies using atomic force microscopy (AFM) demonstrated the presence of fusion pores at the cell plasma membrane in a number of live secretory cells, revealing their morphology and dynamics at nm resolution and in real time. Fusion pores were stable structures at the cell plasma membrane where secretory vesicles dock and fuse to release vesicular contents. In the present study, transmission electron microscopy confirms the presence of fusion pores and reveals their detailed structure and association with membrane-bound secretory vesicles in pancreatic acinar cells. Immunochemical studies demonstrated that t-SNAREs, NSF, actin, vimentin, α-fodrin and the calcium channels α1c and β3 are associated with the fusion complex. The localization and possible arrangement of SNAREs at the fusion pore are further demonstrated from combined AFM, immunoAFM, and electrophysiological measurements. These studies reveal the fusion pore or porosome to be a cup-shaped lipoprotein structure, the base of which has t-SNAREs and allows for docking and release of secretory products from membrane-bound vesicles.

Cuticular structures and epidermal glands of Echinoderes cantabricus and E. hispanicus (Kinorhyncha, Cyclorhagida) with special reference to their taxonomic value.

The body wall of two species of kinorhynchs, Echinoderes cantabricus and E. hispanicus, was examined with transmission and scanning electron microscopy, to determine accurately the nature, arrangement, and consistency of characters used for taxonomic purposes. Structural details of cuticular hairs, pectinate fringes, spines, tubules, and different cuticular scars are described and discussed. Two kinds of epidermal glands, types 1 and 2, are characterized according to the nature of their secretory products, their exact location and distribution in the trunk segments, and their possible value as taxonomic characters. The generally assumed function of tubules for the release of secretory product is analyzed and finally rejected in light of the different distribution of tubules and glands along the trunk and because of the absence of a clear anatomic relation between both structures. The cuticular features studied can be useful for taxonomic purposes because of their consistency, but some of them are difficult to access and evaluate and therefore must be used with caution.

Toxines bactériennes : outils pour l’étude des protéines G impliquées dans les mécanismes de l’exocytose dans les cellules neuroendocrines

In neuroendocrine cells, regulated exocytosis is a multistep process that comprises the recruitment and priming of secretory granules, their docking to the exocytotic sites, and the subsequent fusion of granules with the plasma membrane leading to the release of secretory products into the extracellular space. Using bacterial toxins which specially inactivate subsets of G proteins, we were able to demonstrate that both trimeric and monomeric G proteins directly control the late stages of exocytosis in chromaffin cells. Indeed, in secretagogue-stimulated chromaffin cells, the subplasmalemmal actin cytoskeleton undergoes a specific reorganization that is a prerequisite for exocytosis. Our results suggest that a granule-bound trimeric Go protein controls the actin network surrounding secretory granules through a pathway involving the GTPase RhoA and a downstream phosphatidylinositol 4-kinase. Furthermore, the GTPase Cdc42 plays a active role in exocytosis, most likely by providing specific actin structures to the late docking and/or fusion steps. We propose that G proteins tightly control secretion in neuroendocrine cells by coupling the actin cytoskeleton to the sequential steps underlying membrane trafficking at the site of exocytosis. Our data highlight the use of bacterial toxins, which proved to be powerful tools to dissect the exocytotic machinery at the molecular level.

Ca2+-independent and Ca2+/GTP-binding protein-controlled exocytosis in a plant cell.

Exocytosis allows the release of secretory products and the delivery of new membrane material to the plasma membrane. So far, little is known about the underlying molecular mechanism and its control in plant cells. We have used the whole-cell patch-clamp technique to monitor changes in membrane capacitance to study exocytosis in barley aleurone protoplasts. To investigate the involvement of Ca2+ and GTP-binding proteins in exocytosis, protoplasts were dialyzed with very low (<2 nM) and high (1 microM) free Ca2+ and nonhydrolyzable guanine nucleotides guanosine 5'-gamma-thio]triphosphate (GTP[gammaS]) or guanosine 5'-[beta-thio]diphosphate (GDP[betaS]). With less than 2 nM cytoplasmic free Ca2+, the membrane capacitance increased significantly over 20 min. This increase was not altered by GTP[gammaS] or GDP[betaS]. In contrast, dialyzing protoplasts with 1 microM free Ca2+ resulted in a large increase in membrane capacitance that was slightly reduced by GTP[gammaS] and strongly inhibited by GDP[betaS]. We conclude that two exocytotic pathways exist in barley aleurone protoplasts: one that is Ca2+-independent and whose regulation is currently not known and another that is stimulated by Ca2+ and modulated by GTP-binding proteins. We suggest that Ca2+-independent exocytosis may be involved in cell expansion in developing protoplasts. Ca2+-stimulated exocytosis may play a role in gibberellic acid-stimulated alpha-amylase secretion in barley aleurone and, more generally, may be involved in membrane resealing in response to cell damage.

Annual cycle of sperm storage in spermathecae of the red-spotted newt, Notophthalmus viridescens (Amphibia: Salamandridae).

