Formation of junctional complexes at sites of contact of hemocytes with tissue elements in degenerating nerves of the crayfish, Orconectes virilis

Hemocytes, which contain large cytoplasmic granules, invade the multilamellate glial sheath of ventral ganglion nerve roots of the crayfish following surgical interruption of these nerves. Electron microscopic examination of sections of plasticembedded tissues and replicas of freeze-cleaved ganglion roots reveals numerous slender cytoplasmic extensions of the hemocytes present in damaged nerve sheaths. Many of these microvillous extensions contact glial cells and filamentous extracellular masses. At sites of contact, the microvilli are flattened and occasionally electron-dense material is present in the hemocyte cytoplasm subjacent to the plasma membrane that is closely apposed to a glial cell or connective tissue. Intramembranous surfaces of hemocyte plasmalemmae exposed by freeze-fracture, exhibit particle aggregates 700–2500 Å in diameter. Individual particles are 95–105 Å in diameter. Since the particle aggregates correspond in overall dimension and position in the cell to the sites of contact of hemocyte processes with other sheath components, it is assumed that the two structures are equivalent and represent a junctional complex very similar in structure to some hemidesmosomes. Results from this study strongly suggest that granulated crustacean hemocytes, in response to surgical injury of nerves, invade the damaged nerve sheath and identify damaged glial cells and connective tissue by forming slender cytoplasmic processes which contact elements of the sheath. Tissue components contacted by the hemocytes may subsequently be phagocytosed by them. This is the first report of an invertebrate hemocyte-mediated response to tissue damage in which evidence is presented that the hemocyte may identify necrotic cells and extracellular matrix by forming junctional complexes with them. Crustacean hemocytes, therefore, are likely much more complex functionally than has been previously estimated.     

Comparative studies of the defense mechanism against Schistosoma japonicum of schistosome-susceptible and -resistant Oncomelania nosophora

The amphibious snail Oncomelania nosophora is an intermediate host of Schistosoma japonicum. Previously we reported that there are two strains of the snail, one resistant and one susceptible to a Mindoro, the Philippines, strain of S. japonicum. The resistant snails were collected from Nirasaki and susceptible snails from Kisarazu, Japan. To determine early cellular responses in the two snail strains, we examined histologic alterations in the snails for up to 18 h after the initial exposure to miracidia. In both strains, the penetrating miracidia were distributed in the foot, mantle, gills, heart, stomach, and kidney, and the mean number of penetrating miracidia was similar in both strains. After penetration, snail hemocytes migrated toward the larvae, and by 12 h after exposure, substantial numbers of penetrated larvae were surrounded and encapsulated by hemocytes. The percentage of larvae encapsulated by hemocytes during 12-18 h after the exposure was significantly higher in the resistant Nirasaki strain (60.9+/-19.8%) than in the susceptible Kisarazu strain (42.3+/-15.0%). In a few snails of the Nirasaki strain, all the larvae found were encapsulated by hemocytes. The differences in hemocyte responses between the two strains may explain the susceptibility of the snails to schistosome larvae.     

Structure of hematopoietic nodules in the ridgeback prawn,Sicyonia ingentis: Light and electron microscopic observations

The architecture and fine structure of the epigastric hematopoietic nodules of the ridgeback prawn, Sicyonia ingentis, are described. The nodules consist of a highly branched series of tubules that contain the maturing hemocytes within a connective tissue stroma. Hemocytes can exit the hematopoietic nodules by penetrating through fenestrations in the endothelial cell layer into the central hemal space or by migrating through the outer later of capsular cells and associated collagen fibrils. Four hemocyte categories were observed: agranular, small granule with cytoplasmic deposits, small granule without cytoplasmic deposits, and large granule hemocytes. This classification was based upon the presence, size, and type of cytoplasmic granules and the presence of cytoplasmic deposits. Only agranular cells and small granule hemocytes without cytoplasmic deposits appeared capable of division. Intermediate stages were observed between agranular hemocytes and small granule hemocytes with deposits and between small granule hemocytes without deposits and large granule hemocytes, suggesting existence of two distinct hemocyte lines.     

