Isolation and characterization of invertebrate smooth septate junctions

1983 ◽  
Vol 62 (1) ◽  
pp. 351-370
Author(s):  
C.R. Green ◽  
C. Noirot-Timothee ◽  
C. Noirot
Keyword(s):  
Molecular Weight ◽  
Gap Junctions ◽  
Freeze Fracture ◽  
Intercellular Junctions ◽  
Septate Junction ◽  
Major Protein ◽  
Septate Junctions ◽  
Isolation And Characterization ◽  
Significant Difference ◽  
Freeze Fracture Electron Microscopy

Using modifications of techniques used for the isolation of macula type intercellular junctions (gap junctions and desmosomes) the arthropod smooth septate junction has been isolated from insect midgut tissue. Midguts from cockroaches or mealworms were used and membrane fractions were obtained by sucrose gradient and ultracentrifugation techniques. Preparations with reasonable concentrations of septate junction were obtained and have been studied by thin-section, negative-stain and freeze-fracture electron microscopy. The junctions appeared to be well preserved, although there was evidence that the junction strands were able to slide within the plane of the membrane. Septa were seen to have a cross-striated appearance when viewed after negative staining but their exact structure remained difficult to determine. Polyacrylamide gel electrophoretic studies demonstrated the reproducibility of the isolation procedure and showed that septa may have a 47 000 molecular weight glycoprotein component. Gel electrophoresis also gave some indication of the intramembrane biochemistry of the smooth septate junction, with proteins of 31 000 and 32 000 molecular weight always occurring in the junction fractions. The junctions were, however, very sensitive to both mechanical and chemical treatments, the septa were destroyed by rough homogenization or by treatment with urea at a concentration as low as 1 M. Freeze-fracture of untreated, isolated junctions demonstrated no differences from junctions in intact tissue, while replicas of urea-treated material were more difficult to interpret as the component parts of the junctions became separated once the septa had been destroyed. Gap junctions were also obtained and resisted both mechanical and chemical treatment, which destroyed the septate junctions. Their major protein component appeared to have a molecular weight of 36 000. Attempts to isolate pleated septate junctions (from insects, molluscs and annelids) by the same techniques failed, implying a significant difference in the structures of the two types of septate junction.

Morphological Changes of Intercellular Junctions in the Rat Submandibular Gland Treated by Long-term Repeated Administration of Isoproterenol

1987 ◽  
Vol 66 (8) ◽  
pp. 1303-1309 ◽  
Author(s):  
T. Inoue ◽  
H. Yamane ◽  
T. Yamamura ◽  
M. Shimono
Keyword(s):  
Gap Junctions ◽  
Tight Junctions ◽  
Morphological Changes ◽  
Freeze Fracture ◽  
Acinar Cells ◽  
Intercellular Junctions ◽  
Repeated Administration ◽  
Experimental Conditions ◽  
Significant Difference

Long-term repeated administration of isoproterenol (lPR) 2 mg/100 g bw, once daily for ten days, resulted in morphological changes in the intercellular junctions of rat submandibular glands, which were investigated by means of the freeze fracture technique. A significantly increased number of tight-junctional strands was present. These junctional strands extended much deeper toward the basal membrane than those in normal acinar cells. The basal frontier strands that branched from the networks of tight junctions were elongated and had either free-endings or terminal loops, which were more frequently observed in the IPR-treated acinar cells than in untreated acinar cells. Some of the strands of tight junctions were connected to small gap junctions. The diameters of gap junctions were not significantly different from those of control acinar cells. However, smooth areas devoid of particles were found intermingling with the usual packed particles in irregularly shaped small gap junctions. There was no significant difference between the desmosomes of IPR-treated and untreated acinar cells, in terms of either morphology or distribution. These changes in junctional morphology in the IPR-treated acinar cells resemble those seen in salivary glands during development, and in some experimental conditions including tumorous changes.

scholarly journals COEXISTENCE OF GAP AND SEPTATE JUNCTIONS IN AN INVERTEBRATE EPITHELIUM

