Morphology of the exocrine glands of the frog skin

Frog skin contains three distinct types of exocrine glands: granular (poison), mucous, and seromucous. The granular gland forms a syncytial secretory compartment within the acinus, which is surrounded by smooth muscle cells. The mucous and seromucous glands are easily identifiable as distinct glands. The serous and mucous secretory cells are arranged in a semilunar configuration opposite the ductal end and are filled with granules. Within the acinus, located at the ductal pole of the gland, are distinct groups of cells with few or no granules in the cytoplasm. In both the mucous and seromucous gland there is a cell type with abundant mitochondria; the one in the mucous gland is located in the region adjacent to the secretory cells. The duct of these glands is two-layered, with the individual cells appearing morphologically similar to the layers of the skin epithelium as the duct traverses the skin. The duct appears to be patent throughout its length. The morphological heterogeneity and distinct distribution of the cell types within the gland acinus may be indicative of a functional heterogeneity that allows the production of distinctly different types of secretion from the same gland type, depending on the type of stimulus.     

Integumentary structure and its relationship to wiping behaviour in the common Indian tree frog, Polypedates maculatus

Skin of the Indian tree frog, Polypedates maculatus (Rhacophoridae), was studied in the context of self-wiping behaviour which functions to expel and distribute cutaneous secretions recently shown to retard evaporative water loss. The secretions contain both mucus and lipids and are derived from a common gland considered to be homologous with characteristic anuran mucous glands. The glands are bipotent and secrete both mucus and lipoid products which are evidently mixed within the glandular lumen. Another type of gland resembling characteristic anuran serous (or granular) glands is found in dorsal but not ventral skin, whereas the lipid-secreting mucous glands are found in skin associated with all body surfaces. There is no distinct, lipid-secreting gland present in the skin of this species other than the mucous glands. These histochemical data complement the earlier finding that resistance to evaporative water loss in this species is relatively small compared with phyllomedusine 'waterproof frogs which also exhibit wiping behaviour associated with secretion of lipids. Thus, wiping behaviour may have evolved in association with mucous secretions before dominant lipoid secretions resulted from strong selection for water conservation.     

Flask cells and flask-shaped glandular cells of amphibian skin specifically produce fucose-rich glycoproteins

A battery of horseradish peroxidase-conjugated lectins has been employed as a cytochemical tool for the labelling of specific cell types in amphibian epidermis. Among the lectins used, onlyUlex europaeus I (UEA I) showed specific reaction with the cytoplasm of flask cells. In addition, UEA I stained flask-shaped secretory cells in dermal glands and a reaction on glandular ductal cells was also observed. At the electron microscopic level, lect-in binding was found in granules distributed among mitochondria in the cytoplasm of flask cells and in larger mucous granules of flask-shaped glandular cells, which were released into the lumen. UEA I also stained the extracellular space above flask cells. The labelling was due mainlty to a glycoprotein of mol. wt. approx. 27 kDa. Structural and cytochemical similarities between flask cells and flask-shaped cells of dermal glands could be a consequence of a common secretory role of both cell types.     

Three types of cutaneous glands in the skin of the salamandrid Pleurodeles waltl. A histological and ultrastructural study

Histological and ultrastructural investigations revealed three different multicellular skin gland types in the salamandrid Pleurodeles waltl. The mucous glands are small, with one layer of secretory cells surrounding a central lumen; they produce the viscous and slippery mucus film that has various functions in amphibians. The serous glands can be divided based on their histological and ultrastructural characters into the granular gland Type I (GGI) and the granular gland Type II (GGII). The first type (GGI) is moderately sized and distributed throughout the body surface, with higher concentrations in the parotoid and back regions. In contrast, the second type (GGII) is very large (for Pleurodeles) and was found only in the tail, with highest concentration in the tail dorsum. Both granular gland types contain mainly proteinaceous materials but differ in their morphological features including size, shape, cellular organization and vesicle distribution, vesicle size and vesicle shape. Both GGI and GGII are especially concentrated in body parts that are presented to an attacking predator and are hypothesized to produce repellent to poisonous substances to thwart potential aggressors. J. Morphol., 2009. © 2009 Wiley‐Liss, Inc.     