Schistosoma mansoni modulation of phagocytosis in Biomphalaria glabrata

Both short-term (3 hr) exposure of Biomphalaria glabrata snails (M-line and 13-16-R1) to Schistosoma mansoni (PR1) miracidia and in vitro incubation of parasite sporocysts with host hemolymph components altered host phagocytic ability. Hemocytes obtained from susceptible (M-line) snails that had been exposed to parasite miracidia for 3 hr showed reduced levels of phagocytosis of yeast cells in vitro compared to hemocytes from unexposed individuals. Incubation of whole hemolymph with sporocysts in vitro also reduced yeast phagocytosis in this susceptible strain. In contrast, resistant (13-16-R1) hemocytes showed increased levels of yeast phagocytosis after in vitro incubation with the parasite, and the opsonic properties of 13-16-R1 plasma were greater after exposure of snails to miracidia. These strain-specific effects of S. mansoni on host hemocyte phagocytosis and plasma opsonization were seen only when both plasma and hemocytes were present at the time of exposure to the parasite.     

Tissue responses exhibited by Biomphalaria alexandrina snails from different Egyptian localities following Schistosoma mansoni exposure

Snails’ susceptibilities to infection with Schistosoma mansoni were determined through observation of infection rates, total cercarial production and tissue responses of the first generation (F1) of Biomphalaria alexandrina snails, originally collected from different Egyptian governorates (Giza, Fayoum, Kafr El-Sheikh, Ismailia and Damietta) and responses were compared between groups. The emergence of cercariae for a 3-month period and the calculation of survival and infection rates, in control (Schistosome Biological Supply Center; SBSC) and infected snails were evaluated. SBSC and Giza snails showed greater susceptibilities to infection and lower mortality rates. In addition, at 6 and 72 h post-exposure to miracidia all the snail groups showed no difference in the anatomical locations of sporocysts. The larvae were found in the head-foot, the mantle collar and the tentacles of the snails. Sporocysts showed normal development with low tissue reactions in SBSC and Giza snail groups infected with S. mansoni miracidia (SBSC). However, in Fayoum, Kafr El-Sheikh, Ismailia and Damietta snail groups, variable tissue responses were observed in which numerous hemocytes made direct contact with S. mansoni larvae forming capsules. The results suggested that, different responses of B. alexandrina snail’s hemocytes towards S. mansoni are related to the degree of susceptibility of these snails. So this is important in planning the strategy of schistosomiasis control.     

The ultrastructure of hemocytes inDactylopius confusus (Cockerell), and the role of granulocytes in the synthesis of cochineal dye

Summary The ultrastructural study of free circulating hemocytes in the adult cochineal scale,Dactylopius confusus (Cockerell), demonstrated five cell types: prohemocytes, typical granulocytes (T-granulocytes), oenocytoids, plasmatocytes, and granulocytes with modified sub-cellular structure to perform a special synthetic and secretory function, which we refer to as modified granulocytes (M-granulocytes). Prohemocytes showed undifferentiated sub-cellular structure of the basic stem cell type (i.e., high cytoplasmic density with numerous ribosomes, centrally located large nucleus with a distinct nucleolus, and poorly developed endoplasmic reticulum). The commonly observed typical granulocytes (T-granulocytes) had several smooth endoplasmic reticulum (SER) with dilated cisternae and many SER-derived membrane bounded granules of different sizes and electron density. Oenocytoids were identified by the presence of many crystals, RER-originated fine secretory granules, and an eccentric nucleus. Plasmatocytes were easily characterized by their variable shapes and irregular outline with pseudopodia-like cytoplasmic extensions, possession of an elongated lobed nucleus, multivesicular bodies, RER-derived membrane bounded, electron-dense, lysosomelike vacuoles, well-developed SER cisternae, and numerous pinocytic and SER-originated vesicles of different sizes along the peripheral region. M-granulocytes comprised the largest proportion of hemocytes in all samples observed. M-granulocytes were distinguished not only by the presence of membrane bounded granules of different sizes and electron density, but by the possession of large nuclei with distinct nucleoli, many mitochondria, and a highly developed network of rough endoplasmic reticulum (RER). M-granulocytes had abundant, rosette-shaped, RER-derived chains of fine secretory granules, which accumulated in the cytoplasm and vacuoles, and were ultimately deposited into the hemolymph by exocytosis. These fine granules gave a positive result with periodic acid-Schiff (PAS) test. Based on RER-synthesized fine secretory granules (M-granulocytes), their ultimate deposition into hemolymph, the red pigmentation of hemolymph, positive PAS histochemical test of these granules, and the high population of these hemocytes, no such cell type has been described in previous studies in insects. The sub-cellular structure of the granulocyte in this insect has been modified to perform a special synthetic and secretory function (i.e., possibly the synthesis of the red pigment found in hemolymph, which has been the source of commercially important cochineal dye).Abbreviations EM electron microscope - ER endoplasmic reticulum - LM light microscopy - MVB multivesicular body - PAS periodic acid-Schiff - RER rough endoplasmic reticulum - SER smooth endoplasmic reticulum - SG secretory granules - TEM transmission electron microscopy - UA uranyl acetate     