1971 ◽  
Vol 50 (1) ◽  
pp. 92-101 ◽  
Author(s):  
A. J. Hudspeth ◽  
J. P. Revel
Keyword(s):  
Electron Microscope ◽  
Gap Junctions ◽  
Gap Junction ◽  
Extracellular Space ◽  
Intercellular Junctions ◽  
Septate Junction ◽  
High Electron ◽  
Septate Junctions ◽  
Electrotonic Coupling ◽  
Lanthanum Tracer

The intercellular junctions of the epithelium lining the hepatic caecum of Daphnia were examined. Electron microscope investigations involved both conventionally fixed material and tissue exposed to a lanthanum tracer of the extracellular space. Both septate junctions and gap junctions occur between the cells studied. The septate junctions lie apically and resemble those commonly discerned between cells of other invertebrates. They are atypical in that the high electron opacity of the extracellular space obscures septa in routine preparations. The gap junctions are characterized by a uniform 30 A space between apposed cell membranes. Lanthanum treatment of gap junctions reveals an array of particles of 95 A diameter and 120 A separation lying in the plane of the junction. As this pattern closely resembles that described previously in vertebrates, it appears that the gap junction is phylogenetically widespread. In view of evidence that the gap junction mediates intercellular electrotonic coupling, the assignment of a coupling role to other junctions, notably the septate junction, must be questioned wherever these junctions coexist.

Phylogenetic Relationships within Theinvertebrata in Relation to Thestructure of Septate Junctions and the development of ‘Occluding’ Junctional Types

1982 ◽  
Vol 53 (1) ◽  
pp. 279-305 ◽  
Author(s):  
COLIN R. GREEN ◽  
PATRICIA R. BERGQUIST
Keyword(s):  
Tight Junction ◽  
Phylogenetic Relationships ◽  
Freeze Fracture ◽  
Intercellular Junctions ◽  
Septate Junction ◽  
Simple Type ◽  
Septate Junctions ◽  
Exterior Surface ◽  
Lanthanum Tracer ◽  
To Come

The structures of 13 variants of invertebrate septate junction are reviewed on the basis offreeze-fracture, lanthanum tracer and thin-section studies. In addition, a simple type ofoccluding junction in the phylum Porifera, a variation of tight junction in the phylum Tunicateand the vertebrate tight junction are covered. All the junctions considered form a belt around the apical circumference of cells lining a lumen or an exterior surface. The large number of these junctions now recognized permits discussion relating to invertebrate classification and suggested phylogenetic relationships, and to the development of intercellular junctions. The relationships revealed are discussed under three headings: Coelenterates and lower invertebrates, Proterostomia (the annelid, molluscan and arthropod lineage) and the Deuterostomia(the echinoderm and chordate lineage). It is proposed that the pleated septate junction of the lower invertebrates resembles that of the hydrozoan rather than anthozoan Coelenterates. This lower invertebrate pleated septate junction occurs in several lower invertebrate phyla including the Annelida (of the proterostome lineage), but also occurs in the Sipunculoidea, a group supposedly on the deuterostome lineage.The proterostome line includes the molluscs and the arthropods, which have the molluscarthropodpleated septate junction. Several variations of the smooth septate junction are alsoseen in Arthropoda. Among the deuterostomes the Chaetognatha have both a paired septatejunction and a pleated junction and are therefore considered to be not very far removed fromthe Sipunculoidea. The echinoderms and hemichordates also have double-septum septatejunctions. In addition however, these two phyla have anastomosing septate junctions thatare very similar, varying only in their final configuration. Of the two, the echinoderm anastomosingseptate junction most closely resembles the tight junction seen in the tunicates, and the Hemichordata are therefore considered to be a lateral development from the main lineof chordate evolution. The tunicates have a tight junction similar to that seen in vertebrates;it is however more ‘leaky’ and has distinctive freeze-fracture characteristics.In the phylum Porifera a form of simple parallel membrane junction appears to serve anoccluding function. This junction has regular intercellular spacing in the absence of any septaand it is suggested that the spacing in septate junctions is probably not dictated by the septa.This interpretation is reasonable particularly when the diversity of septal types in conjunctionwith stable intercellular spacing is considered. Finally, a theory is put forward suggesting thatin evolution a change from the septate to the tight junction could simply involve a modificationof a ‘membrane spacing factor’, which allows the membranes of adjacent cells to come together at intervals, in the normal tight junction pattern.

scholarly journals A freeze-fracture and lanthanum tracer study of the complex junction between Sertoli cells of the canine testis.