Granular cutaneous glands in the frog Physalaemus biligonigerus (Anura, Leptodactylidae): comparison between ordinary serous and 'inguinal' glands

Beside the ordinary granular (or serous) glands, the skin of the leptodactylid frog Physalaemus biligonigerus possesses peculiar clusters of large granular units, the 'inguinal' glands, located in the dorsolateral areas of the pelvic girdle. Both gland types store their specific products within the syncytial cytoplasm of the secretory unit. These secretory materials consist of spheroidal or ellipsoidal bodies (granules) with a repeating substructure. The subcellular features of the immature products of the ordinary serous and inguinal glands are identical. However, these products undergo divergent maturative processes, leading to fluidation on the one hand and condensation on the other. Secretory release into the small gland lumen was observed in both cases, involving merocrine mechanisms. On the basis of the analysis of cutaneous serous gland polymorphism in anurans, the inguinal units in P. biligonigerus do not appear to be an independent line. Rather, these large units belong to the ordinary serous type and represent a gland population specialized in the storage of remarkable amounts of product used in chemical defence of the skin.     

Loss of traditional mucous glands and presence of a novel mucus-producing granular gland in the plethodontid salamander Ensatina eschscholtzii

All amphibians are thought to possess two distinct types of dermal gland: mucous and granular. Mucous glands typically produce an acidic, flocculent and carbohydrate positive secretion that keeps skin moist and helps maintain homeostasis. In this study, we use a battery of histochemical tests to identify these traditional mucous glands in the dermis of the plethodontid salamander Ensatina eschscholtzii . The periodic acid-Schiff, alcian blue, toluidine blue and carmine reactions reveal the absence of both acidic glycosaminoglycans and traditional mucous glands. Instead, E. eschscholtzii produces neutral mucus in specialized granular glands, which we name mucus-producing granular glands (MPGGs). The loss of traditional mucous glands is a derived trait for Ensatina .  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 87 , 469–477.     

Seasonal lectin binding variations of thumb pad in the frog (Pelophylax ridibundus)

The thumb pad is one of the most common secondary sexual characteristics in frogs. Although it is known that amphibian skin has affinity for several lectins, there is no report regarding lectin‐binding affinity of the thumb pad or its structural components. This study investigated localization and seasonal variation of specific carbohydrate moieties of glycoconjugates in both the epidermal and dermal components of the frog thumb pad at the light microscopic level using lectin histochemistry. The study consisted of four seasonal groups of the frog species, Pelophylax ridibundus (Synonym of Rana ridibunda): active, prehibernating, hibernating and posthibernating. Four horseradish peroxidase conjugated lectins were employed. It was found that dolichos biflorus agglutinin (DBA), wheat germ agglutinin (WGA), and ulex europaeus (UEAI) gave positive reactions in both epidermal layers and breeding glands. These three lectins bound specific secretory cells in the breeding glands, and the distribution of the cells and epithelial lectin reactions exhibited seasonal changes. In addition, UEA‐I and peanut agglutinin (PNA) showed an affinity in granular glands and the granular zone of mixed glands. Generally, epidermal lectin binding showed dense affinity during the posthibernation period. DBA, UEA‐I, and WGA‐specific cells in the mucous gland decreased gradually until the posthibernation period. These findings suggest that differences of lectin binding in the thumb pad may be related to functional activities and, thus, seasonal adaptations. Moreover, the presence of specific lectin‐binding cells in the breeding glands indicated that they consisted of heterogeneous secretory cell composition or that the cells were at different secretory stages. J. Morphol. 275:76–86, 2014. © 2013 Wiley Periodicals, Inc.     

