Structure of coupled and uncoupled cell junctions

Cells of Chironomus salivary glands and Malpighian tubules have junctions of the "septate" kind. This is the only kind of junction discerned which is large enough to effect the existing degree of intercellular communication. The electron microscopic observations of the "septate" junction conform to a honeycomb structure, with 80-A-thick electron-opaque walls and 90-A-wide transparent cores, connecting the cellular surface membranes. A projection pattern of light and dark bands (the "septa") with a 150-A periodicity results when the electron beam is directed normal to any set of honeycomb walls. Treatment of the salivary gland cells with media, which interrupt cellular communication (without noticeable alteration of cellular adhesion) by reducing junctional membrane permeability or perijunctional insulation, produces no alterations in the junctional structure discernible in electron micrographs of glutaraldehyde-fixed cell material.     

THE CELL JUNCTION IN A LAMELLIBRANCH GILL CILIATED EPITHELIUM : Localization of Pyroantimonate Precipitate

The junctional complex in the gill epithelium of the freshwater mussel (Elliptio complanatus) consists of an intermediary junction followed by a 2–3 µ long septate junction. Homologous and heterologous cell pairs are connected by this junction. After fixation with 1% OsO₄ containing 1% potassium pyroantimonate, electron microscopy of the gill reveals deposits of electron-opaque precipitate, specifically and consistently localized along cellular membranes. In both junctional and nonjunctional membrane regions, the precipitate usefully outlines the convolutions without obliterating the 150 A intercellular space, which suggests the rarity or absence of either vertebrate-type gap or tight junctions along the entire cell border. The precipitate appears on the cytoplasmic side of the limiting unit membranes of frontal (F), laterofrontal (LF), intermediate (I), lateral (L), and postlateral (PL) cells. The membrane surfaces of certain vesicles of the smooth endoplasmic reticulum, of multivesicular bodies, and of mitochondrial cristae contain precipitate, as does the nucleolus. In other portions of the cell, precipitate is largely absent. The amount of over-all deposition is variable and depends on the treatment of the tissue prior to fixation. Deposition is usually enhanced by pretreatment with 40 mM NaCl as opposed to 40 mM KCl, which suggests that the precipitate is in part sodium pyroantimonate. Treatment with 0.2 mM ouabain does not enhance deposition. Regional differentiation of cell membranes with respect to their ability to precipitate pyroantimonate is found in at least three instances: (a) between the ciliary membranes and other portions of the cell membrane: the precipitate terminates abruptly at the ciliary base, (b) between the LF and I cell borders: the precipitate is asymmetric, favoring the LF side of the junction, and (c) between the septate junctional membrane and adjacent membrane: the precipitate occurs periodically throughout the septate junction region with the periodicity corresponding to the spacing of the septa. This suggests that different regions of the cell membrane may have differing ion permeability properties and, in particular, that the septa may be the regions of high ion permeability in the septate junction.     

Intercellular Attachment in the Epithelium of Hydra As Revealed by Electron Microscopy

In Hydra adjacent epithelial cells are bound firmly to each other by desmosomes of a type not described in detail hitherto. The most prominent feature of these desmosomes is the presence of a series of parallel lamellae which bridge the intercellular space and connect the two apposed cell surfaces directly. These structures, here termed intercellular attachment lamellae, display two peaks of density about 50 A apart. These dense lines appear in some instances to be continuous with the outer dense components of the plasma unit membranes of the attached cells. The presence of prominent lamellae in intercellular attachments is sufficiently distinctive to deserve special terminology; accordingly, the term septate desmosome is proposed. It is noted that septate desmosomes may have been seen in other animals in instances where published electron micrographs show cross-striations or prominent connections in regions of intercellular attachment. It is suggested that septate desmosomes in Hydra, in addition to binding cells firmly to each other, form barriers to the movement of water into intercellular spaces and thus help to protect the organism's internal environment. Observations on the use of phosphotungstic acid for improving contrast in materials embedded in epoxy resins are also recorded.     