1978 ◽  
Vol 76 (1) ◽  
pp. 57-75 ◽  
Author(s):  
C J Connell
Keyword(s):  
Tight Junctions ◽  
Sertoli Cells ◽  
Freeze Fracture ◽  
Intercellular Junctions ◽  
Septate Junction ◽  
Septate Junctions ◽  
Thin Sections ◽  
En Face ◽  
Lanthanum Tracer ◽  
Complex Junction

What appear to be true septate junctions by all techniques currently available for the cytological identification of intercellular junctions are part of a complex junction that interconnects the Sertoli cells of the canine testis. In the seminiferous epithelium, septate junctions are located basal to belts of tight junctions. In thin sections, septate junctions appear as double, parallel, transverse connections or septa spanning an approximately 90-A intercellular space between adjacent Sertoli cells. In en face sections of lanthanum-aldehyde-perfused specimens, the septa themselves exclude lanthanum and appear as electron-lucent lines arranged in a series of double, parallel rows on a background of electron-dense lanthanum. In freeze-fracture replicas this vertebrate septate junction appears as double, parallel rows of individual or fused particles which conform to the distribution of the intercellular septa. Septate junctions can be clearly distinguished from tight junctions as tight junctions prevent the movement of lanthanum tracer toward the lumen, appear as single rows of individual or fused particles in interlacing patterns within freeze-fracture replicas, and are seen as areas of close membrane apposition in thin sections. Both the septate junction and the tight junction are associated with specializations of the Sertoli cell cytoplasm. This is the first demonstration in a vertebrate tissue of a true septate junction.

Topographical variations in the structure of the smooth septate junction

1979 ◽  
Vol 37 (1) ◽  
pp. 373-389
Author(s):  
H.B. Skaer ◽  
J.B. Harrison ◽  
W.M. Lee
Keyword(s):  
Musca Domestica ◽  
Functional Significance ◽  
Freeze Fracture ◽  
Rhodnius Prolixus ◽  
Malpighian Tubules ◽  
Septate Junction ◽  
Lanthanum Hydroxide ◽  
Septate Junctions ◽  
Standard Fixation

Smooth septate junctions in the midgut of Musca domestica and in Malpighian tubules of both Musca and Rhodnius prolixus are described. Details of the structures revealed after standard fixation, fixation in the presence of the stain, lanthanum hydroxide, and after freeze-fracture are discussed in the light of models previously put forward to explain the interrelations of the images obtained by these different methods. The organization of the junction between cells of the midgut varies in the apical-to-basal axis. At the apical border the septa (or ridges in freeze-fracture replicas) are packed tightly and follow an undulating but strictly parallel course. This packing loosens towards the middle of the junction until, at its basal extremity, the septa (ridges in replicas) are widely separated and follow independent meandering courses. That these features are found both in lanthanum-infiltrated specimens and freeze-fracture replicas allows a correlation to be made between the septa and the freeze-fracture ridges. The functional significance of these smooth septate junctions is discussed.

Ultrastructural analysis of the apical ectodermal ridge during vertebrate limb morphogenesis

Development ◽  
1977 ◽  
Vol 41 (1) ◽  
pp. 223-232
Author(s):  
John F. Fallon ◽  
Robert O. Kelley
Keyword(s):  
Electron Microscopy ◽  
Gap Junctions ◽  
Freeze Fracture ◽  
Apical Ectodermal Ridge ◽  
Structural Requirements ◽  
Limb Morphogenesis ◽  
Vertebrate Limb ◽  
The Difference ◽  
Freeze Fracture Electron Microscopy ◽  
Fracture Electron