Functional morphology of the mammiliform penial glands ofLittorina saxatilis (Gastropoda)

Summary The functional morphology of the mammiliform penial glands ofLittorina saxatilis has been investigated with both light and electron microscopy. These penial glands line the ventral edge of the penis and orient with the female mantle during copulation. Secretions are released from the penial glands to this interface where they probably function in adhesion. The penial gland secretions comprise heterogeneous granules as well as apocrine and mucous secretions. The heterogeneous granules are produced in separate multicellular glands arranged in a series of lobes that lie outside a thick smooth muscle layer enclosing the lumen. Each glandular lobe is surrounded by a thin layer of smooth muscle. Secretions are transported in individual cellular processes that pass through the thick smooth muscle layer and empty into the lumen. Surrounding the lumen is an epithelium containing apocrine secretory cells as well as occasional goblet-type, mucous cells. The combined action of the muscles forces secretions out of the lumen through the penial papilla, onto the external surface of the mammiliform penial gland. Longitudinal muscles extend into the penial papilla enabling its protrusion or retraction. Retraction of the penial papilla following secretion release is thought to create negative pressure beneath the penial gland producing suction adhesion. The visco-elastic properties of the penial gland secretion are qualitatively different from foot mucus and may represent specialization to an adhesive function.     

Morphology,histochemistry and physiology of the major salivary glands in the echidna Tachyglossus aculeatus (Monotremata)

The three major salivary glands of the monotreme echidna are described. The parotid is a typical serous gland with tubulo-acinar secretory endpieces and a well-developed system of striated ducts. The mandibular gland, although light microscopically resembling a mucous gland, secretes very little glycoprotein. Its cells are packed instead with serous granules, resembling in fine structure the “bull's eye” granules in the mandibular gland of the European hedgehog Erinaceus europaeus. The sublingual glands secrete an extremely viscous mucous saliva. Expulsion of this saliva through the narrow ducts is probably aided by contraction of the extensive myoepithelial sheaths surrounding the secretory tubules. Application of the glyoxylic acid induced fluorescence method failed to demonstrate adrenergic innervation in any of the glands.     

Expression pattern of glycoconjugates in the integument of Bufo ictericus (Anuran, Bufonidae): biochemical and histochemical (lectin) profiles

Mucous consists of glycoproteins and proteoglycans produced by specific secretory cells (mucocytes). In anurans the cutaneous mucous is produced by intradermal glands and displays both mechanical and chemical protection functions. Indeed, mucous maintains the integument moist and facilitates gas exchange (cutaneous respiration). In this work, the carbohydrate moiety distribution was investigated in the integument of Bufo ictericus using conventional and lectin histochemistry to describe the pattern of cutaneous glycoconjugate expression, including both secretory and structural proteoglycans. As a preliminary step, the descendent chromatography in Whatmann 1MM paper was undertaken to prepare the histochemical trials involving the lectins. In B. ictericus, the integument exhibits the basic morphological structure found in lower terrestrial vertebrates: the epidermis is a keratinized squamous stratified epithelium supported by spongious and compact layers. The spongy dermis contain secretory portion of both mucous and serous (or poison) glands. The paper chromatography identified galactose, fucose and mannose as characteristic sugar residues. The secretory cells of the mucous gland in the dermis, as well as the interstice between the stratum corneum and the subjacent stratum spinosum in the epidermis exhibit alpha-l-fucose and alpha-galactose residues. The serous glands give no reaction. The alpha-mannose residue was detected in the extracellular matrix of spongious dermis, but not in the dermal glands. The different glycoconjugate location reflects in two glycoconjugates categories: the secretory which participate in the water flow regulation, and the structural which is involved in the dermal maintenance.     

