CELL JUNCTIONS IN AMPHIBIAN SKIN

Cell junctions have been investigated in the amphibian epidermis, a stratified squamous epithelium, and compared to those described previously in simple columnar epithelia of mammalian cavitary organs. In adult frogs and toads, and in larvae approaching metamorphosis, belts of membrane fusion or zonulae occludentes of considerable depth are regularly found between adjoining cells of the outermost layer of the stratum corneum, binding the cells together into a continuous, uninterrupted sheet. Another set of occluding zonules appears in the second cornified layer (when such a layer is present), and a third set usually occurs in the outermost layer of the stratum granulosum. Specialized elements described as "modified" and "composite" desmosomes are encountered along the lateral and basal aspects, respectively, of the cornified cells; ordinary desmosomes and maculae occludentes (i.e., spots of membrane fusion) are found in all other strata. The usual 200 A intercellular gap is generally maintained between the cells of the stratum germinativum at the basal ends of the intercellular spaces. Hence, the intercellular spaces of the epidermis form a largely continuous network, closed to the external medium and open to the dermal interstitia. The situation is comparable to that found in columnar epithelia, except that the intercellular spaces are much more extensive, and an extracellular subcompartment (or two) apparently exists in the stratum corneum and between the latter and the stratum granulosum. The last subcompartment is usually filled with a dense substance, probably derived from discharged secretory granules. The tripartite junctional complex characteristic of lumen-lining epithelia (i.e., a zonula occludens followed by a zonula adhaerens, and desmosome) is seen only in early larvae; in adults and in larvae approaching metamorphosis, the occluding zonule is followed directly by a series of modified desmosomes. Interpreted in the light of current physiological data, these findings suggest that the diffusion of water, ions, and small, water-soluble molecules is impeded along the intercellular spaces of the epidermis by zonulae occludentes while it is facilitated from cell to cell within the epidermis by zonulae and maculae occludentes.     

A comparative study on vascularization and the structure of the epidermis of an amphibious mudskipper fish,Scartelaos gigas (Gobiidae,Teleostei), on different parts of the body and the appendages

Epidermal structure of the amphibious mudskipper, Scartelaos gigas (Gobiidae), was investigated in relation to their terrestrial adaptation whereby a histological study on the epidermis of 15 regions including nine body regions, five fins and the sucking disc was carried out. The structure of the epidermis consists of three layers: an outermost layer with polygonal cells or rather flattened cells, small cells and mucous cells; a thick middle layer with voluminous cells swollen by epidermal cells; and the stratum germinativum. A dermal bulge was located at each apical area of the epidermis of almost all body regions, but was not existent in the operculum and the appendages, including none of the fins or the sucking disc. In the epidermis of the body regions, the dermal bulges had numerous dermal capillaries just beneath the stratum germinativum. By contrast, the appendages never had dermal capillaries due to the absence of the dermal bulge. Based on these results, the cutaneous air uptake in S. gigas would seem to be more effective in the upper body regions that are most often exposed to air than in the lower body regions, however, cutaneous air uptake is not likely to occur in the appendages.     

The ultrastructure of the integument of the American eel,Anguilla rostrata

Summary The morphology and ultrastructure of the lateral body integument of the leptocephalus, glass eel, pigmented elver, and adult stages of the American eel, Anguilla rostrata, were examined with light and electron microscopy. The integument consists of an epidermis separated by a basal lamina from the underlying dermis. Three cell types are present in the epidermis in all stages. Filament-containing cells, which are the principal structural cell type, are increasingly numerous at each stage. Mucous cells, which secrete the mucous that compose the mucous surface coat, are also more numerous in each subsequent stage and are more numerous in the anterior lateral body epidermis than in the posterior lateral body epidermis of the adult. Club cells, whose function is unknown, are most numerous in the glass eel and pigmented elver. Chloride cells are common in the leptocephalus which is marine and infrequent in the glass eel. They are not present in the pigmented elver and adult which inhabit estuaries and fresh-water. Lymphocytes and melanocytes are also present in some stages. The dermis comprises two layers: a layer of collagenous lamellae, the stratum compactum, and an underlying layer of loose connective tissue, the stratum spongiosum.There is a progressive increase in epidermal thickness at each stage which is paralleled by an increase in the thickness of the stratum compactum. Rudimentary scales are present in the dermis of the adult. The increase in the number of epidermal filament-containing cells, epidermal thickness and stratum compactum thickness is correlated with an increased need for protection from abrasion and mechanical damage as the eel moves from a pelagic, oceanic habitat to a benthic, freshwater habitat. The increase in mucous cell numbers is likewise correlated with an increased need for the protective and anti-bacterial action of the mucous surface coat in the freshwater environment.This investigation was supported by NIH research grant NS-11276 from National Institute of Neurological Diseases and Stroke to Dr. J.D. McCleave and by N.S.F. Grant GD 38933 to the Bermuda Biological Station, St. Georges West, Bermuda. Bermuda Biological Station Contribution No. 668     