The fine structure of the apical ectodermal ridge of five phylogenetically divergent orders of mammals and two orders of birds was examined using transmission and freeze fracture electron microscopy. Numerous large gap junctions were found in all apical ectodermal ridges studied. This was in contrast to the dorsal and ventral limb ectoderms where gap junctions were always very small and sparsely distributed. Thus, gap junctions distinguish the inductively active apical epithelium from the adjacent dorsal and ventral ectoderms. The distribution of gap junctions in the ridge was different between birds and mammals but characteristic within the two classes. Birds, with a pseudostratified columnar apical ridge, had the heaviest concentration of gap junctions at the base of each ridge cell close to the point where contact was made with the basal lamina. Whereas mammals, with a stratified cuboidal to squamous apical ridge, had a more uniform distribution of gap junctions throughout the apical epithelium. The difference in distribution for each class may reflect structural requirements for coupling of cells in the entire ridge. We propose that all cells of the apical ridges of birds and mammals are electrotonically and/or metabolically coupled and that this may be a requirement for the integrated function of the ridge during limb morphogenesis.

scholarly journals Septate junctions in imaginal disks of Drosophila: a model for the redistribution of septa during cell rearrangement.

1982 ◽  
Vol 94 (1) ◽  
pp. 77-87 ◽  
Author(s):  
D K Fristrom
Keyword(s):  
Freeze Fracture ◽  
Septate Junction ◽  
Lateral Boundary ◽  
Progressive Change ◽  
Septate Junctions ◽  
Thin Sections ◽  
Cell Rearrangement ◽  
Imaginal Disks ◽  
Discrete Domains ◽  
Small Channels

The organization of septate junctions during morphogenesis of imaginal disks is described from freeze-fracture replicas and thin sections with a view to understanding junction modulation during rearrangements of cells in epithelia. The septate junctions of each epithelial cell of the disk are distributed in a number of discrete domains equal to the number of neighboring cells. Individual septa traverse domains of contact between pairs of adjacent cells, turn downwards at the lateral boundary of the domain and run parallel to the intersection with a third cell. This arrangement leaves small channels at three-cell intersections that are occupied by specialized structures termed "tricellular plugs." Cell rearrangement involves a progressive change in the width of contact domains between adjacent cells, until old contacts are broken and new ones established. It is proposed that the septate junction adjusts to the changing width of domains by the compaction or extension of existing septa. This redistribution of septa theoretically allows a transepithelial barrier to be maintained during cell rearrangements. The applicability of this model to other epithelial tissues is discussed.

scholarly journals The intramembrane structure of septate junctions based on direct freezing

1986 ◽  
Vol 80 (1) ◽  
pp. 13-28
Author(s):  
B. Kachar ◽  
N.A. Christakis ◽  
T.S. Reese ◽  
N.J. Lane
Keyword(s):  
Horseshoe Crab ◽  
Freeze Fracture ◽  
Transmembrane Proteins ◽  
Periodic Variation ◽  
Septate Junction ◽  
Fracture Face ◽  
Copper Block ◽  
Septate Junctions ◽  
The Relationship ◽  
Protoplasmic Fracture Face

Smooth septate junctions from the midgut of the cricket, Acheta, and the horseshoe crab, Limulus, as well as Hydra-type septate junctions from the epidermis of Hydra have been studied by freeze-fracture after direct freezing using the liquid helium-cooled copper block/slam freezing method. The exoplasmic fracture face at both types of septate junction exhibits rows of closely packed but irregularly shaped intramembrane particles. Complementary to these particle rows, on the protoplasmic fracture face, are sharply defined grooves with a periodic variation in depth and width that was conspicuous in Hydra but less well defined in arthropods. The closely packed, irregular particles on the exoplasmic faces could represent plastically deformed portions of transmembrane proteins pulled through the bilayer during freeze-fracture. On the basis of this interpretation, the grooves on the protoplasmic faces represent a confluence of the bilayer disruptions occurring during fracturing. The structures observed here are different from those reported in replicas of glutaraldehyde-fixed and glycerol-cryoprotected tissue, in which the intramembrane junctional components partition with the protoplasmic face and often assume the appearance of continuous cylinders. This comparison illustrates some of the artifacts associated with freeze-fracturing and shadowing. On the basis of a comparison of freeze-fracture replicas and sections of lanthanum-infiltrated tissues, the relationship between intramembrane junctional components and intercellular septal elements is analysed.