Hemocytes of Biomphalaria glabrata: Factors affecting variability

Variability in the hemocyte number of two geographic strains of Biomphalaria glabrata was studied. In each strain a logarithmic increase in hemocyte number associated with increasing shell size was observed. A two fold increase in circulating hemocytes occurred 2 hr following the exposure of a susceptible strain of B. glabrata to miracidia of Schistosoma mansoni. The hemocyte number was dependent on the temperatures at which the snails were maintained.     

Ultrastructural description of the spore maturation stages of the clam parasite Minchinia tapetis (Vilela, 1951) (Haplosporida: Haplosporidiidae)

The fine structure of maturing spores of a haplosporidian parasite found in the gill, mantle and foot tissues of Ruditapes decussatus L. (Mollusca, Bivalvia), a species of commercial importance in Portugal, is described. When observed free in suspension, immature spores exhibit one or two epispore cytoplasmic extensions (ECE) which constitute a projection of a portion of the exosporoplasm, sometimes without ultrastructural organisation, surrounded by the plasmalemma. Free spores observed by light microscopy (LM) after 3-5 days of incubation in filtered sea-water exhibit no ECE attached to the spore wall. The mature spore is ovoid to ellipsoid, operculate, uninucleate and measures c. 4.8 microm long and c. 3.9 microm wide. The spore shape and size and the identity of the host living in the same geographical region suggest that this species is the same as previously described using LM observations as Haplosporidium tapetis Vilela, 1951 and later transferred to Minchinia Labbé, 1896.     

In vitro effect of larval Schistosoma mansoni excretory-secretory products on phagocytosis-stimulated superoxide production in hemocytes from Biomphalaria glabrata

The in vitro production of the reactive oxygen metabolite superoxide (O2-) was confirmed in hemocytes from the schistosome intermediate host Biomphalaria glabrata. Active forms of the enzyme superoxide dismutase (SOD) inhibited reduction of nitroblue tetrazolium (NBT) to formazan in cells that had phagocytozed zymosan particles, whereas an inactivated form of SOD did not. Moreover, based on the prevalence of O2(-)-positive hemocytes and the relative intensity of NBT staining reactions, hemocytes from the Schistosoma mansoni-resistant 10-R2 strain of B. glabrata possessed an overall greater capacity for generating superoxide than did those from S. mansoni-susceptible M-line snails. Schistosoma mansoni excretory-secretory (E-S) products, released during in vitro transformation of miracidia to sporocysts, inhibited phagocytosis of zymosan particles and superoxide activity in hemocytes from both snail strains, but 10-R2 hemocytes maintained higher levels of phagocytosis and superoxide production than did M-line hemocytes. The dose-dependent decreases in phagocytosis observed in both snail strains in the presence of E-S products could not account fully for the concomitant decrease in superoxide levels detected, indicating that either a single E-S factor differentially affects phagocytosis and superoxide production, or that different E-S factors are involved in the specific interference of each of these hemocyte functions.     

