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.     

A Study of the Alimentary Canal of the Brachyopterygian Fish Polypterus senegalus with Electron Microscopy and Immunohistochemistry

The gastro-intestinal tract of Polypterus senegalus was investigated by means of electron microscopy and immunohistochemistry. Cilia-bearing cells can be observed over the whole length of the intestine. All enterocytes along the intestinal tract are characterized by apical pinocytotic vesicles. However, a typical intestinal region, which in other fish is characterized by large supranuclear vacuoles, is lacking. By means of electron microscopy, four types of endocrine cells and three types of nerve cell processes can be identified. By means of immunohistochemistry, endocrine cells with immunoreactivity for bombesin-, enkephalin-, G/CCK-, 5-HT-, somatostatin- and substance P-antisera can be found. Nerve cell processes show immunoreactivity for bombesin-, enkephalin-, 5-HT-, substance P- and VIP-antisera. The number of immunoreactive endocrine cells, nerve cells and nerve cell processes is different for each part of the gut.     

南方鲇头肾的组织学和超微结构

采用解剖学、组织学、组织化学方法,通过电镜观察,研究了南方鲇(Silurus meridionulis Chen)头肾的形态和超微结构。南方鲇左右头肾不相连,与肾明显分离。头肾组织包括血管系统、淋巴细胞聚集区、粒细胞聚集区、内分泌组织区。血管系统由头肾动脉、头肾静脉、后主静脉及其分枝所组成。淋巴区细胞染色深,紧密聚集成网状结构,主要包括大、小淋巴细胞等细胞类型。粒细胞区域狭窄,染色浅,主要为粒细胞聚集。内分泌组织区域细胞大型,常规染色极浅,包括肾间组织细胞、肾上组织细胞。肾间组织细胞有丰富的线粒体、内质网及分泌颗粒。肾上组织细胞主要分布于后主静脉及其分枝的周围、静脉壁中,细胞中有密集的分泌颗粒。肾上组织细胞和肾间组织细胞集中于头肾,内分泌功能增强,是与其运动能力强、快速捕食相适应的特征。     

Paraneuronal pseudobranchial neurosecretory cells in scorpion catfish Heteropneustes fossilis: an environment scanning electron microscope and transmission electron microscope study

Yadav, L., Sengar, M., Zaccone, D. and Gopesh, A. 2011. Paraneuronal pseudobranchial neurosecretory cells in scorpion catfish Heteropneustes fossilis: an environment scanning electron microscope and transmission electron microscope study. —Acta Zoologica (Stockholm) 00 : 1–8. Pseudobranchial neurosecretory system (PNS), found in the gill region of certain groups of teleosts, falls under the category of the ‘diffuse neuroendocrine system’ (DNES). The cells belonging to the system share morpho‐functional features with the paraneuronal cells observed in respiratory tract and airway surfaces of higher vertebrates. On the basis of the experimental observations, a role in condition of hypoxia has been recorded for this system. In an attempt to elucidate the ultrastructure of pseudobranchial neurosecretory cells, present investigation was undertaken using environment scanning electron microscope (ESEM) and TEM in an air‐breathing catfish, Heteropneustes fossilis. The external morphology of PNS under ESEM appeared as a mass of cells supplied with nerves and blood capillaries. Each cell mass is made up of numerous pear‐shaped neurosecretory cells, confirmed by neurosecretion‐specific acid violet stain. The TEM investigation of the cells revealed the presence of different sizes of dense‐cored vesicles in the cytoplasm, which was observed as granular cytoplasm under light microscope. Presence of large number of mitochondria in the cytoplasm confirmed active involvement of these cells in the physiology of fishes. Although lacuna prevails regarding the exact function of this system of fish, its probable role in hypoxic condition and surfacing behavior are speculated.     

