Sealing junctions in a number of arachnid tissues.

The junctions present in the central nervous system (CNS), midgut, silk gland and venom gland of arachnids have been investigated. Special care was taken to try to locate tight junctions in tissue other than CNS but they were not found in any of the other tissues. The detailed structure of the junctions present are discussed. The tight junctions present in CNS are somewhat different in appearance and fracturing behaviour to most vertebrate tight junctions and closely resemble only those found in Urochordates (a non-vertebrate chordate). The two types of septate junctions found in the other tissues belong to the pleated septate and smooth septate classes but show some interesting differences. It appears probable that the septate junctions in Arachnida, Merostomata and Myriopoda have different fracturing properties from those found in other arthropods. The finding that only septate junctions are present in most arachnid tissues, although tight junctions are present in CNS, is discussed in the context of the sealing function of septate junctions in invertebrate tissues.

Particle arrays on insect nerve membranes.

Freeze-fracture studies of Lepidopteran antennae have revealed the presence of orthogonal arrays of particles on antennal nerve membranes. Initial impressions were that several different arrays were present. Optical diffraction was used to examine the arrays on the electron micrograph negatives. This technique showed that all the arrays were derived from the same basic structure, suggesting that the superficial differences in appearance were due to shadowing effects. The particles are arranged in a lattice with spacings of 10.7 nm X 9.1 nm. The arrays are not clear-cut but tend to break up, producing a disorganized region around their edges. The particles are shown to have depressions in them. However, the evidence available suggests that the arrays do not have the other characteristics of gap junctions. The arrays appear not to be present on most of the nerve membrane faces, occurring only in localized regions of the nerve membranes where they are present in large numbers. This suggests that the arrays may have a specialized local function.

A new type of gap junction in the phylum Brachiopoda.

Gap junctions in the phylum Brachiopoda are shown to be of the P type as has been previously reported in invertebrates only in molluscs and possibly bryozoans. However, the mollusc gap junctions appear identical to vertebrae gap junctions in the fracture face on which particles remain, in particle spacing and in particle arrangement. The brachiopod gap junction, on the other hand, has general appearance traits which are more commonly seen in other invertebrate gap junction types. The freeze-fracture appearance of the brachiopod gap junction is described and discussed in relation to other types of gap junction which have been reported.

Two new septate junctions in the phylum Coelenterata.

Freeze-fracture of fixed and unfixed tissue, lanthanum tracer and conventional thin-section studies have revealed 2 new types of septate junction in the class Anthozoa, phylum Coelenterata. These new junctions have the 15-18-nm intercellular spacing of all other described septate junctions and are found around the apical circumference of cells lining a lumen or outside edge. However, in freeze-fracture replicas and tangential views of lanthanum-impregnated tissue, they are seen to be quite different from other known septate junction types. One of the new junctions is found in endothelial tissue such as that lining the gut or the inside of the tentacles. In tangential view it is seen to consist of relatively short, straight, double septa, again with lateral projections. In feeeze-fracture of unfixed tissue, the junction consists of double rows of particles on the P face, the particles of one row being rounded, those of the other being elongated at right angles to the line of the septum. This dichotomy in particle size is unexpected, as the 2 halves of the septa as seen in tangential view are symmetrical. In freeze-fracture of fixed material the particle arrays remain on the P face and appear similar to those of unfixed material, but never as clear. In fixed tissue, some distortion had occurred and in extreme cases septa appear as a single broad jumbled row of particles. In this double septa junction, the rows of particles seen in freeze-fracture are occasionally seen to anastomose with a septum dividing into 2 and a third row of particles aligning with the 2 new septa to form their double particle rows. In both fixed and unfixed tissues, the E face of the junction consists of wide, shallow grooves. The second of the new junctions occurs in epithelial tissue, such as around the outer edge of sea-anemone tentacles, and consists of long wavy septa with lateral projections. In views where these projections appear longest, they arise predominantly from one side of the septa. In freeze-fracture of both fixed and unfixed tissue, this junction appears as rows of closely spaced particles on the P face. Occasionally rows of particles are seen on the E face, but usually this face is characterized by shallow grooves. In some aspects these 2 new junctions have features in common with the Hydra type junction also found in the Coelenterata. In all 3 types septa are relatively straight, rather than pleated, and there are lateral projections on the septa.

Rectal papillae in Musca domestica: the cuticle and lateral membranes.

