Biologia 63/3: 444—449, 2008 Section Cellular and Molecular Biology DOI: 10.2478/s11756-008-0071-6 Glycoconjugate histochemistry of mucous glands in the skin of metamorphosing Bufo viridis Mesut Sahin, Ay¸ ca Gumus & Erdal Balcan* Molecular Biology Section, Department of Biology, Celal Bayar University, Faculty of Science and Art, 45047 Muradiye Campus, Manisa, Turkey; e-mail: erdal.balcan@gmail.com Abstract: Mucins are the major glycoprotein secretions of mucous glands and display important functions in amphibian skin such as regulation of water homeostasis and mechanical and chemical protection. In the present study, we evaluated the glycoconjugate contents of developing mucous glands on dorsal regions of metamorphosing Bufo viridis (Amphibia: Anura) tadpoles using an alcian blue-PAS panel and lectin histochemistry. All the conical cells of mucous glands showed weak positivity for alcian blue in 0.025 M MgCl2 at pH 5.7 but only a few cells were positive for 0.3 M MgCl2 at the same pH. In addition, all the conical cells of mucous glands were negative for alcian blue at pH 2.5. In lectin histochemistry, conical cells reacted strongly with Galanthus nivalis agglutinin (GNA), Datura stramonium agglutinin (DSA) and peanut agglutinin (PNA), weakly with Maackia amurensis leucoagglutinin (MAL). These results suggest that they express predom- inantly mannose, galactose and partially α(2→3)-linked sialic acid containing glycoconjugates. We concluded that dorsal mucous glands of metamorphosing Bufo viridis tadpoles contain at least two different conical cell types and glycoconjugate heterogeneity of mucous glands may be related with different functions of mucins. Key words: Bufo viridis; mucous gland; glycoconjugate; lectin histochemistry. Abbreviations: AB, alcian blue; DIG, digoxigenin; DSA, Datura stramonium agglutinin; GalNAc, N-acetylgalactosamine; GlcNAc, N-acetylglucosamine; GNA, Galanthus nivalis agglutinin; MAL, Maackia amurensis leucoagglutinin; PAS, periodic acid-Schiff’s; PNA, peanut agglutinin; TBS, Tris-buffered saline; SNA, Sambucus nigra agglutinin. Introduction Anuran skin is a very complex organ that plays im- portant roles in mechanical protection (Amiche et al. 2000), respiration (Van Driessche & Hillyard 1985), and ion and water balance (Stiffler 1994; Sørensen & Larsen 1996; Jørgensen 1993; 1997). It is comprised of two major parts: a mucus-covered epidermis and un- derlying dermis. During metamorphosis, distinct struc- tural changes occur in the skin paralleled by changes in its functional properties (Takada 1985; Fox 1986), for example, mucous glands develop and dorsal skin of body transforms to adult skin, whereas tail skin disap- pears by tail resorption at metamorphic climax stage (Kaltenbach et al. 2004; Kinoshita et al. 2005). Cuta- neous mucus produced by intradermal mucous glands protects amphibians from excessive water loss or gain, acts as a barrier that protects from bacterial and fun- gal infections and minimizes mechanical damage to the skin out of water (Clarke 1997). Mucins are high molecular weight glycoproteins that contain at least one protein domain (Thornton & Sheehan 2004). Cell surface-bound and -secreted mucins contain a range of N-acetylgalactosamine (GalNAc)-Ser/Thr O-linked carbohydrate residues (O-glycans) (Brockhausen 2006). In addition to GalNAc, mucins may also contain N- acetylglucosamine (GlcNAc), galactose, fucose, sialic acids and sulphate (Thornton & Sheehan 2004). Carbo- hydrates of the cell surface and/or extracellular matrix glycoconjugates and proteoglycans have many diverse and important functions in vertebrates, like cell-cell and cell-matrix adhesion events, cell signalling and develop- ment (Varki 1993; Sharon & Lis 2001). Lectins are non-immune proteins or glycoproteins that bind specific terminal or sub-terminal carbohy- drate residues (Vijayan & Chandra 1999). They are widely used as probes to detect different type glyco- conjugates in cells (Lis & Sharon 1986). In the present study we evaluated the glycoconju- gate alterations in mucous glands in dorsal regions of the skin during metamorphic climax of the Eurasian green toad Bufo viridis (Amphibia: Anura) tadpoles by histochemical and lectin histochemical analysis. Material and methods Animals Fertilized eggs of Eurasian green toad Bufo viridis (Am- phibia: Anura) were collected from natural puddles near the Muradiye Campus area of Celal Bayar University (Manisa, * Corresponding author c 2008 Institute of Molecular Biology, Slovak Academy of Sciences