Giovanni Delfino

Serous cutaneous glands in Phyllomedusa hypochondrialis (Anura, Hylidae): secretory patterns during ontogenesis.

Three syncytial gland types (Ia, Ib, and II ) have been described in the skin of larval, juvenile and adult Phyllomedusa hypochondrialis, which share the ultrastructural traits common to the serous secretory units of anuran skin, although each manufactures a peculiar product. Type la secretion consists of dense granules provided with a peculiar substructure, type Ib of vesicles holding a lucent material, type II of lipid deposits. None of the developmental stages investigated showed intermediate features between any of the three cutaneous products, which accumulate in the syncytial cytoplasms of the secretory units following different biosynthetic pathways, consistent with each gland type. These findings confirm previous results on adult specimens of P. hypochondrialis and P. sauvagei and stress the polymorphism of the serous glands in the genus Phyllomedusa. This morphological variability reflects the wide adaptive flexibility of serous glands in anurans.

Spermathecae of Salamandrina terdigitata (Amphibia: Salamandridae): Patterns of sperm storage and degradation

The spermathecae of female Salamandrina terdigitata were observed using light and transmission electron microscopy during the fallspring period of sperm storage and secretory activity and during the summer stasis. When sperm are stored inside the spermathecae, the product synthesized by spermathecal epithelial cells is exported into the lumen, where it bathes the sperm. During sperm storage some spermatozoa undergo degradation by the spermathecal epithelium. This process, which includes sperm capture by the apical microvilli, formation of endocytic vacuoles and production of lysosomes, becomes prominent shortly after oviposition. In many instances, cells filled with vacuolized spermatozoa and/or residual bodies undergo desquamation from the spermathecal epithelium and enter the lumen together with residual sperm. Desquamated cells, together with residual sperm, are a common feature in the spermathecal lumina at the end of the egg‐laying season.

Serous cutaneous glands of Argentine Phyllomedusa Wagler 1830 (Anura Hylidae): secretory polymorphism and adaptive plasticity

An ultrastructural study has been carried out on the cutaneous serous glands in the Argentine hylids Phyllomedusa sauvagei Boulenger 1882 and P. hypochondrialis Boulenger 1882. Both species possess type I serous units, which are representative of the usual serous glands in anurans, and type II units, peculiar to the genus Phyllomedusa Wagler 1830. Type I secretory units were further characterized as type Ia and Ib, on account of the morphology of their specific products. Type Ia secretory materials consist of spheroidal granules (4–6 μm in diameter), which are compact in P. sauvagei and provided with a mesh-like substructure in P. hypochondrialis. Type Ib serous deposits in P. sauvagei are also granules, which exhibit variable density, irregular shape, and are usually larger (up to 15 μm in diameter), whereas in P. hypochondrialis they are even wider vesicles (up to 20 μm), holding a finely dispersed material. No intermediate forms were detected between these features, and therefore the type I secretory u...

Serous gland dimorphism in the skin of Melanophryniscus stelzneri (Anura: Bufonidae).

Two serous gland types (I and II) in the skin of the Argentine toad Melanophryniscus stelzneri were discovered using light and electron microscopy. Glands of the two types differ in several traits: features of the products (both mature and immature), organelles involved in biosynthesis, and paths of serous maturation. No consistent differences, however, were detected between the myoepithelial sheaths encircling the secretory units. Type I glands manufacture vesicles containing a single dense body with a repeating substructure and conform to the fundamental secretory line of bufonid skin, a secretory line involved in biosynthesis of steroids. Type II glands synthesize granules of varying densities and seem to belong to a line of glands that secrete proteinaceous products. The occurrence of the two serous gland types in Melanophryniscus stelzneri is discussed in a comparison with current literature on the morphofunctional characteristics of anuran poison glands, which perform both regulative and defensive roles. It is suggested that di‐ or polymorphism in serous glands is an adaptive trait that allows differential release of active molecules on the body surface. J. Morphol. 237:19–32, 1998.