Ultrastructural aspects of cat submandibular glands

Submandibular glands of five adult female cats were examined by conventional electron microscopic techniques. All gland acini are mucous secreting and each acinus is capped with mucous secreting demilunar cells. Secretory product of demilunar cells is more electron lucent than that of acinar cells. The demilunes show intercellular tissue spaces and intercellular canaliculi whereas similar specializations are absent between acinar cells. Mitochondria and arrays of granular endoplasmic reticulum are more numerous in demilunar cells than in acinar cells. In acinar and demilunar cells secretory droplets first appear as enlarged Golgi saccules which subsequently become closely related to cisternae of the granular endoplasmic reticulum. Filamentous structures, interpreted as mucin molecules, are present in secretory droplets of acinar cells. Intercalated ducts are short, consisting of several junctional cells between acini and striated ducts. Striated ducts are long and tortuous and contain light cells, dark cells and basal cells. Light cells contain numerous membrane bound granules in their distal ends whereas dark cells show electron lucent vesicles in the same position. Basal cells contain a paucity of organelles and membrane plications but exhibit hemidesmosomes along their basal plasma membranes. Myoepithelial cells are abundant in relation to acinar and demilunar cells. Nerve terminals are present in some instances between acinar cells or between acinar and myoepithelial cells.     

Cytochemistry of the skin of patients with mucopolysaccharidoses.

The distribution of complex carbohydrates has been investigated at the light and electron microscope levels in sweat glands of normal subjects and patients with Hurler's or Hunter's disease. Normal sweat glands examined with a battery of light microscopic histochemical methods revealed sulphated complex carbohydrate in secretory granules of the dark cells. These granules lacked affinity for dialysed iron (DI) at the light and electron microscope levels. The DI method demonstrated acid complex carbohydrates ultrastructurally on the surface of the intercellular canaliculi and central lumen in normal sweat glands. DI-reactive acidic material, presumably of mucopolysaccharide nature, surrounded and extended between collagen bundles in the stroma of normal skin, but was absent from the band which ensheathed the sweat gland and consisted of individual rather than bundled collagen fibrils. DI-reactive mucopolysaccharide lined and partially filled vacuoles of dark cells showing a laminar distribution in vacuoles of clear cells in sweat glands of a Hunter patient. The DI method also visualized mucopolysaccharide distributed throughout vacuoles in fibroblasts of this patient. DI-reactive acid material covered the luminal surface of the sweat gland, coated collagen bundles in the stroma and spared the periglandular collagenous sheath in skin from Hurler and Hunter patients as in that from normal controls. Acid phosphatase was localized ultrastructually in vacuoles and nearby cytoplasm and on plasmalemmae of clear cells, dark cells and myoepithelial cells of sweat glands from Hurler and Hunter patients. Vacuoles of dermal fibroblasts and Schwann cells in these specimens also exhibited strong acid phosphatase activity.     

Innervation of skin glands in the frog

Summary A comparative study was undertaken on the innervation of mucous and granular glands in frog skin. Results obtained by the Falck-Hillarp fluorescence technique and cholinesterase staining indicated that both types of glands receive exclusively adrenergic innervation. Electron microscopy was used to investigate the innervation pattern at the ultrastructural level. The distribution of nerve terminals was found to differ in the two types of glands. In the mucous gland, terminals were found at a distance of about 0.5 m from the basement membrane but never within the gland parenchyma. In the granular gland, the terminals were located between smooth muscle cells and also in direct contact with the secretory epithelium but never outside the basement membrane.This work was carried out in part at King Gustaf V:s Forskningsinstitut, Stockholm, and was supported by a grant from Karolinska Institutet     

Localization of prolactin receptor in the dorsal and ventral skin of the frog (Rana ridibunda)

The dorsal and ventral skin in amphibians plays an important role in osmoregulation. Prolactin hormone is involved in regulation of amphibian skin functions, such as water and electrolyte balance. Therefore, amphibians may be useful as a model for determining the sites of the prolactin receptor. In this study, prolactin receptor was detected in frog dorsal and ventral skin using immunohistochemical staining method. Prolactin receptor immunoreactivity was localized in all epidermal layers except stratum corneum of dorsal skin epidermis, stratum germinativum layer of ventral skin epidermis, myoepithelial cells, secretory epithelium and secretory channel cells of granular glands in both skin regions. The mucous glands and secretory granules of granular glands did not show immunoreactivity for the prolactin receptor. According to our immunohistochemical results, the more widespread detection of prolactin receptor in dorsal skin epidermis indicates that prolactin is more effective in dorsal skin. Presence of prolactin receptors in epidermis points out its possible osmoregulatory effect. Moreover, detection of receptor immunoreactivity in various elements of poison glands in the dermis of both dorsal and ventral skin regions suggests that prolactin has a regulatory effect in gland functions.     