The role of specialized regions of insect rectal papillae in the regulation of water and ion uptake is well documented. Although the apparatus for active uptake of water or ions is located in various cell membranes, the absorbed molecules must first pass through the cuticle which lines the rectal epithelium. Most cuticle (e.g. abdominal) has been shown to be permeable only to molecules soluble in wax, and to be impermeable to water and ions. Obviously if such cuticle lined the rectum, absorption of water and ions would be severely restricted. The present freeze-fracture and lanthanum tracer study was undertaken to investigate in more detail both the morphological features of the rectal papillae cuticle which could be responsible for its anomalous permeability and the various cell membranes involved in this transport. It has been suggested from permeability studies that the anomalous permeability of rectal papillae cuticle could be due to the lack of a complete wax layer over the surface of the rectal cuticle. The present study strongly supports this suggestion. Thus, the freeze-fracture micrographs have shown that a surface layer of the cuticle reacts during fracturing like a lipid bilayer. However, in rectal papilla cuticle this surface bilayer is interrupted at each epicuticular depression by areas of different fracturing behaviour. These discontinuities in the surface bilayer probably allow the rectal contents to contact directly the true cuticular matrix. They could, therefore, explain the case with which water and ions penetrate the rectal cuticle and so gain access to the underlying epithelial cells. Although similar discontinuities are present on some of the rectal cuticle surface external to the rectal papillae, they appear to be filled in by plugs of lipid-like material. The lateral plasma membranes of the rectal papillae cells are generally considered to be the main site of active transport. The present lanthanum tracer and freeze-fracture study has shown that the lateral plasma membranes contain 3 distinct differentiations. Septate junctions are present at the apical and basal surfaces of the epithelial layer; a further membrane differentiation is found adjacent to the septate junctions; and thirdly, an array of short, variable length, non-anastomosing linear structures covers most of the lateral plasma membrane surface. These latter structures, unlike known types of cell junctions do not show equivalent arrays in apposing membranes even when the lateral plasma membranes of adjacent cells are closely apposed. The possible function of these structures is discussed.

Invertebrate gap junctions.

Both glycerol and glutaraldehyde, the two most commonly used chemical aids in freeze-fractue studies, have been shown individually to affect the structure of certain membranes as observed in freeze-fracture replicas. The present investigation studied the effect of glycerol on the gap junctions found in a number of tissues from several invertebrate phyla. Glycerol was shown, in some of these tissues, to affect both the spatial arrangement of the particles within the membrane and their fracturing properties if the tissue had not been prefixed with glutaraldehyde. It is suggested that the effect of glycerol on invertebrate gap junctions may be of diagnostic use in enabling them to be separated into a number, or spectrum, of types. Previously B-type gap junctions have been reported only in Arthropoda. This study has extended knowledge of their occurrence into the phyla Coelenterata, Platyhelminthes and Annelida. The B-type gap junctions appear consistently to have a 12-nm particle separation, which is larger than the 10-nm separation found in vertebrate and molluscan A-type gap junctions.

Junctional structures in hydra.

The sealing and communicating junctions present in hydra have been examined using conventional staining, lanthanum tracer and freeze-fracturing techniques. The presence of distinct types of gap and septate junctions has been confirmed. Combined lanthanum tracer and freeze-fracture results have provided a more detailed understanding of these junctional structures. A model has been constructed which demonstrates the various aspects of the junction seen at different sectioning angles. The probable lengths of septa within septate junctions and the junctional 'maze' formed by them is discussed because of its bearing on the 'sealing' nature of the junction and also, to some extent, on its permeability to tracers such as lanthanum.

Goblet cell membrane differentiations in the midgut of a lepidopteran larva.

So-called goblet cells are present in the midgut of lepidopteran larvae. They are thought to be involved in the active transport of potassium out of the haemolymph and into the gut lumen. A number of plasma membrane differentiations within the goblet cell cavity has been investigated using conventional staining, lanthanum tracer and freeze-etch techniques. Of particular interest are junction-like inter- and intra-membrane differentiations found on the villus-like cytoplasmic projections present at the apical tip of the goblet cell cavities. These cytoplasmic projections appear to act as a valve; in some cases they seem to close off the top of the goblet cell cavity, so isolating it from the gut lumen, while in other cases they are spread apart leaving a wide channel from the cavity into the lumen. The junction-like structures on these cytoplasmic projections are different in structure from the septate-type junctions which seal the midgut cells together at their apical borders, and the 2 types are present on the same plasma membrane, often within one micron of each other. The need for a different type of junction may possibly be related to the fact that it occurs between 2 areas of the same plasma membrane. The morphology of this unusual junction-like structure is discussed and 2 diagrams are presented to illustrate our interpretation of its structure.

Junctional structures in the midgut cells of lepidopteran caterpillars.

The junctional structures present between the midgut cells of 3 lepidopteran caterpillars have been examined using freeze-etching, conventional staining and lanthanum tracer techniques. The bonding junction present in this type of tissue is the so-called continuous junction. Septa have only occassionally been reported in conventionally strained cross-sections of these junctions. During the present study septa have been observed in such sections but were more readily located in tissue treated with lanthanum tracer. Tangential sections of lanthanum-impregnated tissue show that these septa are parallel-sided, in contrast to the honeycomb appearance of the septa in septate junctions. The septa in freeze-etch replicas of glutaraldehyde-fixed tissue often show continuous rods on one membrane face, suggesting that the continuous junction may be more akin to the tight junction than to the normal septate junction. However, freeze-etch replicas of unfixed tissue appear much more like replicas of normal septate junctions. The main differences between septate and continuous junctions appear to be that the inter-membrane septa of the continuous junction are parallel-sided in tangential section, as against the honeycomb appearance of the septate junction, and that the particles which delineate the septa in freeze-etched preparations appear to be both somewhat differentl bonded and closer together in the continuous junction. A diagram is presented showing the internal and inter-membrane structures of the 2 types of junction based on the present study, and suggesting a possible explanation of the fact that septa are more readily seen in cross-sections of septate junctions than in continuous junctions. As septa are present in both types of junction, and because their freeze-etch appearances are not very different, it is suggested that the two types of septate junction be called 'smooth septate' and 'pleated septate' junctions, to indicate their characteristic appearances in tangential section.