Cadmium modulates proliferation and differentiation of human neuroblasts

Cadmium is an environmental pollutant inducing numerous pathological effects, including neurological disorders and brain diseases. However, little is known about the molecular mechanisms of cadmium in affecting neurons and in inducing neurotoxicity in the development of the human brain. We have recently established, cloned, and propagated in vitro a primary long‐term cell culture (FNC‐B4) obtained from the human fetal olfactory neuroepithelium. In the present study, we show that different concentrations of cadmium chloride (CdCl2) induced dose‐dependent biological effects in FNC‐B4 cells. A low concentration (10 μM) of CdCl2 stimulated neuroblast growth, whereas a high concentration (100 μM) inhibited the growth and the viability of neuroblasts inducing morphological and cytoskeletal alterations as well as apoptotic cell death. We also observed that CdCl2 affected, in a dose‐dependent manner, the differentiation of FNC‐B4 neuroblasts, with increased mRNA and protein levels of differentiation markers and decreased expression levels of neuronal stem markers. Furthermore, differentiated cells co‐expressed glial and neuronal markers. We suggest that CdCl2 in FNC‐B4 neuroblasts might represent a selective cue by which, in a heterogeneous primary culture, the more differentiated mature cells die, whereas the undifferentiated cells, at the same time glial and neuronal progenitors, are forced to access a state of differentiation.

Secretory granule-cytoplasm relationships in serous glands of anurans: ultrastructural evidence and possible functional role.

A survey covering the serous (granular) cutaneous glands in several anuran families from the Old and New Worlds (Bombinatoridae, Discoglossidae, Ranidae, Hylidae, Pseudidae and Leptodactylidae) has revealed consistent patterns of complex interactions between the syncytial secretory unit and serous deposits (granules). These relationships involve outgrowths from the syncytial cytoplasm encircling the granules and complex invaginations of the perigranular compartment (halo) into the syncytium. The outgrowths are branched, cytoplasm processes resembling ramified microvilli, or can be larger, dome-like to cylindrical structures. Despite their different features and origins, all these structures are efficient devices for amplifying the cytoplasmic surfaces round the granules, so improving exchange between the secretory syncytium and serous product. These complex secretory granule-cytoplasm interactions affect the product released from the Golgi apparatus and are consistent with the hypothesis of prolonged serous maturation following the initial phase of biosynthesis. Post-Golgian maturation modifies the secretory material on a centripetal gradient, causing condensation and, possibly, the transfer of component molecules from and/or to the cytoplasm.

Secretory polymorphism and serous cutaneous gland heterogeneity in Bufo granulosus (Amphibia, Anura).

Three types of secretory products (a, b and c) in the poison glands of the Argentine toad Bufo granulosus have been detected under light microscope. The type a secretory product consists of granules of homogeneous density, type b of vesicles with a translucent compartment and type c of granules of varying density. Subsequent transmission electron microscope analysis disclosed obvious similarities in the secretory pathways of type a and c granules; the differences detected under light microscope are due to the functional phases observed. On the contrary, production of type b secretory vesicles involves a distinctive pathway. Therefore, two classes of glands (I and II) have been identified. Glands of the first class are typical of bufonid toads and produce granules provided with repeating substructure; glands of the second class, which manufacture a lucent product, are unusual in the family Bufonidae. Ultrastructural differences, consistent with the two gland classes, have also been described in the myoepithelia. The myocytes ensheathing class I secretory units possess striking cytoskeletal specializations, whereas those of class II glands are rich in sarcoplasmic reticulum. The distinctive ultrastructural traits detected in these myoepithelial cells have been compared with the results of previous studies on the dimorphic serous glands of Bombina. Findings point to the use of pharmacological treatment on the skin of anurans with different classes of serous glands to elicit differential secretory discharge.