The morphology of the mucous gland and its responses to prolactin in the skin of the red-spotted newt

The mucous gland of the red-spotted newt, Notophthalamus viridescens viridescens, Rafinesque was examined by histochemical and ultrastructural techniques and its cytological responses to various hormonal conditions were studied. Its secretory epithelial cells produce and release in merocrine fashion a neutral, unsulphated mucosubstance. The secretory epithelium is bounded peripherally by a thin, but apparent non-functional, myo-epithelium. The duct of this mucous gland consists of a single keratinized tubular cell that extends from the neck region of the gland to the surface of the epidermis. Mucous secretion is absent or greatly reduced on the skins of newts maintained under laboratory conditions for a few weeks but reappears after injection of ovine prolactin. Mucous glands in laboratory conditioned animals show a 4-fold increase in volume brought about by the engorgement of their epithelial cells with secretory granules. Ovine prolactin reduces the volume of the glands to unconditioned levels with a corresponding reduction in granular content, suggesting that prolactin functions in the release of the granules. This view is reinforced by the findings that autotransplantation of the pituitary gland prevents the conditioning effect and that glandular volume increases in auto-transplanted animals given ergocornine. Granular accumulation begins also in hypophysectomized newts but ceases after a week, indicating the need for some hypophyseal factor in the synthesis as well as the release of the granules. Ovine prolactin restores mucous glands of hypophysectomized newts to the unconditioned state. Contrary to earlier findings, ovine prolactin induces a reduction in the volume of the mucous gland in thyroidectomized newts.     

Anatomy and ultrastructure of dermal glands in an adult water mite,Teutonia cometes (Koch, 1837) (Acariformes: Hydrachnidia: Teutoniidae)

Organization of dermal glands in adult water mites Teutonia cometes (Koch, 1837) was studied using light-optical, SEM and TEM methods for the first time. These glands are large and occur in a total number of ten pairs at the dorsal, ventral and lateral sides of the body. The slit-like external openings of the glands (glandularia) are provided with a cone-shaped sclerite, and are combined with a single small trichoid seta (hair sensillum), which is always situated slightly apart from the anterior aspect of the gland opening. Each gland is formed by an epithelium encompassing a very large lumen (central cavity) normally filled with secretion that stains in varying intensity on toluidine blue stained sections. The epithelium is composed of irregularly shaped secretory cells with an electron-dense cytoplasm and infolded basal portions. The cells possess a large irregularly shaped nucleus and are filled with tightly packed slightly dilated cisterns and vesicles of rough endoplasmic reticulum (RER) with electron lucent contents. Dense vesicles are also present in the apical cell zone. Some cells undergo dissolution, occupy an upper position within the epithelium and have a lighter cytoplasm with disorganized RER. Muscle fibers are regularly present in the deep folds of the basal cell portions and may serve to squeeze the gland and eject the secretion into the external milieu. The structure of these dermal glands is compared with the previously described idiosomal glands of the same species and a tentative correlation with the glandularia system of water mites is given. Possible functions of the dermal glands of T. cometes are discussed.     

Immunocytochemical localization of renin in the submandibular gland of the mouse.

Renin was localized in the submandibular gland of the adult mouse at light and electron microscopic levels by the unlabeled antibody enzyme method of Sternberger. At the light microscopic level, renin was confined to the granular convoluted tubule (GCT) segment of the gland with considerable variation among GCT cells in intensity of staining. Some GCT cells failed to stain for renin. The pattern of staining was the same in the gland of male and female mice, but in the glands of females GCT segments were smaller and less numerous. At the electron microscopic level, staining for renin was also confined to the GCT cells, and was localized exclusively to the secretory granules. The intensity of staining of the secretory granules within a given GCT cell varied; some cells contained only minimally reactive or negative secretory granules. All other organelles within the GCT cell, except condensing vacuoles, failed to stain.     