Fine structure of the epidermis of the mudskipper,Periophthalmus modestus (Gobiidae)

The ultra-structure of the epidermis of the mudskipper,Periophthalmus modestus, was examined by both light and transmission electron microscopies. The epidermis is exceptionally not well endowed with mucous or granular cells. Filament-containing cells occur in three distinct layers of the surface, middle and basal epidermis. The surface layer is further subdivided into two layers, an outermost and less superficial one. Two different cell types were identified in the epidermis. Type I cells are fiat cells in a single stratum. Type II cells are enormous cells, characterized by having a large vacuole in the cytoplasm. The outermost layer is composed of a free surface of Type I cells and numerous microridges covered with a fuzzy, fibrillar substance. The “fuzz” forms a cuticule-like structure, but keratinization as found in terrestrial animals does not occur. The superficial layer contains Type I cells and intraepithelial blood capillaries. When Type I cells become senescent, numerous intercellular spaces are formed in the plasma membranes of adjacent cells, with the senescent cells finally falling off. Just beneath these cells, however, young cells of Type I are always found. The blood capillaries are usually reinforced with young Type I cells. A large volume of oxygen may be absorbed through the skin using the blood capillary network. The middle layer contains several strata of Type II cells. The special corky structure of these cells seems to play an important role in thermal insulation and protection against ultraviolet light in relation to life out of water. However, by comparison with terrestrial animals, the histological design of the epidermis of this goby appears incomplete, so as to reduce desiccation on land, owing to the epidermis lacking a keratinized stratum. The differentiation of the epidermis seems to be an adaptation for a terrestrial habit in this species.     

Structure and Histochemistry of the Skin of a Torrent Catfish, Liobagrus mediadiposalis

The epidermis of the torrent catfish, Liobagrus mediadiposalis, consists of three layers: the outermost layer, middle layer and stratum germinativum. The epidermis consists of two types of skin glands, small mucus cell and voluminous club cell. The unicellular mucus cell contains acid sulfomucins (some sialomucins) and the club cell, sometimes binucleate, is proteinaceous. Well-developed vascularization is one of the characteristics of epidermis of L. mediadiposalis. Well-developed lymphatic spaces contain lymphocytes in the epidermis. The dermis lacks scales and consists mostly of a thick, dense connective tissue; its superficial region just below the basal membrane is supplied with fine blood capillaries. These histological features of the skin in L. mediadiposalis are consistent with that required for cutaneous respiration.     

Light and electron microscopic observations on the normal structure of the vomeronasal organ of garter snakes

The vomeronasal epithelium of adult garter snakes (Thamnophis sirtalis and T. radix) was studied by light and electron microscopy. The sensory epithelium is extraordinarily thick, consisting of a supporting cell layer, a bipolar cell layer, and an undifferentiated cell layer. The supporting cell layer is situated along the luminal surface and includes supporting cells and the peripheral processes (dendrites) of bipolar neurons. The luminal surfaces of both supporting cells and bipolar neurons are covered with microvilli. Specializations of membrane junctions are always observed between adjacent cells in the subluminal region. Below the supporting cell layer, the epithelium is characterized by a columnar organization. Each column contains a population of bipolar neurons and undifferentiated cells. These cells are isolated from the underlying vascular and pigmented connective tissue by the presence of a thin sheath of satellite cells and a basal lamina. Heterogeneity of cell morphology occurs within each cell column. Generative and undifferentiated cells occupy the basal regions and mature neurons occupy the apical regions. Transitional changes in cell morphology occur within the depth of each cell column. These observations suggest that the vomeronasal cell column is the structural unit of the organ and may represent the dynamic unit for cell replacement as well. A sequential process of cell proliferation, neuronal differentiation, and maturation appears to occur in the epithelium despite the adult state of the animal.     