Distribution of goblet and endocrine cells in the intestine: A comparative study in Amazonian freshwater Tambaqui and hybrid catfish

Goblet cells (GCs) and endocrine cells (ECs) play an important role in intestine physiology, and few studies currently exist for Amazonian fishes. This study aimed to quantify the distribution of GCs and ECs producing cholecystokinin-8 and neuropeptide Y, assessed by mucin histochemistry and peptides immunohistochemistry, in the intestine of two Amazonian species with different feeding habits Tambaqui (Colossosoma macropomum) and hybrid catfish (Pseudoplatystoma reticulatum × Leiarius marmoratus), an omnivore and carnivore, respectively. A systematic literature review correlating feeding habit and GC and EC distribution was also included to contribute to the comparative study. The results of this study provided novel information about the gut cells of Tambaqui and hybrid catfish. Both, GCs and ECs can be found sweeping the entire intestine of Tambaqui and hybrid catfish although the cells can be more concentrated in certain segments. The GCs and ECs in Tambaqui were more uniformly distributed in the midgut segments (T1, T2, and T3). Unlike, in hybrid catfish GCs were more concentrated in the hindgut (C4) and ECs mainly in the two midgut segments (C1 and C2) of hybrid catfish. Based on the comparison between Tambaqui, hybrid catfish, and other fishes in the literature review, we suggest that cell distribution can be partially explained by feeding habits, carnivorous vs. omnivorous.     

Electron microscopic study on the innervation of the pancreas of the domestic fowl

Summary The innervation of the pancreas of the domestic fowl was studied electron microscopically. The extrapancreatic nerve is composed mostly of unmyelinated nerve fibers with a smaller component of myelinated nerve fibers. The latter are not found in the parenchyma. The pancreas contains ganglion cells in the interlobular connective tissue. The unmyelinated nerve fibers branch off along blood vessels. Their synaptic terminals contact with the exocrine and endocrine tissues. The synaptic terminals can be divided into four types based on a combination of three kinds of synaptic vesicles. Type I synaptic terminals contain only small clear vesicles about 600 Å in diameter. Type II terminals are characterized by small clear and large dense core vesicles 1,000 Å in diameter. Type III terminals contain small clear vesicles and small dense core vesicles 500 Å in diameter. Type IV terminals are characterized by small and large dense core vesicles. The exocrine tissue receives a richer nervous supply than the endocrine tissue. Type II and IV terminals are distributed in the acinus, and they contact A and D cells of the islets. B cells and pancreatic ducts are supplied mainly by Type II terminals, the blood vessels by Type IV terminals.This work was supported by a scientific research grant (No. 144017) and (No. 136031) from the Ministry of Education of Japan to Prof. M. Yasuda     

Semicystic spermatogenesis and biflagellate spermatozoon ultrastructure in the Nile electric catfish Malapterurus electricus (Teleostei: Siluriformes: Malapteruridae)

Spermatogenesis and spermatozoon ultrastructure in the Nile electric catfish Malapterurus electricus are described using scanning and transmission electron microscopy. Although the testis organization conforms to the ‘unrestricted’ spermatogonial type, the species has a rare type of spermatogenesis not previously described among catfishes, ‘semicystic’, in which the cyst ruptures before the spermatozoon stage. Spermiogenesis also involves some peculiar features such as condensation of the chromatin in the posterior part of the nucleus to form a compact electron‐dense mass with some irregular electron‐lucent lacunae, while the uppermost part of the nucleus is a loose electron‐lucent area, absence of the nuclear rotation and, as a consequence, the centriolar complex and the initial segment of each flagellum arise directly in a position perpendicular to the basal pole of the nucleus, and occurrence of numerous vesicles in the midpiece. In addition, spermiogenesis includes migration of the diplosome and mitochondria to the basal pole of the nucleus, formation of two moderate nuclear fossae, each of which contains the centriolar complex, development of two independent flagella and elimination of the excess cytoplasm. The mature spermatozoon has a more or less round head with no acrosome or acrosomal vesicle, a long midpiece with numerous mitochondria and vesicles and two long tails or flagella having the classical axoneme structure of 9 + 2 microtubular doublet pattern and with no lateral fins and membranous compartment. These findings suggest that the ultrastructural features of spermiogenesis and spermatozoa of M. electricus are synapomorphies of types I and II spermiogenesis and spermiogenesis is closely similar to the type described in the Nile catfish Chrysichthys auratus.     

