Ultrastructure of the Rectum Epithelial Cells in the Mosquito Larvae, Anopheles sinensis Wiedemann

ABSTRACT The Ultrastructure of rectum epithelial cells in the mosquito larvae, Anopheles sinensis Wiedemann, was studied using electron microscope. The rectal epithelium forms rectal papillae composed of the absorptive cells and the surrounding basal cells. Moreover, rectal epithelium was covered with thin cuticular intima. Apical plasma membrane of the epithelial cells had infoldings and in between them, mitochondria developed into elongated shape were attached. In addition, the membrane infoldings reach down into the cell cytoplasm to form several layers of leaflet-like prolongations. On both sides of these prolongations were also large, well-developed mitochondria. Their formation was that mitochondria were attached to 3 μm length and 4–13 layers of membrane wrinkle lump. Many spherites, which are lamelated crystals that form an illusory structure in concentric circles inside of the cytoplasm of epithelial cell were observed. Basal plasma membrane in the epithelial cells was also wrinkled to promulgate into the cytoplasm to become basal infoldings producing canaliculi in basal labyrinth formation. There were many mitochondria scattered in these formations as well. On the bottom of the epithelial cell, basal lamina was attached and between basal lamina and muscle bundle was subepithelial space, which is connective tissue. Inside the space, tracheal and nerve cells were observed.     

Structure of the larval midgut of the fly Chironomus thummi and its relationship to sites of cadmium sequestration

The midgut structure of 4-7 day old fourth instar Chironomus thummi larvae was investigated with the light arid electron microscopes. Four regions are present which may be identified by the following major features: (1) Anterior I: this is the region under the esophageal invagination. (2) Anterior II: short microviili characterize these cells. Long narrow, basal plasma membrane infoldings associated with mitochondria are conspicuous. This region is hypothesized to be important in ion and fluid transport. (3) Anterior III: numerous crystals are seen in these cells. Storage is proposed to be a major function of this region. (4) Posterior: these cells have long microviili, extensive RER, many Golgi, and short basal plasma membrane infoldings. Posterior cells probably function in secretion of digestive enzymes and absorption of nutrients. Cadmium is sequestered by the midgut, almost exclusively in the posterior cells. This phenomenon is discussed.     

Ultrastructural localization of carbonic anhydrase activity in the rat choroid plexus epithelial cell

Summary Localization of carbonic anhydrase activity was studied electron microscopically on cells of the rat choroid plexus epithelium. For the ultracytochemical detection of these activities, Yokota's technique (1969), which is the modification of Hansson's method (1967) was employed. Numerous electron dense reaction products were observed in the microvilli of the choroidal epithelial cell. The reaction deposits were also remarkably present in the infoldings of the basal plasmalemma but to a lesser extent than in the microvilli. The localization sites were mainly on the plasma membrane, but some reaction products were also observed in the cytoplasm near the plasma membrane. Hardly any reaction product was found in the intracellular organelles except for the mitochondria in which reaction products were occasionally observed on the cristae. These activities were completely inhibited by acetazolamide. As the carbonic anhydrase activity was histochemically seen in the microvilli and the basal infoldings, it is likely that carbonic anhydrase is related to an active transport process in the secretion of cerebrospinal fluid as is Na⁺, K⁺-ATPase (Masuzawa et al. 1980).     

Dense-cored membranous structures in smooth muscle cells of the vas deferens of the rat

Summary Smooth muscle cells from rat vas deferens were studied by electron microscopy. Vesicular and tubular membranous structures containing an electron-opaque material were found in the smooth muscle cells. Similar structures were also found in a subfraction (F3) of microsomes of vas deferens smooth muscle which was shown to be rich in both plasma membrane and putative endoplasmic reticulum markers. Treatment of the tissues with calcium-free Krebs solution containing EGTA prior to fixation eliminated almost completely the presence of these dense-cored membranous structures (DMS), whereas incubation of the subcellular membrane fraction with EGTA solution had no effect on the appearance of the DMS. Plasma membrane infoldings were found in the smooth muscle cells extending well into their interior. Horseradish peroxidase penetrates vesicles in a location similar to that of DMS in smooth muscle cells, suggesting that some of the DMS may be connected to the extracellular space. We conclude that the dense-core material within the DMS is calcium dependent. We also suggest that some of the DMS represent infoldings of the plasma membrane extending into the cell's interior.     