Ultrastructure of enterochromaffin cells and associated neural and vascular elements in the mouse duodenum

Summary Enterochromaffin cells of adult mouse duodenum were studied with light- and electron-microscopical techniques. They were distinguished from other enteroendocrine cells by their pleomorphic, electron-dense secretory granules in the basal cytoplasm. At the apices of enterochromaffin cells, tufts of short microvilli bordered the gut lumen. At their bases, irregular cytoplasmic extensions were either in contact with or passed through the basal lamina. The presence of cytoplasmic extensions in close proximity to fenestrated capillaries and subepithelial nerves suggested an endocrine or paracrine function. Electron micrographs of serial thin sections were used to reconstruct an enterochromaffin cell from the crypt epithelium in three dimensions and to determine its relationship with the underlying neural plexus. Although extensions from the serially sectioned and reconstructed cell and other enterochromaffin cells studied in crypt epithelia protruded through the basal lamina, no synaptic contacts were seen. Evidence of a synaptic contact between a neurite and another type of enteroendocrine cell (possibly an intestinal A cell), suggested a neurocrine role for some of the basally-granulated cells. Possible functions of enterochromaffin cells are discussed in the light of recent literature on the system of enteroendocrine cells, also known as APUD (amine precursor uptake and decarboxylation) cells and/or paraneurons.     

Effect of miracidial dose on adoptively transferred resistance to Schistosoma mansoni in the snail intermediate host, Biomphalaria glabrata

Adoptively transferred resistance to Schistosoma mansoni in the snail intermediate host Biomphalaria glabrata was measured as a function of miracidial challenge dose. Schistosome-susceptible snails implanted with the amebocyte-producing organ (APO) from resistant donors showed 29 and 39% prevalences of infection after challenge with 5 and 10 miracidia, respectively, but 68-83% prevalences when exposed to 25-200 miracidia. Prevalences in control (untampered) susceptible snails ranged from 97 to 100% at the different miracidial doses. Higher infection prevalences at elevated doses suggest that a range of transferred resistance occurs and possibly that low levels of APO-derived plasma factors or hemocytes in some recipients can be overwhelmed by larger numbers of parasites.     

Observations on the process of wound repair in penaeid shrimp

The Petersen disk tag is a standard mark for penaeid shrimp, and attachment of the tag involves the insertion of a stainless steel pin through the shrimp's abdomen, resulting in a relatively large puncture wound. The wound healing process first observed at 24 hr post-tagging showed a pronounced hemocytic infiltration of the wound area. Hemocytes in contact with the pin became fusiform, began adhering to one another, and formed several concentric layers around the pin. Scattered foci of bacteria or nercrotic tissue in the vicinity of the would also became encapsulated by concentric layers of fusiform hemocytes, thereby forming nodules. Melanin appeared in association with the layers of hemocytes nearest the pin and in the nodules. Hemocytic infiltration was lollowed by the appearance of fibrocytes and the deposition of collagenlike fibers along the would channel 48 hr after wounding. Involution of epidermis and consequential cuticular involution into the would channel began at 96 hr after wounding. Complete epidermal and cuticular formation along the wound occurrd by 384 hr post-tagging.     

On the origin of the Biomphalaria glabrata hemocytes

A histologic, morphometric and ultrastructural study performed on Biomphalaria glabrata submitted to infection with Schistosoma mansoni miracidia failed to provide significant evidences that the so-called amebocyte-producing organ (APO) is really the central organ for hemocyte production. In infected snails no general reactive changes appeared in the APO, the mitoses were seen only occasionally, and the possibility of cellular hyperplasia was ruled out by morphometric measurements. Under the electron microscope the APO cells presented an essentially epithelial structure, without features indicative of transition toward hemocytes. On the other hand, the present findings pointed to a multicentric origin for the mollusc hemocytes, as earlier studies had indicated. Dense foci of hemocyte collections appeared sometimes around disintegrating sporocysts and cercariae in several organs and tissues of the infected snails, including a curious accumulation of such cells inside the ventricular cavity of the heart. In the heart and other sites, features suggestive of transformation of vascular space endothelial lining cells into hemocytes were apparent. To some extent, the postulated multicentric origin for B. glabrata hemocytes recapitulates earlier embryologic findings in vertebrates, when mesenchymal vascular spaces generate the circulating and phagocytic blood cells.     