Fine structure of diffused pseudobranchial neurosecretory cells associated with carotid labyrinth in an air‐breathing catfish Clarias batrachus

Fish are known to have branchial chemoreceptors and even extrabranchial chemoreceptors to meet the challenges of aquatic environment. The pseudobranchial neurosecretory system associated with carotid labyrinth (CL) is one such example. CL – a chemosensory organ is well known in amphibians. The homologous structure also exists in fish. Clusters of neurosecretory cells, close to the CL and the first two efferent branchial arteries occur in catfish and a few other groups of teleosts. These cells belong to the pseudobranchial neurosecretory system (PNS). To reveal the ultrastructure of CL and the pseudobranchial neurosecretory cells (PNSCs), environmental scanning electron microscope (ESEM) and transmission electron microscope (TEM) investigations were made in an Asian air‐breathing catfish Clarias batrachus. Under ESEM, the PNS appeared as a mass of cells innervated by nerves and supplied by blood capillaries. The CL appeared to have a network of blood capillaries. The transmission electron microscopic investigations showed pear shaped PNSCs having different sizes of dense cored vesicles (DCVs), numerous mitochondria, nerve varicosities, indicating a secretory function of the cells. The CL shows a close association with PNSCs and smooth muscles. Although the exact function of the CL and associated PNSCs in the biology of fish is far from clear, their morphology suggests they are involved in a stress response such as to hypoxia.     

Immunohistochemically detected ontogeny of prolactin and growth hormone cells in the African catfish Clarias gariepinus

The chronological appearance of selected endocrine cells in the pituitary of African catfish Clarias gariepinus (Burchell, 1822) was studied morphologically, histologically and immunohistochemically by using antisera raised against catfish growth hormone (cgGH) and recombinant tilapia prolactin I (tPRL). cgGH- and tPRL-like immunoreactive cells were visible from day 1 post fertilisation (hatching) throughout the juvenile and the adult stage. From 1 to 90 days after hatching, the larval pituitary is oval in shape with a distinctly shaped rostral pars distalis, proximal pars distalis and pars intermedia. From day 120 onwards allometric growth of the rostral and proximal pars distalis extended the prolactin and growth hormone cells anteriorily and posteriorily, respectively. Size and activity of the prolactin and growth hormone cells, measured by the ratio of cell surface to nuclear surface remained constant until day 40 and showed a growth spurt thereafter. Growth hormone content, measured with a catfish-specific radio-immunoassay from hatching until 60 h post hatching, increased exponentially between 30 and 60 h.     

The Pituitary Gland of the Roach Leuciscus rutilus

Three endocrine cell types were recognized in the proximal pars distalis (PPD) of the roach, Leuciscus rutilus, by electron microscopy. Two of these cell types are basophilic and assumed to be gonadotropic. The third cell type is acidophilic and its similarity to somatotropic (STH) cells is evident. Green fluorescent cells corresponding to cells with dense core vesicles of about 85 nm are scattered between the PPD and the proximal neurohypophysis (PNH). Their possible content of a catecholamine was demonstrated with microspectrofluorometric analyses. Four axon types with granules 65, 81, 110 and 137 nm in average diameter terminate at the basement membrane. Only fibers with granules of 81 nm enter the PPD to make synapses with the STH cells. Extensions of the basement membrane are continuous with the perivascular space and penetrate the endocrine tissue. It is postulated that axon terminals on the basement membrane are functionally equivalent to synaptic contacts on the endocrine cells. Axon type 4 has granules which in sizes correspond to those of cells in lateral and ventrolateral parts of the nucleus lateralis tuberis (NLT). A similar correlation is possible between axon type 3 and other cells in the NLT. This is in accordance with earlier conclusions of a regulation of the gonadotropic cells from the lateral (posterior) parts of the NLT.     