Structure of the skin of an air-breathing mudskipper, Periophthalmus magnuspinnatus

The epidermis of the mudskipper Periophthalmus magnuspinnatus consisted of three layers: the outermost layer, middle layer and stratum germinativum. Extensive vascular capillary networks were present near the superficial layer of epidermis and outermost layer. The diffusion distance between the vascular capillaries and the surface of epidermis was c . 1.5 ± 0.9μm. The middle layer consisted of small or voluminous cells swollen by epidermal cells. Due to the swollen cells, the thickness of the epidermis increased and the epidermis appeared web-like. The swollen cells contained tonofilaments, lucent contents and desmosomes. Fine blood capillaries were also discernible in this layer. Well-developed lymphatic spaces containing lymphocytes existed in the stratum germinativum. Numerous blood capillaries were present under the basement membrane. The dermis consisted of a stratum laxum and stratum compactum, and there was a definite area with acid mucopolysaccharides and a small scale in the stratum laxum. The skin had an epidermal pigment cell, dendritic melanophores (-cytes) containing melanin granules within their cytoplasm, and two kinds of dermal pigment cells, melanophores and colourless pigments containing reflecting platelets.     

Structure and development of spines over the skin surface of the river puffer Takifugu obscurus (Tetraodontidae,Teleostei) during larval growth

The histological structure and development of spines on the skin surface of Takifugu obscurus were studied during larval development conducted artificially with an average 30‰ salinity and 18.0–20.3°C water temperature. The epidermis comprises an outermost layer, middle layer, and the stratum germinativum, and contains three types of gland cells: small spherical or flask‐shaped mucous cells, larger sacciform mucous cells, and large granular cells. The dermis and subcutis follow. The spines first appear over the ventral region at 10 days after hatching and consist of two parts: a central long tapering portion which projects into the epidermis and eventually outside of the body, and a short supporting basal portion that is embedded within the stratum compactum layer of the dermis. The central, long tapering portion has two very short processes on top until 25 days after hatching, but these two separate spines fuse into one 30 days after hatching. In contrast, the short supporting spines rooted at the base consist of three to six small spines (usually four to five spines) and are present even in the adult stage. Therefore, calcareous spines consisting of one central long spine and three to six smaller supporting spines form tetra‐ and septaradiate spines (mainly penta‐ and hexaradiate). The spines first appear over the ventral region.     

Cellular responses of the skin and changes in plasma cortisol levels of trout (Oncorhynchus mykiss) exposed to acidified water

The skin structure and the plasma cortisol levels of trout, Oncorhynchus mykiss, were examined during 7 days of exposure to water of pH 5. By day-4 and-7, the thickness of the epidermis was significantly (P<0.05) less in acid exposed fish than in controls, and degenerative cells were common in the upper epidermal layers. Many epidermal cells exhibited signs of necrosis, and by day-7 many apoptotic cells were also present. Secretory vesicles of high electron density were abundant in the filament cells of the 3–4 outermost layers of epidermis, and intercellular spaces had increased. Mitotic figures occureed throughout the epidermis, with the exception of the outermost cell layer. Mucous cells became elongated after day-1, and later, newly differentiating mucous cells could be seen close to the skin surface, and many mucocytes contained mucosomes of high electron density. Rodlet cells were occasionally seen. Chloride cells appeared similar to those of control fish. Many leucocytes, mainly macrophages and lymphocytes, had penetrated the epidermis via the highly undulating basal lamina, and at day-7, numerous apoptotic lymphocytes were found. In the dermis, melanosomes became dispersed in the cytoplasmic extensions of melanocytes which were present in the epidermis of all acid-exposed fish. Iridocytes were rate after day-4, while fibroblasts were abundant and secreted large amounts of collagen. After 1 day of exposure to acidified water, a significant (P<0.05) elevation of the plasma cortisol level had occurred, but this subsequently declined, and had returned to control values by day-7. The changes in skin structure, however, remained throughout the whole exposure period.     