Fine structure of the larval midgut of the fly Rhynchosciara and its physiological implications

The midgut of Rhynchosciara americana larvae consists of a cylindrical ventriculus from which protrudes two gastric caeca formed by polyhedral cells with microvilli covering their apical faces. The basal plasma membrane of these cells is infolded and displays associated mitochondria which are, nevertheless, more conspicuous in the apical cytoplasm. The anterior ventricular cells possess elaborate mitochondria-associated basal plasma membrane infoldings extending almost to the tips of the cells, and small microvilli disposed in the cell apexes. Distal posterior ventricular cells with long apical microvilli are grouped into major epithelial foldings forming multicellular crypts. In these cells the majority of the mitochondria are dispersed in the apical cytoplasm, minor amounts being associated with moderately-developed basal plasma membrane infoldings. The proximal posterior ventriculus represents a transition region between the anterior ventriculus and the distal posterior ventriculus. The resemblance between the gastric caeca and distal posterior ventricular cells is stressed by the finding that their microvilli preparations display similar alkaline phosphatase-specific activities. The results lend support to the proposal, based mainly on previous data on enzyme excretion rates, that the endo-ectoperitrophic circulation of digestive enzymes is a consequence of fluid fluxes caused by the transport of water into the first two thirds of midgut lumen, and its transference back to the haemolymph in the gastric caeca and in the distal posterior ventriculus.     

Morphometric analysis of the translocation of lumenal membrane between cytoplasm and cell surface of transitional epithelial cells during the expansion-contraction cycles of mammalian urinary bladder

The flow of membrane between the cytoplasm and the lumenal surface during the expansion-contraction cycle of urinary bladder was estimated by stereological examination of electron micrographs of urothelial cells from guinea pigs, gerbils, hamsters, rabbits, and rats. The quantitative data obtained allowed an approximation of the surface area, volume, and numbers of lumenal membranelike vesicles and infoldings per unit volume of cytoplasm. Depending upon the species, approximately 85 to approximately 94% of the membrane surface area translocated into and out of the cytoplasm was in the form of discoidal vesicles. The remainder was accounted for by infoldings of the lumenal plasma membrane. The density of vesicles involved in transfer of membrane was quite similar in all the species examined, except guinea pigs which yielded lower values. In contrast, the densities of the total cytoplasmic pools of discoidal vesicles potentially available for translocation varied greatly among the different species. In general, species of animals with a highly concentrated urine had a greater density of discoidal vesicles than species with a less concentrated urine. This correlation may indicate an authentic relationship between lumenal membranes and the tonicity of urine, such as increased membrane recycling or turnover with increasingly hypertonic urine; or it may signify the existence of some other, more obscure relationship.     

High pressure freezing of intact plant tissues. Evaluation and characterization of novel features of the endoplasmic reticulum and associated membrane systems

We have used plant root tips frozen under high pressure in conjunction with freeze-fracture electron microscopy a) to evaluate the quality of freezing of unfixed, non-cryoprotected tissues obtainable with this method, b) to examine the structure of cells frozen under high pressure, c) to evaluate the usefulness of high pressure freezing to preserve transient membrane events, and d) to look for artifacts caused by the high pressure. A single artifact of high pressure, possibly related to the collapse of air spaces during pressurization before freezing, manifested itself as long tears or folds in the plasma membrane. Excellent freezing, as evidenced by the smooth, turgid appearance of all membrane systems and the lack of aggregated cytosolic materials was observed in 10 to 20% of samples. In the best preserved specimens freezing was uniform throughout the sample volume and all organelles were readily identified. In the remaining ones, a gradient of ice crystal sizes was seen; cells within 50 to 100 microns of the surface being better preserved than those in the interior. Cortical microtubules appeared well preserved as were close associations of endoplasmic reticulum (ER) with nuclear, Golgi and plasma membranes. Junctions between the ER and nuclear membrane were constricted and much thinner (30 nm in diameter) than in chemically-fixed, thin-sectioned tissue, and although no continuities between the ER and Golgi membranes were observed, many Golgi stacks had an adjacent ER cisterna either at the cis or trans face. Both Golgi and ER cisternae exhibited distinct, round dilations indicative of vesicle blebbing or vesicle fusion events. Characteristic disc- and horseshoe-shaped infoldings of the plasma membrane corresponding to fused secretory vesicle and/or membrane recycling structures were also prominent in many cells. Short extensions of the cortical ER cisternae were regularly observed appressed against these plasma membrane infoldings suggesting a functional role for the ER in vesicle-mediated secretion and/or membrane recycling. Many lipid bodies were intimately associated with the ER, some with their surface monolayer fused with the cytoplasmic leaflet of the ER membrane. Our findings demonstrate that high pressure freezing can provide excellent morphological preservation of intact tissues and can preserve fast, transient membrane events such as those associated with vesicle fusion and vesicle blebbing. We conclude that this is the best available method for freezing relatively large (up to 0.6 mm thick) tissue samples for study by electron microscopy.     