Elaboration and ultrastructural changes in the pore canal system of the mineralized cuticle of Carcinus maenas during the moulting cycle

Two basic structural components are concerned in the elaboration of the pore canal system in the mineralized cuticle of the decapod crab Carcinus maenas: tubular cytoplasmic extensions originating from epidermis and vertical fibres. These components are present from the moment the first procuticular materials of the new cuticle are laid down but their organization varies according to a precise schedule during the further moult cycle stages. Cytoplasmic extensions form a complicated branching system connecting the epidermal layer with all regions of the cullcular compartment, at least transitorily. During the moult cycle the prolongation of this cellular system appears to result from two concomitant but opposite phenomena. Before ecdysis the growth of cell extensions in the proximal cuticular layers prevails over their regression at the distal level. During the post-moult period these phenomena are reversed in importance so that the pore canal system is without cytoplasmic material as soon as intermoult starts. The depositing of vertical fibres takes place in close contact with the proximal cell extension plasma membrane, which never bears dense plaques. As moult stages progress, they are gradually organized into twisted sheaths that persist throughout the intermoult. Incidentally, some fibres invade the pore canal lumen freed from cell extensions. Some aspects regarding the fine organization, the chemical composition and the functional significance of both epidermal tubular extensions or vertical fibres are also discussed in the light of previous investigations carried out on crustaceans and in other arthropods.     

Stretch-activated neuronal pathways to longitudinal and circular muscle in guinea pig distal colon

The role of the longitudinal muscle (LM) layer during the peristaltic reflex in the small and large intestine is unclear. In this study, we have made double and quadruple simultaneous intracellular recordings from LM and circular muscle (CM) cells of guinea pig distal colon to correlate the electrical activities in the two different muscle layers during circumferential stretch. Simultaneous recordings from LM and CM cells (<200 microm apart) at the oral region of the colon showed that excitatory junction potentials (EJPs) discharged synchronously in both muscle layers for periods of up to 6 h. Similarly, at the anal region of the colon, inhibitory junction potentials (IJPs) discharged synchronously in the two muscle layers. Quadruple recordings from LM and CM orally at the same time as from the LM and CM anally revealed that IJPs occurred synchronously in the LM and CM anally at the same time as EJPs in LM and CM located 20 mm orally. Oral EJPs and anal IJPs were linearly related in amplitude between the two muscle layers. Spatiotemporal maps generated from simultaneous video imaging of the movements of the colon, combined with intracellular recordings, revealed that some LM contractions orally could be correlated in time with IJPs in CM cells anally. N(omega)-nitro-L-arginine (L-NA; 100 microM) abolished the IJP in LM, whereas a prominent L-NA-resistant "fast" IJP was always observed in CM. In summary, in stretched preparations, synchronized EJPs in both LM and CM orally are generated by synchronized firing of many ascending interneurons, which simultaneously activate excitatory motor neurons to both muscle layers. Similarly, synchronized IJPs in both LM and CM anally are generated by synchronized firing of many descending interneurons, which simultaneously activate inhibitory motor neurons to both muscle layers. This synchronized motor activity ensures that both muscles around the entire circumference are excited orally at the same time as inhibited anally, thus producing net aboral propulsion.     

中国鲎和圆尾鲎血淋巴细胞分类和特征的比较研究

为了更好地了解中国鲎(Tachpleus tridentatus)和圆尾鲎(Carcinoscorpius rotundicauda)血淋巴细胞的种类组成和特征差异,综合运用光学显微镜、扫描电镜和粒度仪,较为系统地对两种鲎的血淋巴细胞进行了分类和特征研究,从而为两种鲎的血淋巴细胞和分子生物学研究提供基础资料。根据血淋巴细胞大小、核质比、细胞着色特点、细胞中颗粒存在与否、颗粒的密集程度等,中国鲎和圆尾鲎的血淋巴细胞均可分为大颗粒细胞、小颗粒细胞和透明细胞三种主要类型,且两种鲎的血淋巴细胞均以颗粒细胞为主,透明细胞在血淋巴细胞中所占比例最小,但具有高核质比。两种鲎的同类血淋巴细胞在染色和形态上无显著性差异,但在同一种鲎中,血淋巴细胞密度存在显著的雌雄差异。       

The fine structure of the cysticercoid of Hymenolepis diminuta. III. The scolex.