Characterization of metamorphic changes in anuran larval epidermis using lectins as probes

The skin of an adult frog of Xenopus laevis was characterized by the reactivity of 20 lectins. The lectins were classified into six groups in their binding to the epidermal cells: Lycopersicon esculentum lectin (LEL)-type which was positive for all epidermal cells; Pisum sativum agglutinin (PSA)-type for stratum germinativum; succinylated wheat germ agglutinin (sWGA)-type for strata spinosum, granulosum and corneum; Dolichos biflorus agglutinin (DBA)-type for strata germinativum and spinosum; peanut agglutinin (PNA)-type for stratum spinosum; and Ulex europaeus agglutinin (UEA-I)-type for strata granulosum and corneum. PSA and sWGA were utilized as markers of mitotically active germinative cells and the differentiated cells of the epidermis, respectively, to describe the metamorphic conversion of larval epidermal cells to adult type. PSA stained all epidermal cells of tadpoles before metamorphic climax. At the end of metamorphosis, PSA-positive cells were restricted to cells in the basal layer of body epidermis while all the tail epidermis remained PSA-positive. The other cell marker, sWGA, only stained apical cells in tadpole epidermis. During the metamorphic climax, sWGA-positive cells appeared in the cells beneath the stratum corneum of the body region, but not in the tail region. The present study demonstrates that PSA and sWGA are useful to investigate metamorphic changes in tadpole epidermal cells.     

Formation of desmosomes and other contact specializations in cultured skin of the frog (Rana esculenta)

Summary Small trypsinized explants from ventral skin of frogs (Rana esculenta) were maintained in culture for 4 days during which a newly formed epithelium differentiated along the cut edges of the dermis. During the first 6 h adjacent cells produced numerous interdigitating lamellipodia. After 2 days, epithelial polarity was restored by the formation of zonulae occludentes and the epithelial cells were joined by a few small newly formed desmosomes and by numerous interdigitations. Bipartite junctional complexes consisting of a zonula occludens, followed by a series of typical desmosomes, and characteristic of adult frog epidermis were formed only after 4 days. When cultured in the presence of an inhibitor of protein synthesis (cycloheximide) the trypsinized epidermis no longer formed desmosomes. Therefore pools of one or more crucial desmosomal proteins must be very low or non-existent. However, cycloheximide did not prevent the formation of cell contact specializations, consisting of a highly developed system of complex lamellar interdigitations, between adjacent cells.     

Comparative study of the olfactory epithelium of the three-spined stickleback (Gasterosteus aculeatus) and the nine-spined stickleback (Pungitius pungitius)

Summary The olfactory epithelium of the three-spined stickleback (Gasterosteus aculeatus) and the nine-spined stickleback (Pungitius pungitius) has been studied with a conventional histochemical and a novel immunological staining technique. In both species, the sensory epithelium is arranged in folds separated by non-sensory epithelial tissue. In the nine-spined stickleback, intrinsic folds consisting of non-sensory cells are found in the apical part of the sensory epithelium where they divide the surface of the sensory epithelium into small islets. These non-sensory cells are non-ciliated, flattened and piled on top of each other; they contain numerous electron-translucent vesicles. The intrinsic folds are absent from the sensory epithelium of the three-spined stickleback. In both species, axons of receptor cells form a layer of fibers in the sensory epithelium immediately above the basal cells. In the three-spined stickleback, thick branches of the olfactory nerve are frequently found in this layer. These branches are only occasionally observed in the sensory epithelium of the nine-spined stickleback. Thus, the three-spined stickleback and the nine-spined stickleback show considerable differences in the organization of the sensory regions of the olfactory epithelium.     