Arthropod immune system. III. Septate junctions in the hemocytic capsule of the german cockroach, Blattella germanica

Granulocytes (GRs) and/or plasmatocytes (PLs), the two major immunocytes in arthropods, participate in cellular encapsulation of foreign tissue. Although gap and desmosome junctions have been reported in insect capsules, smooth septate junctions are being reported for the first time by both thin section and freeze-fracture techniques in Blattella germanica. In 7-day-old capsules, the septa are 23 nm thick, faintly 'scalloped' and slightly curved in appearance; the interseptal space has a periodicity of about 5 nm. In freeze-fractured capsules, the septa are associated on both sides with the corresponding intramembranous structures, belonging to the plasma membranes of the two junction-forming GRs. The intercellular space is 27 nm wide. There are 36-40 septa/1 mum junctional length. The junctions show furrows on the extracellular fracture face (E) and the complementary regular rows of intramembranous particles on the cytoplasmic face (P). The septate junctions often occur in the region of the capsule that also shows the presence of gap junctions, but only rarely that of desmosomes. The septate junctions are in close proximity with mitochondria. It is suggested that the function of these junctions is to produce compact capsules.     

Fine structure of sporogenesis and septum formation in Micromonospora globosa Kriss and M. fusca Jensen.

Sporogenesis in two species of Micromonospora M. globosa and M. fusca (Actinomycetes) was quite similar. As in fungi, spore formation began as a blowing-out of a hyphal tip with the subsequent centripetal invagination of the plasma membrane. Septal wall material was deposited in a typical three-layered pattern, i.e., two electron-opaque layers separated by an electron-transparent layer. A second electron-opaque wall layer was later formed within the spore and finally a third, less electron-opaque wall was produced. Spore dihiscence was facilitated by the fragmentation of the first-formed wall surrounding the spore. Sporogenesis in Micromonospora is blastic in nature producing terminal, thick-walled spores. In M. fusca, a sporulation process was observed which closely resembled sporangial formation. The process appeared similar to that described for the genus Actinoplanes. Swollen, multiseptate structures were also present. Also in M. fusca, perforate septa with flared pore margins were observed. These septa were similar in appearance to the dolipore septa of Basidiomycetes although they lack a parenthesome and pore plug. Although an extensive membrane system (mesosome) was associated with the finishing septum, its function in the process of septum formation was not determined.     

Surface specializations of Fundulus cells and their relation to cell movements during gastrulation

Cell movements in Fundulus blastoderms during gastrulation were studied utilizing time-lapse cinemicrography and electron microscopy. Time-lapse films reveal that cells of the enveloping layer undulate and sometimes separate briefly but remain together in a cohesive layer. During epiboly, the marginal enveloping layer cells move over the periblast as it expands over the yolk sphere. Movement occurs as a result of ruffled membrane activity of the free borders of the marginal cells. Deep blastomeres become increasingly active during blastula and gastrula stages. Lobopodia project from the blastomeres in blastulae and adhere to other cells in gastrulae, giving the cells traction for movement. Contact specializations are formed by the lateral adjacent plasma membranes of enveloping layer cells. An apical junction is characterized by an intercellular gap of 60–75 A. Below this contact, the plasma membranes are separated by 120 A or more. In mid-gastrulae, cytoplasmic fibrils occur adjacent to some apical junctions, and small desmosomes appear below the apical junction. Septate desmosomes also appear at this time. A junction with an intercellular gap of 60 A occurs between marginal enveloping layer cells and periblast. Contacts between deep blastomeres become numerous in gastrulae and consist of contacts at the crests of surface undulations, short areas of contact in which the plasma membranes are 60 or 120 A apart, and long regions characterized by a 200-A intercellular gap. Lobopodia contact other blastomeres only in gastrulae. These junctions contain a 200-A intercellular space. Some deep blastomeres are in contact with the tips of periblast microvilli. The mechanism of epiboly in Fundulus is discussed and reevaluated in terms of these observations. The enveloping layer is adherent to the margin of the periblast and moves over it as a coherent cellular sheet. Periblast epiboly involves a controlled flow of cytoplasm from the thicker periblast into the thinner yolk cytoplasmic layer with which it is continuous. Deep cells move by adhering to each other, to the inner surface of the enveloping layer, and to the periblast.     