Transmission electron microscopy of the scolex of the 8-day-old Hymenolepis diminuta cysticercoid demonstrates its resemblance to the scolex of the adult. A syncytial tegument composed of external and internal layers is connected by cytoplasmic extensions. Fully developed microtriches are present. Furthermore, a basement membrane, muscle layers, and medullary region containing flame cells, nerve tissue, and other cell bodies are observed. Of particular interest is the presence of discrete sensory endings whose function is discussed.     

The hemocytes of Panstrongylus megistus (Hemiptera: Reduviidae)

Five hemocyte types were identified in the hemolymph of Panstrongylus megistus by phase contrast and common light microscopy using some histochemical methods. These are: Prohemocytes, small cells presenting a great nucleus/cytoplasm ratio; Plasmatocytes, the most numerous hemocytes, are polymorphic cells mainly characterized by a large amount of lysosomes; Granulocytes, hemocytes very similar to plasmatocytes which contain cytoplasmic granules and are especially rich in polysaccharides; Oenocytoids, cells presenting a small nucleus and a thick cytoplasm; they show many small round vacuoles when observed in Giemsa smears and many cytoplasmic granules under phase microscopy; Adipohemocytes, very large hemocytes, presenting many fat droplet inclusions which could correspond to free fat bodies which entered the hemolymph. Only prohemocytes and plasmatocytes can be clearly classified; all the other hemocyte types have a more ambiguous classification.     

Differences between larval and pupal hemocytes of the tobacco hornworm, Manduca sexta, determined by monoclonal antibodies and density centrifugation

Insect hemocytes play a major role in developmental processes where they disassociate and rebuild metamorphosing tissues while undergoing physiological changes themselves. We identified hemocyte changes from the last larval to the beginning of the pupal stage of the tobacco hornworm, Manduca sexta. Larval and pupal hemocytes behaved differently in a 40% Percoll density gradient. Larval granular cells were found in almost all density layers, pupal granular cells were abundant in high density layers; larval plasmatocytes occurred in dense layers, pupal plasmatocytes became enriched in less dense layers of the gradient. Using a panel of monoclonal antibodies generated against purified hemocytes, several different antibody binding patterns were identified. Quantitative differences in staining intensities were observed more often than qualitative changes, e.g. a loss or a gain of staining. Both phenomena were related to both plasmatocytes and granular cells. The distribution of the corresponding antigens in tissues was tested on cross sections of larvae and pupae as well as in Western blot analyses using organ homogenates. Several antibodies were specific for hemocytes only, among which two antibodies bound to molecules of the hematopoietic organ. Other antibodies had an additional reactivity to other tissues, mainly to the basal lamina.     

The cytology and cytochemistry of the hemocytes of Mytilus edulis and their responses to experimentally injected carbon particles

Hemocytes of Mytilus edulis were examined cytologically and cytochemically. On the basis of structure, staining reactions, and phagocytic behavior, they were divided into two main groups: basophilic hemocytes and eosinophilic granular hemocytes (granulocytes). The basophilic cells were further divided into small lymphocytes and larger phagocytic macrophages reactive for lysosomal hydrolases. Mitosis was observed in granulocytes and in small lymphoid cells, believed to be the stem cells for the basophilic cell line. A few cells appeared to be intermediate between lymphocytes and small granulocytes. Macrophages were the main cell type involved in the clearance of injected carbon particles. However, granulocytes did show some phagocytic activity. Brown cells displaying apparent amoebocytic behavior were found to contain Fe³⁺ and Pb²⁺ in cytoplasmic inclusions, some of which were also reactive for β-glucuronidase and glucosaminidase. These cells appear to have a separate origin from the hemocytes.