Alkaline phosphatase, 5'-nucleotidase and magnesium-dependent adenosine triphosphatase activities in the transitional epithelium of the rat urinary bladder.

The cerium-based method was used to demonstrate cytochemically the ultrastructural localization of alkaline phosphatase (ALPase), 5'-nucleotidase (5'-Nase) and magnesium-dependent adenosine triphosphatase (Mg-ATPase) on the transitional epithelium of the rat urinary bladder. The reaction product for ALPase was found on the plasma membrane of all epithelial cells, except the luminal surface of superficial cells. The activity of 5'-Nase appeared on the plasma membrane of all bladder transitional epithelial cells, including the free surface of superficial cells. The Mg-ATPase reaction product was seen on the plasma membrane of superficial, intermediate and basal cells, but never on the luminal surface of superficial cells and it was only occasionally seen on the basal surface. The possible functions of these phosphatases have been discussed, and it was emphasized that the 5'-Nase activity present on the luminal surface of superficial cells may play a special role in the membrane movement of these cells in the transitional epithelium.     

The Fine Structure of the Retina Studied with the Electron Microscope : IV. Morphogenesis of Outer Segments of Retinal Rods

The morphogenesis of the outer segments of retinal rods was studied mainly in the kitten before the opening of the eye, and the probable sequence of the morphogenetic stages is deduced. Since the development of retinal rods is not synchronous, the deductions were based on observations of many single and serial sections. One centriole extends ciliary tubules of about 0.5 µ long, in the growing primitive cilium. Beyond this length, each ciliary tubule becomes a row of small vesicles (called "ciliary vesicles" in this paper), which penetrate into the distal region of the cilium. Where the ciliary vesicles establish contact with the plasma membrane of the distal region of the cilium, more or less deep infoldings of the plasma membrane are observed. In the distal region can be seen rows of tubular or vesicular structures. A few of these membranous structures are continuous with the bottoms of the infoldings. At the following stage, the infoldings disappear and the ciliary vesicles lose contact with the distal plasma membrane. Nonetheless, the formation of the tubular structures continues in the distal region of the primitive outer segment. The tubular structures appear to be transformed into the primitive rod sacs by sidewise enlargement. At a subsequent time, presumably, these primitive rod sacs flatten and are rearranged into a position perpendicular to the long axis of the outer segment. The detailed structure of the basal body of the connecting cilium was also studied by means of serial sections.     

Light and electron microscopic study of the hindgut of the ant (Formica nigricans, hymenoptera): I. Structure of the ileum

The study of the ileum of the ant Formica nigricans by light and electron microscopy revealed the existence of three differentiated regions: proximal, middle, and distal ileum. The middle region constitutes most of the length of the organ. Its wall is made up by a folded simple epithelium lined by a cuticle, which is surrounded by an inner circular muscle layer and various external longitudinal muscle fibers adjacent to the hemolymph. A subepithelial space is present between the epithelium and the circular muscle layer. Epithelial cells show extensive infoldings of the apical, and to a lesser extent the basolateral plasma membrane. Apical infoldings are characterized by the presence of 10-nm particles (portasomes) covering the cytoplasmic side of the membrane. Mitochondria are abundant throughout the cytoplasm, although they mainly are present underneath the apical infoldings. Lateral borders of epithelial cells display an apical junctional complex, mainly constituted by a long and convoluted pleated septate junction. These features support the view that epithelial cells in the middle ileum are specialized in ion solutes and water transport. The proximal ileum connects with the ampulla into which the Malpighian tubules drain. As opposed to the middle ileum, epithelial cells of the proximal ileum show less developed basolateral infoldings, and the apical plasma membrane is devoid of portasomes and only occasionally invaginates. These features suggest that the proximal ileum plays no relevant role in ion and water transport. The distal ileum penetrates into the rectal sac, forming a valve-like structure; this region presumably controls the amount of urine reaching the rectum.     