New insights into sexually dimorphic skin glands of anurans: The structure and ultrastructure of the mental and lateral glands in Hypsiboas punctatus (Amphibia: Anura: Hylidae)

Many anuran species are characterized by sexually dimorphic skin glands. These glands often are concentrated on specific areas, such as the mental region, flanks, or the nuptial pads. We studied the histology and histochemistry of mental and lateral glands in Hypsiboas punctatus, and compared them to skin from other body regions. We describe four types of dermal glands, two types of mucous and two types of serous glands. The mucous glands are formed by a single layered epithelium. The mucocytes surrounding a central lumen are filled with polyhedral granules. Ordinary mucous glands are small sized glands with cubical epithelium, mucoid content, and small granules. Specialized mucous glands are characterized by a larger size, a columnar epithelium, a proteinaceous content and larger granules. Both types of serous glands are syncytial and share some structural features including size, shape, and morphology of secretory granules. However, ordinary and specialized serous glands differ in their histochemical properties, size and appearance of secretory granules, and glandular outlets. The specialized type of mucous glands in H. punctatus resembles most SDSGs described in anurans, whereas the presence of specialized serous glands that are sexually dimorphic is less common. Both specialized glands occur only in mental and lateral regions of males, whereas ordinary mucous and ordinary serous glands occur in males and females. J. Morphol. 2012. © 2012 Wiley Periodicals, Inc.     

Morphological characterization of the glandular system in the salamander Plethodon shermani (Caudata,Plethodontidae)

Amphibians have evolved a wide variety of mechanisms that provide a certain degree of protection against predators, including camouflage, tail autonomy, encounter behavior and noxious or toxic skin secretions. In addition to these strategies, some amphibians release a glue-like secretion onto the surface of their skin when threatened. While some information regarding the origin and production of these adhesive secretions is available for frogs such as Notaden bennetti, these aspects are only partially understood in salamanders. We contribute to an earlier study and provide additional information regarding the origin, production, and characterization of the adhesive secretion in the red-legged salamander (Plethodon shermani) at a microanatomical level. When stressed, this salamander secretes a milky, viscous liquid from its dorsal and ventral skin. This secretion is extremely adhesive and hardens within seconds upon exposure to air. This study describes two cutaneous gland types (mucous and granular) in the dorsal and ventral epithelial tissue that differ considerably in their secretory content. While the smaller mucous glands contains flocculent to granular material, mostly acidic glycoproteins, the granular glands synthesize various granules of differing size and density that consist of basic proteinaceous material. The results strongly indicate that the secretions of both gland types from the dorsal as well as the ventral side form the adhesive mucus in Plethodon shermani, consisting of basic and acidic glycoproteins, glycoconjugates with mannose and α-l-fucose residues as well as lipid components.     

Distribution of prolactin receptor in frog (Rana ridibunda) dorsal skin during hibernation

The role of prolactin in the regulation of frog skin functions is still unclear particularly during environmental changes. In this study, prolactin receptor (PRLR) was detected in active and hibernating frog dorsal skin using immunohistochemical method. PRLR immunoreactivity in active frogs was observed in the epidermis, in the secretory epithelium of granular glands and the secretory channel cells of the glands. Myoepithelial cells of granular glands that started accumulating secretory material or those with a full lumen were PRLR immunoreactive, while some myoepithelial cells of empty granular glands were negative for PRLR. In hibernating frogs, this immunoreactivity was observed in the same regions; however, immunoreactivity was more intense than that in active frogs. PCNA was employed for detection of proliferative activity of PRL in the dorsal skin, and immunoreactivity was detected in the nuclei of a few epidermis cells and in the duct of glands of active frogs. The number of immunoreactive nuclei in these regions increased in hibernating and in prolactin injected groups. We conclude that prolactin provides morphological and functional integrity of skin stimulating the proliferation and regulating the function of granular glands and plays an important role in the adaptation of amphibians to the long winter period.