The structure and function of the smooth septate junction in a transporting epithelium: The malpighian tubules of the new zealand glow-worm Arachnocampa luminosa

Junctional complexes between the epithelial cells in the four distinct regions of the glow-worm Malpighian tubule were investigated by electron microscopy using thin sectioning, freeze-fracturing, osmotic disruption and tracer techniques. The lateral plasma membranes of all four cell types are joined by smooth septate junctions but the extent of the complex across the cell depth varies in the four different regions. The width of the septa, the interseptal spacing and the separation between the outer leaflets of the adjacent plasma membranes are different for each cell type. Gap junctions were identified only in the junctional complex between Type IV cells and were intercalated amongst large lateral sinuses. In oblique sections of lanthanum infiltrated tissue, the electron-lucent septa at the basal side of the junction are outlined by the tracer as it penetrates. In the Junctional complexes of all four regions the septa appear as short, distinct, linear bars. In tangential sections of gap junctions between Type IV cells, the junctions appear as a hexagonal array of intermembrane particles with a centre to centre spacing of 18 nm. Horseradish peroxidase did not penetrate the junctional complexes very far but readily passed through the basal lamina into the spaces between extracellular invaginations of the basement membrane of the cells. Junctional complexes in all four areas of the tubule have similar freeze-fracture faces. In freeze-fracture replicas of fixed tissue continuous ridges of fused particles are seen on the P face and complementary furrows are found on the E face. Junctional response to osmotically adjusted Ringer solutions was similar in all four cell types. Distortion or ‘blistering’ of the intercellular space between the septa of the junction occurred when the tissue was bathed in or injected with a hypertonic Ringer solution. The structure of these junctions, visualized by the different techniques, and the role of the septate junction in a transporting epithelium, are discussed.     

JUNCTIONAL COMPLEXES IN VARIOUS EPITHELIA

The epithelia of a number of glands and cavitary organs of the rat and guinea pig have been surveyed, and in all cases investigated, a characteristic tripartite junctional complex has been found between adjacent cells. Although the complex differs in precise arrangement from one organ to another, it has been regularly encountered in the mucosal epithelia of the stomach, intestine, gall bladder, uterus, and oviduct; in the glandular epithelia of the liver, pancreas, parotid, stomach, and thyroid; in the epithelia of pancreatic, hepatic, and salivary ducts; and finally, between the epithelial cells of the nephron (proximal and distal convolution, collecting ducts). The elements of the complex, identified as zonula occludens (tight junction), zonula adhaerens (intermediary junction), and macula adhaerens (desmosome), occupy a juxtaluminal position and succeed each other in the order given in an apical-basal direction. The zonula occludens (tight junction) is characterized by fusion of the adjacent cell membranes resulting in obliteration of the intercellular space over variable distances. Within the obliterated zone, the dense outer leaflets of the adjoining cell membranes converge to form a single intermediate line. A diffuse band of dense cytoplasmic material is often associated with this junction, but its development varies from one epithelium to another. The zonula adhaerens (intermediate junction) is characterized by the presence of an intercellular space (~200 A) occupied by homogeneous, apparently amorphous material of low density; by strict parallelism of the adjoining cell membranes over distances of 0.2 to 0.5 µ; and by conspicuous bands of dense material located in the subjacent cytoplasmic matrix. The desmosome or macula adhaerens is also characterized by the presence of an intercellular space (~240 A) which, in this case, contains a central disc of dense material; by discrete cytoplasmic plaques disposed parallel to the inner leaflet of each cell membrane; and by the presence of bundles of cytoplasmic fibrils converging on the plaques. The zonula occludens appears to form a continuous belt-like attachment, whereas the desmosome is a discontinuous, button-like structure. The zomula adhaerens is continuous in most epithelia but discontinuous in some. Observations made during experimental hemoglobinuria in rats showed that the hemoglobin, which undergoes enough concentration in the nephron lumina to act as an electron-opaque mass tracer, does not penetrate the intercellular spaces beyond the zonula occludens. Similar observations were made in pancreatic acini and ducts where discharged zymogen served as a mass tracer. Hence the tight junction is impervious to concentrated protein solutions and appears to function as a diffusion barrier or "seal." The desmosome and probably also the zonula adhaerens may represent intercellular attachment devices.     