Ultrastructural,cyto- and biochemical observations during turnover of plasma membrane in duck salt gland

Summary The mechanism of plasma membrane turnover was investigated using the duckling salt gland as a model system. Feeding fresh water to saltstressed ducklings results in a decrease in the Na, K-ATPase in salt gland to nonstressed levels in about 7 days, as measured by ATP hydrolysis and ³H-ouabain binding. Electron micrographs reveal that this is accompanied by a decrease in plasma membrane infoldings on the basal and lateral borders of gland secretory cells. Simultaneously there is an increase in filamentous material and a rise in acid phosphatase and peptidase activities in these cells. Cytochemistry shows that the acid phosphatase activity is mostly associated with the basal or basolateral regions of secretory cells. These observations could indicate that the removal of plasma membrane components is accomplished by internalization and digestion within the secretory cells.     

Light and electron microscopic study of the hindgut of the ant (Formica nigricans, hymenoptera): II. Structure of the rectum

The rectum of the ant Formica nigricans is composed of six ovoid rectal papillae inserted into a rectal pouch. The wall of the rectal pouch is made up of a flat epithelium of simple rectal cells lined by cuticle, and surrounded by a circular muscle layer. Each rectal papilla is comprised by a simple columnar epithelium of principal cells facing the lumen, and a simple cuboid epithelium of secondary cells towards the hemolymph; a group of 20-25 slender junctional cells lies laterally between both epithelia enclosing an intrapapillar sinus. The muscle layer of the rectal wall also surrounds the base of the papillae. Principal cells do not exhibit extensive infoldings at the apical and basal plasma membranes. Lateral membranes, in contrast, develop highly folded mitochondria-scalariform junction complexes enclosing very narrow intercellular canaliculi between adjacent cells. These canaliculi open to wider intercellular sinuses that ultimately drain into the intrapapillar sinus at the sites of entry of tracheal cells. The lateral plasma membranes do not link to the apical or basal plasma membrane, thus originating a syncytium throughout the principal cells. The apical plasma membrane of secondary cells shows invaginations in relation with an apical tubulovacuolar system, bearing portasomes to the cytoplasmic side of the membrane. Secondary cells unite by convoluted septate junctions, and basolateral infoldings are also developed. These ultrastructural traits, some of them different from those found in other insects, are discussed and examined in relation to their role in water and solute absorption. A route for rectal transport in F. nigricans is proposed.     

Regulation of fish gill Na(+)-K(+)-ATPase by selective sulfatide-enriched raft partitioning during seawater adaptation

Na(+)-K(+)-ATPase is arguably the most important enzyme in the animal cell plasma membrane, but the role of the membrane in its regulation is poorly understood. We investigated the relationship between Na(+)-K(+)-ATPase and membrane microdomains or "lipid rafts" enriched in sulfatide (sulfogalactosylceramide/SGC), a glycosphingolipid implicated as a cofactor for this enzyme, in the basolateral membrane of rainbow trout gill epithelium. Our studies demonstrated that when trout adapt to seawater (33 ppt), Na(+)-K(+)-ATPase relocates to these structures. Arylsulfatase-induced desulfation of basolateral membrane SGC prevented this relocation and significantly reduced Na(+)-K(+)-ATPase activity in seawater but not freshwater trout. We contend that Na(+)-K(+)-ATPase partitions into SGC-enriched rafts to help facilitate the up-regulation of its activity during seawater adaptation. We also suggest that differential partitioning of Na(+)-K(+)-ATPase between these novel SGC-enriched regulatory platforms results in two distinct, physiological Na(+) transport modes. In addition, we extend the working definition of cholesterol-dependent raft integrity to structural dependence on the sulfate moiety of SGC in this membrane.     