Ontogenesis and ultrastructure of seminal vesicles of the catfish,Clarias gariepinus

The ontogenesis and structural characteristics of the seminal vesicles in Clarias gariepinus (sharptooth catfish) were studied by light and electron microscopy and are described in detail. The seminal vesicles, beginning as simple protrusions from the vas efferentia, becomes more complex with age. Their distal ends become fingerlike and the bases form palm-like extensions. Juvenile male organs do not reveal any signs of seminal vesicles although spermatogenic tissue is already well delineated. The developing gonads contain clusters of large cells, close to the sperm duct and cysts of the testis, from which seminal vesicles are formed. Secretory epithelium lines the tubules of the seminal vesicles and becomes columnar as the tissue matures. Electron micro-graphs of these epithelial cells reveal two types of cells: opaque cells and cells with very vacuolized cytoplasm. Dense pinocytotic vesicles are present between the membranes of neighbouring seminal tubules and apical cell membranes facing the lumen. Maturation and onset of secretion by the secretory cells is accompanied by morphological changes. Protruding cylindrical cells become shortened, modified to cuboidal, rounded cells that send tubular extensions into the lumen. In the final stage of differentiation, only connective tissue membranes supporting the tubule walls remain intact. At the points of contact between the testis, seminal vesicles, and sperm duct, the epithelia of these organs often become confluent. The distal parts of the seminal vesicles, rarely contain sperm; during spawning sperm accumulated in the proximal tubules of the vesicles. © 1994 Wiley-Liss, Inc.     

An Ultrastructural and Immunocytochemical Study of Gastrodermal Cell Types in Microstomum lineare (Turbellaria,Macrostomida)

The gastrodermal cell types of Microstomum lineare (Turbellaria, Macrostomida) were studied by electron microscopy. Their immunoreactivity (IR) to bovine pancreatic peptide (BPP), FMRF-amide and vasotocin, somatostatin, neurotensin, ACTH, CCK, bombesin, secretin, gastrin/CCK and insulin antisera was tested by light microscopic immunocytochemical methods. In addition to granular club cells and phagocytic cells, neurons and neoblasts occur in the gastroderm of this turbellarian species. This is the first observation of neurons in the gastroderm of a flatworm. Dense-core vesicles (70–100 nm diameter), electron lucent cytoplasm and numerous Golgi complexes characterize the neurons. Unpolarized two-way synapses, neuromuscular junctions and polarized chemical synapses can be observed in the gastroderm. Neoblasts with large nuclei and scanty cytoplasm and differentiating cells containing clusters of basal bodies occur next to the basal lamina of the gastroderm. BPP-like, FMRF-amide-like and vasotocin-like immunoreactivity is demonstrated in the gastroderm. Both BPP and FMRF-amide IR is restricted to the basal cytoplasm of the granular club cells, while a different location for IR to vasotocin antiserum is observed. The status of the neuronal cell in the gastroderm of M. lineare is discussed in relation to endocrine (paracrine) cells and neurons in the gastroderm of invertebrates.     

Susceptibility of European catfish (Silurus gianis) to Edwardsiella ictaluri

European catfish (Silurus glanis) were tested for their susceptibility to the bacterium Edwardsiella ictaluri. The LD₅₀ of E. ictaluri when injected into European catfish was 5.4 × 10⁶ compared to 7.1 times; 10⁴ for channel catfish (Ictalurus punctatus). E. ictaluri was isolated from dead and moribund European catfish and the bacterium was also detected in kidney smears by an indirect fluorescent antibody (FA) technique. The bacterium was not isolated or detected by FA from surviving fish 15 days after injection. No clinical signs of E. ictaluri infection were noted in European catfish, but these were prevalent in the channel catfish. These experiments indicate that under experimental conditions European catfish are not as susceptible to E. ictaluri as channel catfish.     

Identification and characterization of a FasL-like protein and cDNAs encoding the channel catfish death-inducing signaling complex