Tight and gap junctions in the intestinal tract of tunicates (Urochordata): a freeze-fracture study

1986 ◽  
Vol 84 (1) ◽  
pp. 1-17
Author(s):  
N.J. Lane ◽  
R. Dallai ◽  
P. Burighel ◽  
G.B. Martinucci
Keyword(s):  
Gap Junctions ◽  
Tight Junctions ◽  
Thin Section ◽  
Intestinal Tract ◽  
Freeze Fracture ◽  
Intestinal Cells ◽  
Fracture Profile ◽  
Tracer Studies ◽  
Septate Junctions ◽  
Fracture Study

The intestinal tracts from seven different species of tunicates, some solitary, some colonial, were studied fine-structurally by freeze-fracture. These urochordates occupy an intermediate position phylogenetically between the vertebrates and the invertebrates. The various regions of their gut were isolated for examination and the junctional characteristics of each part investigated. All the species examined exhibited unequivocal vertebrate-like belts of tight-junctional networks at the luminal border of their intestinal cells. No septate junctions were observed. The tight junctions varied in the number of their component strands and the depth to which they extended basally, some becoming loose and fragmented towards that border. The junctions consisted of ridges or rows of intramembranous particles (IMPs) on the P face, with complementary, but offset, E face grooves into which IMPs sometimes fractured. Tracer studies show that punctate appositions, the thin-section correlate of these ridge/groove systems, are sites beyond which exogenous molecules do not penetrate. These junctions are therefore likely to represent permeability barriers as in the gut tract of higher chordates. Associated with these occluding zonular junctions are intermediate junctions, which exhibit no identifiable freeze-fracture profile, and macular gap junctions, characterized by a reduced intercellular cleft in thin section and by clustered arrays of P face particles in freeze-fractured replicas; these display complementary aggregates of E face pits. The diameters of these maculae are rarely very large, but in certain species (for example, Ciona), they are unusually small. In some tissues, notably those of Diplosoma and Botryllus, they are all of rather similar size, but very numerous. In yet others, such as Molgula, they are polygonal with angular outlines, as might be indicative of the uncoupled state. In many attributes, these various junctions are more similar to those found in the tissues of vertebrates, than to those in the invertebrates, which the adult zooid forms of these lowly chordates resemble anatomically.

Plasma membranes, cell junctions and cuticle of the rectal chloride epithelia of the larval dragonfly Aeshna cyanea

1983 ◽  
Vol 59 (1) ◽  
pp. 159-182
Author(s):  
J. Kukulies ◽  
H. Komnick
Keyword(s):  
Gap Junctions ◽  
Structural Control ◽  
Plasma Membranes ◽  
Freeze Fracture ◽  
Septate Junction ◽  
Cell Junctions ◽  
Junctional Complex ◽  
Apical Region ◽  
Thin Sections ◽  
Aeshna Cyanea

The cell membranes and cell junctions of the rectal chloride epithelia of the larval dragonfly Aeshna cyanea were examined in thin sections and by freeze-fracture. These epithelia function in active ion absorption and maintain a high concentration gradient between the haemolymph and the fresh-water environment. Freeze-fracturing reveals fine-structural differences in the intramembraneous particles of the luminal and contraluminal plasma membranes of these epithelia, reflecting the functional diversity of the two membranes, which are separated by the junctional complex. The particle frequency of the basolateral plasma membranes is reduced after transfer of the larvae into high concentrations of environmental salinity. The junctional complex is located in the apical region and composed of three types of cell junctions: the zonula adhaerens, seen in freeze-fracture as a nearly particle-free zone; the extended and highly convoluted pleated septate junction and randomly interspersed gap junctions of the inverted type. Gap junctions also occur between the basolateral plasma membranes. They provide short-cuts in the diffusion pathway for direct and rapid co-ordination of the interconnected cell processes. Colloidal and ionic lanthanum tracer solutions applied in vivo from the luminal side penetrate through the cuticle via epicuticular depressions, but invade only the apical portion of the junctional complex. This indicates that the pleated septate junction constitutes a structural control of the paracellular pathway across the chloride epithelia, which are devoid of tight junctions. The structure of the pleated septate junctions is interpreted as a device for the extension of the diffusion distance, which is inversely related to the net diffusion. A conservative estimate of the total length of the junction, and the number and extension of septa reveals that the paracellular route exceeds the transcellular route by a factor of 50.

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