A study on the vascularization and structure of the epidermis of the air-breathing mudskipper, Periophthalmus magnuspinnatus (Gobiidae, Teleostei), along different parts of the body

A histological study on the epidermis of eight body regions, five fins and the sucking disc was performed on the mudskipper, Periophthalmus magnuspinnatus. The study aimed to determine the role of the skin in respiration and to assess which region of the skin was most effective. The structure of the epidermis, consisting of the superficial layer, middle layer and the stratum germinativum, was the same in all regions. Large numbers of blood capillaries were situated at the superficial layer and occasionally at the middle layer. The mean diffusion distance between the capillary endothelial cells and the surface of the epidermis ranged from 2.6 to 15.4 μm: the lowest value was on the back (mean 2.0 μm) and the highest value was at the base of the anal fin (mean 15.4 μm). Relative surface area of respiratory epithelium in 14 regions was highest in the 1st and 2nd dorsal fins with a thinner epidermis and a lower diffusion distance (mean value 3.2% and 2.5% respectively), whereas the lowest was found at the base of the anal fin (mean 0.7%). Among the 14 regions of the epidermis, it can be surmised that the two dorsal fins toward the upper region may often be more exposed to air and for longer time periods than the other body regions during the amphibious life phase of Periophthalmus magnuspinnatus.     

Changes in structure of ventral epidermis of Rana ridibunda during metamorphosis

Summary The organisation of the ventral epidermis organisation was followed throughout ontogenesis in Rana ridibunda. Epidermis of tadpoles with 2–3 limbs was organised into two layers: a stratum germinativum consisting of elongated columnar cells, and an outer stratum corneum consisting of two types of cuboid cells. Two types of cells can be distinguished; they are a light (clear) cell and a dark (dense) cell. In the 4-legged tadpoles the stratum corneum cells start to flatten and a replacement layer appeared underneath. A well-defined stratum germinativum is found and within it, epidermal glands. Moulting took place for the first time in tadpoles just before metamorphosis, and a well-organised stratum granulosum was formed still containing the two main types of epidermal glands. The flask cells appear in the juveniles for the first time, greatly increasing in numbers in the adult epidermis.     

Keratinization of the outer surface of the avian scutate scale: Interrelationship of alpha and beta keratin filaments in a cornifying tissue

Summary The outer surface of adult Gallus domesticus scutate scale was studied as a model for epidermal cornification involving accumulation of both alpha and beta keratins. Electron-microscopic analysis demonstrated that the basal cells of the adult epidermis contained abundant lipid droplets and that filament bundles and desmosomes were distributed throughout the cell layers. Indirect immunofluorescence microscopy and double-labeling immunogold-electron microscopy confirmed that the stratum germinativum contained alpha keratin but not beta keratin. Beta keratins were first detected in the stratum intermedium and were always found intermingled with filament bundles of alpha keratin. As the differentiating cells moved into the outer regions of the stratum intermedium and the stratum corneum, the large mixed keratin filament bundles labeled increasingly more with beta keratin antiserum and relatively less so with alpha keratin antiserum. Sodium dodecyl sulfate-polyacrylamide gel analysis of vertical layers of the outer surface of the scutate scale confirmed that cells having reached the outermost layers of stratum corneum had preferentially lost alpha keratin. The mixed bundles of alpha and beta keratin filaments were closely associated with desmosomes in the lower stratum intermedium and with electron-dense aggregates in the cytoplasm of cells in the outer stratum intermedium. Using anti-desmosomal serum it was shown that these cytoplasmic plaques were desmosomes.     

Functional organization of the skin of the ‘Green-puffer fish’Tetraodon fluviatilis (Ham.-Buch.) (Tetraodontidae,Pisces)