To elucidate cytolytic mechanisms in the channel catfish, lysates from catfish lymphoid and fibroblast cell lines were screened by Western blot analysis using a panel of antibodies reactive with components of the mammalian apoptotic pathway. Strong reactivity with three proteins (approximate M_r 70,000, 37,000, and 15,000) was seen using an antibody targeted to mammalian Fas ligand (FasL). The sizes of the two smaller proteins are consistent with their tentative designation as membrane-bound (37,000 M_r) and soluble (15,000 M_r) FasL. Treatments known to induce FasL in mammalian systems (e.g., PMA/calcium ionophore, UV-irradiation) induced expression of the 37,000-M_r protein in catfish T-cell lines. Moreover, expression of the 37,000-M_r protein in clonal T cells was up-regulated by increasing cell density. At the nucleotide level, homologues of Fas receptor (FasR), FADD, and caspase 8 were identified and characterized. These gene products likely constitute the teleost equivalent of the death-inducing signaling complex (DISC). FADD was constitutively expressed in all (T, B, macrophage, and fibroblast) cell lines examined as well as in peripheral blood lymphocytes (PBL), whereas FasR and caspase 8 were expressed in all cell lines except CCO, a FasL-positive fibroblast line. In contrast to FasL, expression of FasR and caspase 8 was inversely proportional to cell density. Collectively these studies identified four membrane-proximal proteins involved in the initiation of apoptosis in channel catfish and suggest that mechanisms of cell-mediated cytotoxicity in teleosts are similar to those used by mammals.     

Amino Acid-Activated Channels in the Catfish Taste System

Membrane vesicles derived from external taste epithelia of channel catfish (Ictalurus punctatus) were incorporated into lipid bilayers on the tips of patch pipettes. Consistent with previous experiments (Teeter, J. H., J. G. Brand, and T. Kumazawa. 1990. Biophys. J. 58:253–259), micromolar (0.5–200 μM) concentrations of l-arginine (l-Arg), a potent taste stimulus for catfish, activated a nonselective cation conductance in some bilayers, which was antagonized by d-Arg. Two classes of l-Arg-gated receptor/channels were observed in reconstituted taste epithelial membranes: one with a unitary conductance of 40–60 pS, and the other with a conductance of 75–100 pS. A separate class of nonselective cation channels, with a conductance of 50–65 pS, was activated by high concentrations of l-proline (l-Pro) (0.1–3 mM), which is the range necessary to elicit neural responses in catfish taste fibers. The l-Pro-activated channels were not affected by either l- or d-Arg, but were blocked by millimolar concentrations of d-Pro. Conversely, neither l- nor d-Pro altered the activity of either class of l-Arg-activated channels, which were blocked by micromolar concentrations of d-Arg. These results are consistent with biochemical, neurophysiological, and behavioral studies indicating that taste responses of channel catfish to l-Arg are mediated by high-affinity receptors that are part of or closely coupled to nonselective cation channels directly gated by low concentrations of l-Arg, while responses to l-Pro are mediated by distinct, low-affinity receptors also associated with nonselective cation channels.     

Distinct Cell Clusters Touching Islet Cells Induce Islet Cell Replication in Association with Over-Expression of Regenerating Gene (REG) Protein in Fulminant Type 1 Diabetes

Background

Pancreatic islet endocrine cell-supporting architectures, including islet encapsulating basement membranes (BMs), extracellular matrix (ECM), and possible cell clusters, are unclear.

Procedures

The architectures around islet cell clusters, including BMs, ECM, and pancreatic acinar-like cell clusters, were studied in the non-diabetic state and in the inflamed milieu of fulminant type 1 diabetes in humans.

Result

Immunohistochemical and electron microscopy analyses demonstrated that human islet cell clusters and acinar-like cell clusters adhere directly to each other with desmosomal structures and coated-pit-like structures between the two cell clusters. The two cell-clusters are encapsulated by a continuous capsule composed of common BMs/ECM. The acinar-like cell clusters have vesicles containing regenerating (REG) Iα protein. The vesicles containing REG Iα protein are directly secreted to islet cells. In the inflamed milieu of fulminant type 1 diabetes, the acinar-like cell clusters over-expressed REG Iα protein. Islet endocrine cells, including beta-cells and non-beta cells, which were packed with the acinar-like cell clusters, show self-replication with a markedly increased number of Ki67-positive cells.

Conclusion

The acinar-like cell clusters touching islet endocrine cells are distinct, because the cell clusters are packed with pancreatic islet clusters and surrounded by common BMs/ECM. Furthermore, the acinar-like cell clusters express REG Iα protein and secrete directly to neighboring islet endocrine cells in the non-diabetic state, and the cell clusters over-express REG Iα in the inflamed milieu of fulminant type 1 diabetes with marked self-replication of islet cells.     