Junctional specializations in the seminiferous epithelium of an insect (Triatoma infestans): a freeze fracture and lanthanum tracer study

Intercellular junctions in the seminiferous epithelium of the testis of Triatoma infestans were examined by conventional electron microscopy, tannic acid fixation, electron-opaque tracers, and freeze-fracture techniques. Distinctive aspects of the intercellular junctions are described in different regions of the testis follicles. In the basal region, close to the basal lamina, smooth septate junctions intermingled with gap junctions were observed between sustentacular cells. In the parabasal regions of the seminiferous epithelium, plated septate junctions, 'molluscous and arthropod' type (according to the classification of Green, 1981), were observed. Over the above junctions, in the central regions, also located between sustentacular cell membranes, parallel rows of intramembrane particles form successive belts of widely spaced septate junctions in an atypical configuration. Invertebrate gap junctions were also observed between adjacent spermatocyte membranes.     

FINE STRUCTURE AND MORPHOGENIC MOVEMENTS IN THE GASTRULA OF THE TREEFROG, HYLA REGILLA

The blastoporal groove of the early gastrula of the treefrog, Hyla regilla, was examined with the electron microscope. The innermost extension of the groove is lined with invaginating flask- and wedge-shaped cells of entoderm and mesoderm. The distal surfaces of these cells bear microvilli which are underlain with an electron-opaque layer composed of fine granular material and fibrils. The dense layer and masses of vesicles proximal to it fill the necks of the cells. In flask cells bordering the forming archenteron the vesicles are replaced by large vacuoles surrounded by layers of membranes. The cells lining the groove are tightly joined at their distal ends in the region of the dense layer. Proximally, the cell bodies are separated by wide intercellular spaces. The cell body, which is migrating toward the interior of the gastrula, contains the nucleus plus other organalles and inclusions common to amphibian gastrular cells. A dense layer of granular material, vesicles, and membranes lies beneath the surface of the cell body and extends into pseudopodium-like processes and surface undulations which cross the intercellular spaces. A special mesodermal cell observed in the dorsal lining of the groove is smaller and denser than the surrounding presumptive chordamesodermal cells. A long finger of cytoplasm, filled with a dense layer, vesicles and membranes, extends from its distal surface along the edge of the groove, ending in a tight interlocking with another mesodermal cell. Some correlations between fine structure and the mechanics of gastrulation are discussed, and a theory of invagination is proposed, based on contraction and expansion of the dense layer and the tight junctions at distal cell surfaces.     

Ultrastructural features of the constricted region of dividing plastids

Summary The ultrastructure of the constricted region of dividing plastids of spinach, bean, turnip, tobacco, and wheat has been studied. In these species, an electron-opaque, ring-like structure (RS) girdles the constricted region of plastids in advanced stages of division. The RS is a compound entity composed of two concentric rings of electron-opaque materials; one on the stromal face of the inner membrane and the other on the cytoplasmic face of the outer membrane. It was concluded that the compound nature of the RS is highly conserved in angiosperms being present in some cereal grasses and in plants representing four different orders of dicotyledonous plants. Evidence indicating that the electron-opaque materials of the RS are integrated into the envelope membranes was also provided and it was suggested that the envelope in the region of the RS may have unique properties. For spinach, it was also noted that plastids with deeply constricted necks tend to have RSs with lower volumes than those from wider necks and that endoplasmic reticulum was frequently present in the cytoplasm of the constriction region.Abbreviations RS ring structure - ER endoplasmic reticulum     

THE FINE STRUCTURE OF THE ELECTRIC ORGAN OF TORPEDO MARMORATA

The fine structure of the electric organ of the fish Torpedo marmorata has been examined after osmium tetroxide or potassium permanganate fixation, acetone dehydration, and Araldite embedment. This organ consists of stacks of electroplaques which possess a dorsal noninnervated and a ventral richly innervated surface. Both surfaces are covered with a thin basement membrane. A tubular membranous network whose lumen is continuous with the extracellular space occupies the dorsal third of the electroplaque. Nerve endings, separated from the ventral surface of the electroplaque by a thin basement membrane, contain synaptic vesicles (diameter 300 to 1200 A), mitochondria, and electron-opaque granules (diameter 300 A). Projections from the nerve endings occupy the lumina of the finger-like invaginations of the ventral surface. The cytoplasm of the electroplaques contains the usual organelles. A "cellular cuff" surrounds most of the nerve fibers in the intercellular space, and is separated from the nerve fibre and its Schwann cell by a space containing connective tissue fibrils. The connective tissue fibrils and fibroblasts in the intercellular space are primarily associated with the dorsal surface of the electroplaque.     