The Plasma Membrane and Peripheral Cytoplasm of the Green Algal Flagellate, Gloeomonas kupfferi

A morphological analysis of the plasma membrane and peripheralendomembrane components of the unicellular chlamydomonad flagellate,Gloeomonas kupfferi, was performed. Conventional fixation, freezesubstitution, and rapid freeze-deep etch processing protocolsfor electron microscopic analyses revealed the following. Theplasma membrane is highlighted by distinct infoldings whichdo not appear to be sensitive to changes in osmotic or cellcycle conditions. These infoldings are irregularly-spaced androughly 200-225 nm apart. Each elliptical infolding is 400-440nm long and 90-115 nm wide. The exoplasmic face (EF) of eachinfolding is highlighted by an aggregation of 130-160 nm intramembranousparticles (imps) that are 9.1-10·5 nm in size. Theseinfoldings are associated with the peripheral endoplasmic reticulumnetwork and microtubular network internally and the inner walllayer externally. It is suggested that these infoldings maybe associated with cell wall maintenance.Copyright 1994, 1999Academic Press Gloeomonas, plasma membrane, infoldings, freeze fracture     

Isolation and characterization of granules of the toad bladder

Summary The electron-dense granules that lie just below the apical plasma membrane of granular epithelial cells of toad urinary bladder contribute glycoproteins to that apical membrane. Also, exocytosis of granules (and tubules) elicited by antidiuretic hormone potentially doubles that apical surface, during the same period the transport changes characteristic of the hormonal response occur.Granules separated from other membrane systems of the cells provide the material to assess the importance of the granules as glycocalyx precursors and in hormone action. We used isosmotic media to effect preliminary separations by differential centrifugation. Then granules were isolated by centrifugation on self-forming gradients of Percoll of decreasing hypertonicity.We find qualitative and quantitative changes in protein composition and enzymic activities in the isolated fractions. The primary criterion for granule purification was electron microscopic morphology. In addition, polypeptide species found in the granule fraction are limited in number and quantity. The granules are enzymically and morphologically not lysosomal in nature. Granules may provide the glycoproteins of the apical glycocalyx but they differ from the isolated plasma membrane fraction enzymically, in protein composition and in proportion of esterified cholesterol.We conclude that the granules are not average plasma membrane precursors. Their role in the membrane properties of the toad urinary bladder may now be evaluated by characterizing permeability and other properties of the isolated organelles.     

Ultrastructure of the Malpighian Tubule Cells in the Mosquito Larvae, Anopheles sinesis

Ultrastructure of epithelial cells constituting the Malpighian tubule of Anopheles sinesis last instar larvae was observed with electron microscope. Malpighian tubule consists of four long and narrow tubule structures with principal cells in typical absorptive cells and regenerative cells forming the simple epithelium. Apical plasma membrane of the principal cell is differentiated into microvilli with one mitochondrion in each microvilli. Basal plasma membrane had extreme infolding to form a canaliculi and a well developed mitochondria was attached in the infoldings. And, rER, ribosomes, and vacuoles were well developed inside the cells. However, there were two main cell types depending on the differentiation of cell organelles. Type 1 cell was cubic, forming the distal portion of Malpighian tubule. The length of microvilli was approximately 4 μm and the basal infoldings were introjected to the depth of 2 μm inside the cell. On the other hand, Type II cell that formed the main proxinal portion was a low squamous type cells with shorter 2 μm of microvilli and the basal infoldings were introjected to the depths of 4 μm inside the cell. As for vacuoles scattered inside the cells, they were regularly observed in both Type I and II and the Type II cells had better developed cellular organelles. Although regenerative cells were extremely small, their cellular organelles were developed and their overall electron density was high that they appeared darker than the principal cells.     

Ultrastructural studies and Na+,K+-ATPase immunolocalization in the antennal urinary glands of the lobster Homarus gammarus (Crustacea, Decapoda).