Female sperm storage was studied in a population of Notophthalmus viridescens from South Carolina. Spermathecae initiate production of a glycoprotein secretory product in October. At this time ovarian follicles are immature (0.5-0.9 mm dia), and mating does not occur despite spermiation in males. Six of the 10 females collected in December had sperm in their spermathecae, indicating onset of mating. Unmated females collected in October and sacrificed in February and March possessed mature ovarian follicles (1.3-1.4 mm dia), and the spermathecae contained large secretory vacuoles 2-3 μm dia. Release of secretory product is concomitant with the appearance of sperm in the spermathecae. Thus mated females lack secretory vacuoles in the spermathecal epithelium, and additional synthesis of secretory product does not occur. All females collected in February and March have mated. Sperm are embedded in the spermathecal epithelium and are undergoing degradation in February. Degradation of sperm in the lumen and epithelium is evident in specimens examined from May and June. Atresia of ovarian follicles begins in April in captive specimens, and specimens captured from the bay in May are spent. A general postbreeding emigration from the pond occurs in summer. Fourteen females collected 7 March were injected with human chorionic gonadotropin (hCG) on 9 March and laid fertile eggs 10-18 March. Two of these females were sacrificed each month from April-September; all retained some sperm in their spermathecae, but further oviposition did not occur. Four females were kept 1 year after oviposition of fertile eggs, and oviposition again was induced with hCG; these eggs were infertile, and spermathecae lacked sperm. Spermathecae are inactive from June-September in captive and wild-caught specimens. Sperm may be stored effectively up to 6 months (December-May), and no evidence was found for retention of viable sperm from one breeding season to the next. © 1996 Wiley-Liss, Inc.

Annual cycle of sperm storage in spermathecae of the red‐spotted newt, Notophthalmus viridescens (Amphibia: Salamandridae)

Female sperm storage was studied in a population of Notophthalmus viridescens from South Carolina. Spermathecae initiate production of a glycoprotein secretory product in October. At this time ovarian follicles are immature (0.5–0.9 mm dia), and mating does not occur despite spermiation in males. Six of the 10 females collected in December had sperm in their spermathecae, indicating onset of mating. Unmated females collected in October and sacrificed in February and March possessed mature ovarian follicles (1.3–1.4 mm dia), and the spermathecae contained large secretory vacuoles 2–3 μm dia. Release of secretory product is concomitant with the appearance of sperm in the spermathecae. Thus mated females lack secretory vacuoles in the spermathecal epithelium, and additional synthesis of secretory product does not occur. All females collected in February and March have mated. Sperm are embedded in the spermathecal epithelium and are undergoing degradation in February. Degradation of sperm in the lumen and epithelium is evident in specimens examined from May and June. Atresia of ovarian follicles begins in April in captive specimens, and specimens captured from the bay in May are spent. A general postbreeding emigration from the pond occurs in summer. Fourteen females collected 7 March were injected with human chorionic gonadotropin (hCG) on 9 March and laid fertile eggs 10–18 March. Two of these females were sacrificed each month from April-September; all retained some sperm in their spermathecae, but further oviposition did not occur. Four females were kept 1 year after oviposition of fertile eggs, and oviposition again was induced with hCG; these eggs were infertile, and spermathecae lacked sperm. Spermathecae are inactive from June-September in captive and wild-caught specimens. Sperm may be stored effectively up to 6 months (December-May), and no evidence was found for retention of viable sperm from one breeding season to the next. © 1996 Wiley-Liss, Inc.

The fusion pore interface: a new biological frontier

The development of micro-voltammetry to detect the release of secretory products from single cells has yielded surprising information, which suggests that the release of secretory products is regulated after the fusion of secretory vesicles with the plasma membrane. This technique has also been used to demonstrate that the release of secretory products can occur during transient fusion events, which leads one to question the current models for membrane recycling. In the past year, strong evidence has emerged in support of a role for rab3 and Gα i3 proteins in regulating a putative scaffold of proteins that cause bilayer fusion during exocytosis. These findings parallel the biochemical identification of several new cytosolic, secretory vesicle and plasma membrane proteins that may also play a role in regulating fusion.

Neurotransmitter release through the exocytotic fusion pore

The exocytotic fusion pore transiently connects the secretory granule and plasma membrane during secretory vesicle fusion. Recent evidence on the properties of the fusion pore measured electrophysiologically suggest that the fusion pore is a lipid structure formed by a tension-driven fusion mechanism. We propose that a macromolecular scaffold of proteins directs and regulates the fusion process by drawing a characteristic plasma membrane dimple into close proximity with the secretory granule. Although the identity of the proteins making up the scaffold are largely unknown, studies on the regulation of exocytosis implicate the GTP binding protein Rab3 and unknown Ca2+ binding proteins as part of the scaffold, which appears to behave as a coincidence detector. Simultaneous measurement of fusion and the release of secretory products, along with some unusual properties of secretory granule matrices, have led to the unexpected conclusion that the release of secretory products from the matrix may also be regulated.