Ditylenchus dipsaci Infestation of Trifolium repens. II. Dynamics of Infestation Development

Trifolium repens (white clover) stolons were inoculated with Ditylenchus dipsaci (stem nematode), and the development of resulting infestations was monitored. Nematodes initially remained confined to superficial locations, concentrating in petiole axils near inoculation points. They were able to migrate slowly from the inidal inoculation points and infest adjacent axils, especially in regions near the stolon tip. As time progressed, in some axils, nematodes migrated through the stolon epidermis and colonized slowly expanding subepidermal pockets of host tissue (ca. 0.2-mm length of stolon/day). In these loci nematodes established exponentially increasing populations, but the rates of locus expansion remained constant, indicating that locus expansion was limited by unidentified host-dependent factors. As a result of increasing population pressure within subepidermal loci, J4 entered a "diapause" state and the rate of egg production by adults declined, thereby reducing rate of population growth to more sustainable levels. Typically, these populations peaked at ca. 10,000 individuals in ca. 160 days occupying 3-cm lengths of stolon. Thereafter, heavily infested regions of stolons started to die, leading to the formation of longitudinal splits in their epidermis. In other axils, nematodes did not migrate into the stolons but remained confined to axils. Some of these populations increased a hundred-fold in 95 days, with population growth ending when petioles started to die. Host plant stolon morphology was affected only when subepidermal stolon populations developed high population levels (>100 nematodes) within close proximity (<2 cm) to active terminal meristems. This occurred either when axillary buds became active on previously infested nodes or when nematodes established endoparasitic populations at locations near the stolon tip during winter and spring, when the rate of stolon extension was limited by low light intensity. Affected stolon tips could "escape" from the influence of such infestations when light intensity and temperature increased. Nematode activity was limited by low temperature rather than light intensity. Global warming is likely to lead to greater damage to infested plants during the winter and early spring because the predicted milder winter temperatures will enhance nematode activity but not necessarily promote stolon growth.     

Formation of the septate junction in regenerating planarian gastrodermis

Septate junctions develop initially just basad from apical junctional complexes at the apical ends of regenerating gastrodermal cells. The first morphological indication of differentiation of the junction is the appearance of gentle undulations of the plasma membranes of apposing cells. Subsequently dense dots develop at fairly regular intervals at the cytoplasmic surface of one cell, while SER cisternae become localized opposite them near the surface of the apposing cell. The dense dots are associated with bulges which narrow the intercellular space. Later the dense dots are replaced by filaments aligned along the inner leaflet of the parent cell. Strands of amorphous deposits form connections between SER cisternae and the sister membrane on the opposite side of the junction. Ruthenium red staining provides information on precursors which occupy the intercellular space between the apposed plasma membranes. As development of the junction progresses, ruthenium red stains only the newly formed septa but not the interseptal matrix. Regular arrangement of individual septa seems to be completed under the control of V-projections from both of their surfaces. Precursors for the structural material of the septa may be a secretory product derived from the SER. Dense dots and their derived filaments probably serve as reinforcing material for strengthening the cell membrane of the junction.     

Respiration and plasma membrane ATPase in strawberry stolons

The biochemical, physiological and anatomical properties of strawberry (Fragaria ananassa Duch.) cv. 'Cambridge Favourite' stolons were studied during growth. ATPase activity was measured, in microsomal and plasma membrane fractions, along with chlorophyll determination, in-situ photosynthesis measurements and scanning electron microscopy (SEM) and X-ray microanalysis of stolon cross-sections. Potassium-stimulated ATPase activity and proton-pumping, both together indicating the presence of plasma membrane ATPase, was greatest in the stolon tip, the tissue with the fastest growth and respiratory activity. The enzyme activity and respiration gradient from the tip of the stolon to the base was concomitant with xylem development which was more differentiated in the base than in the tip. These cross-sections also showed 30% greater amounts of calcium and potassium of the cryo-preserved basal part relative to the stolon tip. This gradient existed independent of the presence of daughter plants. A hypothesis is presented which suggests that for the long-distance longitudinal transport of nutrients this gradient between stolon tip and base is likely to be involved in stolon growth.     