Evaluation of an in vitro cell assay to select attenuated bacterial mutants of Aeromonas hydrophila and Edwardsiella tarda to channel catfish

Aims: To evaluate the feasibility of using an in vitro cell assay to select attenuated bacterial mutants. Methods and Results: Using catfish gill cells G1B, the feasibility of using an in vitro assay instead of in vivo virulence assay using live fish to select attenuated bacterial mutants was evaluated in this study. Pearson correlation analysis between in vitro virulence to G1B cells and in vivo virulence of Aeromonas hydrophila and Edwardsiella tarda revealed that there was a significant correlation between the two (r = ?0·768, P value = 3·7 × 10^?16). Conclusions: The in vitro cell assay might be initially used to screen large quantities of bacteria to select attenuated mutants of catfish pathogens. Significance and Impact of the Study: The in vitro cell assay using catfish gill cells to identify attenuated mutants of catfish pathogens will reduce cost involved in the in vivo virulence assay that requires many fish and aquariums.     

Nutritive phagocytosis in animal cells

Summary Nutritive phagocytosis in the hydroid Clava squamata was studied with the electron microscope, using carbon particles of 0.6 as an indicator.An early step in phagocytosis is the transformation, in many cells, of the free border from a type with cylindrical microvilli to one with a complicated system of cytoplasmic folds.Particles fixed at the actual stage of ingestion are found (a) between two cytoplasmic folds, (b) between a fold and a relatively straight portion of the cell surface, or (c) in a depression of an otherwise straight portion of the cell surface.Ingested carbon particles were always found enclosed by a membrane, with a layer of moderate electron density between the carbon particle and the membrane.The ingested carbon particles are localized apically in small vesicles each containing one particle (interpreted as primary phagocytic vesicles) or at deeper levels of the cell, in larger vesicles containing many carbon particles (interpreted as secondary phagocytic vesicles).Other cytoplasmic changes during phagocytosis relate to the distribution of mitochondria and the occurence and distribution of flattened vesicles of a characteristic appearance.With the technical assistance of Birgitta af Burén.Financial support from Swedish Natural Science Research Foundation is gratefully acknowledged.     

Granular “endocrine” cells in avian respiratory epithelia

Summary An electron microscopic investigation of the extrapulmonary respiratory tract of embryos and chick of the domestic fowl (Gallus domesticus) has demonstrated for the first time in birds the presence here of a small number of epithelial cells characterised by an aminecontaining type of granule. These granular cells were scattered singly throughout the trachea, syrinx and primary bronchi and seemed more numerous in the caudal part of the airway. In favourable planes of section a small part of the cell was in contact with the luminal surface of the epithelium. The characteristic granular vesicles (approximate diameter 140 nm) appeared to be randomly distributed in the cytoplasm and there was no concentration of vesicles close to the plasma membrane. One of the cells was closely associated with an intraepithelial axon. By fluorescence microscopy, a small number of cells with a similar shape and distribution to the granular cells was observed after administration of 3,4-dihydroxyphenylalanine which may indicate the presence of an amine handlign mechanism in these cells. It is suggested that the granular cells belong to the APUD system of endocrine cells and that they may be modulated by the concentration of gas in the airways.     

The ultrastructure of the midgut glands inLigia italica (Isopoda) under different nutritional conditions

After a period of food deprivation,Ligia italica were refed for 2 days with different diets and their midgut glands were examined under the electron microscope with special reference to the large cells. The predominant features are the following: extended glycogen fields after sucrose-diet; numerous lipid droplets and peroxisome-like vesicles after lipid-diet (butter); swollen mitochondria and a great number of pinocytotic vesicles after protein diet (curds); electron dense vesicles and myelin bodies after the uptake ofEscherichia coli. In contrast to amphipods, the intertidal isopodL. italica is not able to digest cellulose, as the cell ultrastructure exhibits all features of starved animals, as well as that following feeding with lignin.Supported by Deutsche Forschungsgemeinschaft (Sto 75/4–9)