Unlike in crustacean freshwater species, the structure and ultrastructure of the excretory antennal gland is poorly documented in marine species. The general organization and ultrastructure of the cells and the localization of Na(+),K(+)-ATPase were examined in the antennal gland of the adult lobster Homarus gammarus. Each gland is composed of a centrally located coelomosac surrounded ventrally by a labyrinth divided into two parts (I and II) and dorsally by a voluminous bladder. There is no differentiated nephridal tubule between them. The labyrinth and bladder cells have in common a number of ultrastructural cytological features, including basal membrane infoldings associated with mitochondria, apical microvilli, and cytoplasmic extrusions, and a cytoplasm packed with numerous vacuoles, vesicles, lysosome-like bodies, and swollen mitochondria. Each type of cell also presents distinctive characters. Na(+),K(+)-ATPase was detected through immunofluorescence in the basal part of the cells of the labyrinth and in the bladder cells with an increasing immunostaining from labyrinth I to the bladder. No immunoreactivity was detected in the coelomosac. The cells of the labyrinth and of the bladder present morphological and enzymatic features of ionocytes. The antennal glands of the lobster thus possess active ion exchanges capabilities.     

Endocytic vesicles and surface invaginations in cultured vascular endothelium: a morphometric comparison

Ruthenium red staining of plasma membrane glycoproteins of confluent cultured arterial endothelial cells revealed that the limiting membrane of many apparently discrete cytoplasmic vesicles was continuous with the plasmalemma. Surface invaginations accessible to ruthenium red appeared as vesicles when sectioned out of the plane of attachment to the cell surface, Morphometric analysis of ruthenium red-positive (RR+) and ruthenium red-negative vesicles (RR-) indicated that 47.2% of the total apparent vesicle population was RR+ and that those infoldings accounted for 19.6 +/- 1.4% of the cell surface in transverse sections. Whereas 14.9% of the true vesicles (ruthenium red-negative) were coated vesicles, only 1.1% of RR+ "vesicles" were coated pits. These studies show that although many deep infoldings of the cell surface may be misinterpreted as vesicles, almost all are uncoated. The existence of discrete coated vesicles (independent of coated pits) in vascular endothelium in vitro is readily apparent.     

Characterization of the column and autocellular junctions that define the vasculature of gill lamellae.

Novel adhesion junctions have been characterized that are formed at the interface between pillar cells and collagen columns, both of which are essential constituents of the gill lamellae in fish. We termed these junctions the "column junction" and "autocellular junction" and determined their molecular compositions by immunofluorescence microscopy using pufferfish. We visualized collagen columns by concanavalin A staining and found that the components of integrin-mediated cell-matrix adhesion, such as talin, vinculin, paxillin, and fibronectin, were concentrated on plasma membranes surrounding collagen columns (column membranes). This connection is analogous to the focal adhesion of cultured mammalian cells, dense plaque of smooth muscle cells, and myotendinous junction of skeletal muscle cells. We named this connection the "column junction." In the cytoplasm near the column, actin fibers, actinin, and a phosphorylated myosin light chain of 20 kDa are densely located, suggesting the contractile nature of pillar cells. The membrane infoldings surrounding the collagen columns were found to be connected by the autocellular junction, whose components are highly tyrosine-phosphorylated and contain the tight junction protein ZO-1. This study represents the first molecular characterization and fluorescence visualization of the column and autocellular junctions involved in both maintaining structural integrity and the hemodynamics of the branchial lamellae.     

Fine structure of the rectal pads in the desert locust Schistocerca gregaria with reference to the mechanism of water uptake

The rectal pads of Schistocerca gregaria are composed of three different cell types: epithelial, secondary and junctional cells. The rectal pads are interconnected by simple rectal cells and both are lined internally by a articular intima. The epithelial cells exhibit extensive infoldings of the apical plasma membranes that are closely associated with mitochondria. Their lateral plasma membranes are highly folded around large mitochondria and enclose intercellular channels and spaces. They are united by belt and spot desmosomes, septate junctions, gap junctions and scalariform junctions, but terminate in a basal syncytium without contacting the basal plasma membranes. The apical and basal cytoplasm contain coated vesicles, dense tubular elements, multivesicular bodies and lysosomes, suggesting receptor-mediated endocytosis of small peptide molecules into the epithelial cells. The apical membrane infoldings of the secondary cells are also associated with large mitochondria. Their basal plasma membranes are covered by connective cell processes and connected with them by spot desmosomes which may be involved in solute recycling. The presence of neurosecretory-like axons near the secondary cells suggests that they exert local control on the function of these cells. The ultrastructural details are examined in relation to their role in solute and water transport.