THE FINE STRUCTURE OF STALKED BACTERIA BELONGING TO THE FAMILY CAULOBACTERACEAE

The fine structure of a series of stalked bacteria belonging to the genera Caulobacter and Asticcacaulis has been examined in thin sections. The cell wall has the multilayered structure typical of many Gram-negative bacteria, and continues without interruption throughout the length of the stalk. The core of the stalk, continuous with the cytoplasmic region of the cell, is enclosed in an extension of the cell membrane, and contains a system of internal membranes: it is devoid of ribosomes and nucleoplasm. A membranous organelle occupies the juncture of stalk and cell, separating the ribosomal region from the core of the stalk. Typical mesosomes also occur in the cell, being particularly frequent at the plane of division. The secreted holdfast is located at the tip of the stalk in Caulobacter, and at the pole of the cell adjacent to the stalk in Asticcacaulis.     

STUDIES ON AN EPITHELIAL (GLAND) CELL JUNCTION : II. Surface Structure

The surface structure of a gland epithelium (Drosophila salivary gland), particularly that at the junction between cells, was examined under the electron microscope. The junctional surface, which in the preceding paper was shown to be highly permeable to ions, has the following structural characteristics. About two-thirds of it are profusely infolded; the surface membranes of adjoining cells interdigitate and present desmosomes. The width of the intercellular space varies considerably. The remainder of the junctional surface, the third that abuts on the lumen, is rather straight. Here, the cell membranes are aligned parallel at a distance of 150 A, and interconnected at regular intervals of 100 A. The connecting material has a high electron opacity, and is about as thick as the cell membranes, but, unlike the latter, has no resolvable unit membrane structure. The surface at the cell base, which in the preceding paper was shown to be rather impermeable, is infolded and resembles the infolded junctional region. The luminal surface exhibits microvilli. Critical surface dimensions are given, and the implications of surface structure in intercellular permeability are discussed.     

Fine-structural Correlates of Growth in Hyphae of Ascodesmis sphaerospora

Mycelial mats of Ascodesmis sphaerospora were fixed and embedded for electron microscopy, and thin sections of 1-mm blocks, taken from the 1st to the 7th mm behind the hyphal tips, were cut parallel to the long axis of the hyphae. The hyphal tip region is characterized by an outer zone of electron-transparent vesicles, 500 to 1,000 A in diameter, and is apparently associated with wall elaboration. Immediately behind this region, dense granules become evident along convoluted membrane systems and along the plasma membrane; in the same region are numerous small lomasomes in the lateral wall. As the hypha grows, septa are laid down at 3- to 7-min intervals at a distance of 200 to 250 μ behind the hyphal tip. A cylinder of endoplasmic reticulum is intimately involved in cross-wall deposition from its earliest stages; as the wall grows in, it becomes increasingly constricted in the pore region, finally assuming a torus-like configuration. Woronin bodies are shown to have a crystalline substructure and to originate in pouch-like membrane systems. Cross-walls from a 7- to 13-hr-old mycelium frequently show highly ordered structures in the vicinity of the pore. These structures may appear either as laminar stacks of discs to one side of the pore or as series of stubby concentric rings within the pore area itself. In the latter case, a mass of granular material is frequently seen plugging the pore. Other unusual organelles and inclusions in 7- to 13-hr hyphae are vesicles containing swirls of beaded or dilated membrane, membrane-enclosed rods, and stacks of unit membranes associated with spherical, electron-transparent vesicles.     

An ultrastructural study and stereological analysis of the colon wall in the cockroach Periplaneta americana

The colonic epithelium has been examined for ultrastructural evidence of physiological activity. The cells show extensive folding of the apical plasma membrane, associated with mitochondria and an internal coating of particles about 120 Å diameter. Anteriorly many apical infoldings are dilated at the tip to form substantial extracellular spaces up to 0.8 μm wide. Narrow intercellular channels are present, opening to the haemolymph side of the epithelium. Pinocytosis is seen frequently at the basal surface. The surface densities of apical plasma membranes were not significantly different in the posterior mid-gut and colon. Similarly the volume densities of mitochondria were equal in the colonic epithelium and rectal pads, but the surface density of outer mitochondrial membranes was greater in the colon. It is suggested that the colon may absorb organic solutes from the gut lumen.