Summary The present study concerns the functional organization of the skin ofTetraodon fluviatilis. The epidermis consists of five different types of cells — the flask-shaped mucous cells, the eosinophilic granular cells, the sacciform granulated cells, the vesicle containing granulated cells, and the polygonal cells. A thin noncellular layer, the cuticle found on the surface of the skin, is probably secreted from the polygonal cells in the outermost layer of the epidermis. A,well-defined lymphatic plexus exists between the cells of the basal layer.Numerous triradiate calcareous spines are embedded within elastic connective tissue pockets in the thick dermis. These pockets are filled with an amorphous, acellular, PAS positive material, and are richly supplied with fine blood capillaries. A histomorphologic basis for the erection of the spines and various structural modifications in the skin facilitating its enormous stretching under inflated conditions of the fish are discussed.Abbreviations Used BCA blood capillary - BM basement membrane - BC basal cell - BL basal layer - CFB collagen fiber bundle - CTB connective tissue band - DER dermis - EGC eosinophilic granular cell - EPD epidermis - FB fibroblasts - FC fat cell - L lymphocyte - LS lymphatic space - MC mucous cell - ML middle layer - MUS muscle - MYS myocommata - NV nerve - OL outermost layer - PCB black pigment cell - PCY yellow pigment cell - PEC polygonal epidermal cell - SCT subcutis - SGC sacciform granulated cell - SP spine - STC stratum compactum - STL stratum laxum - VGC vesicle containing granulated cell - VS vertical strand This investigation was supported by a research grant No. 38(131)/72-GAU-II from the Council of Scientific and Industrial Research and a financial assistance grant for teachers No. F. 6(4626) 72-(SF-1), from the University Grants Commission, Government of India, New Delhi.     

Ultrastructure of the colon of Locusta migratoria

The ultrastructure of the colon of Locusta migratoria is described. The colon is lined by a thick cuticle that, for the most part, adheres to the underlying epithelium. The cuboid epithelial cells are characterized by moderate invaginations of the apical and, to a lesser extent, basal plasma membranes; the lateral plasma membranes are relatively flat. The bulk of the mitochondria are located in the apical region of the cell and are not particularly associated with any of the plasma membranes. The basal region of the cells contains much rough endoplasmic reticulum, glycogenlike granules, and a predominance of spherical, electron-dense bodies of various sizes. Where muscle fibers make contact with the epithelium, the cells are much reduced; the cytoplasm is usually less electron-dense, and, typically, the nucleus has a thick layer of granular material associated with the inner nuclear membrane. The apical and basal plasma membranes of the reduced epithelial cells contain numerous hemidesmosomes. The apical hemidesmosomes occur in pairs around an extracellular space that contains electron-opaque material. The latter forms tonofibrillae that extend into the endocuticle. Bundles of microtubules are associated with the hemidesmosomes. The tubules traverse the cell from the apical to the basal region. The possible significance of these findings is discussed.     

Histology and Ultrastructure of the Body Wall in the Phoronid Phoronopsis harmeri

The histology and ultrastructure of the body wall in Phoronopsis harmeriwere studied using light microscopy and TEM. The ectoderm epithelium of tentacles, anterior body region, and ampulla consists of monociliary cells. Gram-negative bacteria were found between microvilli, in the protocuticle of the anterior region, and in the ampulla. The epithelium of the posterior body region lacks both monociliary cells and bacteria. The bundles of nerve fibers run between the layer of epithelial cells and basal membrane. The musculature of the body wall comprises circular and longitudinal muscles. The circular muscle fibers are applied to the basal membrane and constitute a solid layer extending almost throughout the length of the body. This pattern is broken in the posterior body region, where there is no solid layer of circular musculature, and the latter is arranged in isolated muscle bands. In the ampullar (terminal) body region, the inversion of circular and longitudinal muscle layers takes place, so that the latter appears to be pressed against the basal membrane. The apical surfaces of longitudinal muscle cells bear cytoplasmic processes; some of the cells have a flagellum. The basal portion of the longitudinal muscle cells forms a cytoplasmic process containing bundles of tonofilaments. The processes of all cells making up the muscle bands are interwoven and anchored to the basal membrane.     

Ionic gradients within mantle-collar epithelial cells of the land snail Otala lactea

Summary Aestivating Otala lactea have been shown to reduce the rate of evaporative water loss from the cells of the mantle-collar epithelium to a level comparable to that of an insect. X-ray microanalyses of ultrathin frozen sections from aestivating and non-aestivating snails have shown gradients of chloride and potassium ions in the apical microvillus region of the regulating mantle collar epithelium. The greatest difference in osmotic concentration occurs in the apical 2 m of the cell. There appears to be a barrier at that level that prevents water being mobilised from the underlying tissues. Methods of presenting data generated by X-ray microanalysis are also discussed.