ECL Cells: Biology and Pathobiology

During recent years, the so-called ECL cells of the acid-producing part of the stomach have attracted much attention, mainly due to the fact that mice and rats were found to develop gastric carcinoids (ECL cell tumors) following life-long treatment with blockers of acid secretion. These observations touched off concern about the safety of the long-term clinical use of such drugs. The ECL cells are the predominant endocrine cell population in the oxyntic mucosa. They produce histamine, chromogranin A/pancreastatin and an as yet unidentified peptide hormone. They respond to gastrin by the release of secretory products; more long-term responses include adaptation to the gastrin stimulus, hypertrophy and hyperplasia. Intravenous infusion of maximally effective doses of gastrin promptly reduced the number of cytoplasmic vesicles in the ECL cells and their content of histamine and pancreastatin. Despite the ongoing infusion of gastrin, the number of vesicles and the content of histamine and pancreastatin were back to normal 4-6 h after the start of the infusion. The histidine decarboxylase (HDC) activity and HDC mRNA level increased progressively until plateaus were reached after 6-8 h of gastrin infusion. The size of the ECL cells started to increase about 4 days after the start of a subcutaneous gastrin infusion (resulting in half-maximally effective serum gastrin concentrations). The ECL cell size reached maximum after about 2 weeks and then remained at this level.The number of cytoplasmic vesicles was increased; this effect seemed to reach a maximum after 1-2 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)

Release of secretory products during transient vesicle fusion.

Patch-Camp experiments have shown that fusion of secretory granules with the plasma membrane does not always occur as an all-or-none event, but can develop slowly in a fluctuating manner or can be transient. These observations suggested that release could be detected during such incomplete fusion events. To test this hypothesis we have combined patch-clamp measurements of the activity of single exocytotic fusion pores in beige mouse mast cells with the electrochemical detection of serotonin released during the exocytotic events. We report here that on fusion pore opening there is a small release of serotonin which is directly proportional to the pore conductance. We also show that a significant release occurs during transient fusion events. These results demonstrate, to our knowledge for the first time, release of a neurotransmitter from a secretory vesicle that did not undergo complete fusion.

Mucin Exocytosis

Mucins produced by goblet cells of the respiratory mucosa are condensed while stored in secretory granules. Mucin condensation and its decondensation upon exocytosis can be explained by the theory of polymer gel phase transition. After the opening of a secretory pore, Ca2+ inside the granule is exchanged for extracellular Na+. Na/Ca exchange triggers a polymer gel phase transition whereby the mucin polymer matrix undergoes massive swelling and thereby changes from a condensed to a hydrated phase. Swelling of the granular content is driven by a Donnan potential and results in the release of secretory product and the formation of small mucin gels, which later anneal to each other to form the respiratory mucus. Because of the tangled rather than cross-linked topology of the mucin network, the rheologic properties of the respiratory mucus depend primarily on hydration. As mucins are polyionic, the hydration of mucus is controlled by a Donnan equilibrium. Hence, mucus hydration and rheology are determined by two factors: the quantity, chain length, and charge density of the secreted mucins, and the amount and the ionic and polyionic composition of the water transported across the respiratory mucosa.

Morphology and ultrastructure of the venom glands of the northern pacific rattlesnake Crotalus viridis oreganus.

The venom gland of Crotalus viridis oreganus is composed of two discrete secretory regions: a small anterior portion, the accessory gland, and a much larger main gland. These two glands are joined by a short primary duct consisting of simple columnar secretory cells and basal horizontal cells. The main gland has at least four morphologically distinct cell types: secretory cells, the dominant cell of the gland, mitochondria-rich cells, horizontal cells, and "dark" cells. Scanning electron microscopy shows that the mitochondria-rich cells are recessed into pits of varying depth; these cells do not secrete. Horizontal cells may serve as secretory stem cells, and "dark" cells may be myoepithelial cells. The accessory gland contains at least six distinct cell types: mucosecretory cells with large mucous granules, mitochondria-rich cells with apical vesicles, mitochondria-rich cells with electron-dense secretory granules, mitochondria-rich cells with numerous cilia, horizontal cells, and "dark" cells. Mitochondria-rich cells with apical vesicles or cilia cover much of the apical surface of mucosecretory cells and these three cell types are found in the anterior distal tubules of the accessory gland. The posterior regions of the accessory gland lack mucosecretory cells and do not appear to secrete. Ciliated cells have not been noted previously in snake venom glands. Release of secretory products (venom) into the lumen of the main gland is by exocytosis of granules and by release of intact membrane-bound vesicles. Following venom extraction, main gland secretory and mitochondria-rich cells increase in height, and protein synthesis (as suggested by rough endoplasmic reticulum proliferation) increases dramatically. No new cell types or alterations in morphology were noted among glands taken from either adult or juvenile snakes, even though the venom of each is quite distinct. In general, the glands of C. v. oreganus share structural similarities with those of crotalids and viperids previously described.