Ultrastructural patterns of secretory activity in poison cutaneous glands of larval and juvenile Dendrobates auratus (Amphibia, Anura)

A transmission electron-microscope study has been performed on larval and juvenile skin of the Central American arrow-frog Dendrobates auratus to investigate early secretory processes and maturational changes in the serous (poison) glands. Poison biosynthesis involves the endoplasmic reticulum (both smooth and rough types), as well as Golgi stacks which release early serous product as secretory vesicles (or pre-granules). These vesicles contain fine-grained material, along with single electron-opaque bodies, spheroidal in shape, that accompany the grained product throughout its post-Gogian, maturational change. The first steps of this process involve condensation and lead to the formation of secretory granules with a glomerular-like substructure, resulting from a thick, random aggregation of rods (secretory granule subunits). Advanced maturational activity causes the loss of peculiar granule substructure: the dense bodies split into fragments, whereas the thick glomerular arrangement becomes looser, until the secretory product changes into a dispersed material. This ultrastructural study revealed biosynthesis and maturation processes in close sequence, suggesting the poison of D. auratus contains proteins and/or peptides as well as lipophilic compounds. Molecules of both these classes are known to perform several roles relevant to survival strategies in extant anurans. Furthermore, the ephemeral granules with a glomerular-like substructure detected in tadpoles and froglets exhibit the complex patterns of mature poisons in adult specimens of other anurans: Hylidae and related families. This agrees with current trends in the taxonomy of these advanced frogs and underlines the pertinence of an ontogenetic approach in investigating anuran phylogenesis.

Development of serous cutaneous glands in Scinax nasica (Anura, Hylidae): patterns of poison biosynthesis and maturation in comparison with larval glands in specimens of other families.

We examined the development of serous (poison) cutaneous glands in larval and juvenile Scinax nasica (Hylidae) at the ultrastructural level. We describe the biosynthesis and maturation of the cutaneous poison in comparison with the corresponding processes in representatives of Discoglossidae, Leptodactylidae, Pelobatidae and Pipidae. Serous biosynthesis in S. nasica starts in discrete adenoblasts and continues in the syncytial secretory unit. Biosynthetic processes involve rough endoplasmic reticulum and the Golgi apparatus, that releases membrane-bounded material, varying from fine grained to flocculent. During the post-Golgian secretory phase, this material undergoes initial maturation, and two products are formed: dense granules and larger vesicles holding a thin substance that will later be structured into a three-dimensional, honeycomb-like net. Both the secretory granules and vesicles change into glomerular-like aggregates of bowed, rod-shaped subunits (modules). In adult frogs, formation of dense granules is bypassed. The modular granule substructure seems to be related to the merocrine release of small amounts of poison, involved in regulating skin homeostasis. Comparison with maturational changes in larval glands of species representing four anuran families discloses similar patterns in the Leptodactylidae, but production of opaque homogeneous granules occurs in the Discoglossidae, clear vesicles in the Pelobatidae and aggregates of dense bars in the Pipidae.

A BACTERICIDAL PROTEIN IN BOMBINA VARIEGATA PACHYPUS SKIN VENOM

The skin venom of the yellow bellied toad Bombina variegata pachypus has an antimicrobial activity which seems to be correlated to the presence of a 6700 mol. wt polypeptide. This polypeptide was purified by electroelution from SDS-urea-polyacrylamide gels and characterized for its antimicrobial activity. A bactericidal action was detected at concentrations with little or no cytolytic effect. The determination of the Minimal Inhibitory Concentration showed that there was activity against gram positive and gram negative bacteria and also against yeasts. The skin secretions of three other anuran species (Bufo viridis, Hyla arborea and Discoglossus pictus) were examined for the presence of antimicrobial activities. Only the Hyla arborea secretion exhibited antimicrobial properties. A small amount of a 6700 mol. wt polypeptide was detected among the Hyla secreted products.