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Comparative and Functional Analysis of the Digital Mucus Glands and Secretions of Tree Frogs


Comparative and Functional Analysis of the Digital Mucus Glands and Secretions of Tree Frogs

Julian K A Langowski et al. Front Zool.


Background: Mucus and mucus glands are important features of the amphibian cutis. In tree frogs, the mucus glands and their secretions are crucial components of the adhesive digital pads of these animals. Despite a variety of hypothesised functions of these components in tree frog attachment, the functional morphology of the digital mucus glands and the chemistry of the digital mucus are barely known. Here, we use an interdisciplinary comparative approach to analyse these components, and discuss their roles in tree frog attachment.

Results: Using synchrotron micro-computer-tomography, we discovered in the arboreal frog Hyla cinerea that the ventral digital mucus glands differ in their morphology from regular anuran mucus glands and form a subdermal gland cluster. We show the presence of this gland cluster also in several other-not exclusively arboreal-anuran families. Using cryo-histochemistry as well as infrared and sum frequency generation spectroscopy on the mucus of two arboreal (H. cinerea and Osteopilus septentrionalis) and of two terrestrial, non-climbing frog species (Pyxicephalus adspersus and Ceratophrys cranwelli), we find neutral and acidic polysaccharides, and indications for proteinaceous and lipid-like mucus components. The mucus chemistry varies only little between dorsal and ventral digital mucus in H. cinerea, ventral digital and abdominal mucus in H. cinerea and O. septentrionalis, and between the ventral abdominal mucus of all four studied species.

Conclusions: The presence of a digital mucus gland cluster in various anuran families, as well as the absence of differences in the mucus chemistry between arboreal and non-arboreal frog species indicate an adaptation towards generic functional requirements as well as to attachment-related requirements. Overall, this study contributes to the understanding of the role of glands and their secretions in tree frog attachment and in bioadhesion in general, as well as the evolution of anurans.

Keywords: Cryo-histochemistry; Hyla cinerea; Infrared spectroscopy; Lubrication; Macrogland; Mucosubstance; Sum frequency generation spectroscopy; Synchrotron micro-computer-tomography; Wet adhesion.

Conflict of interest statement

Competing interestsThe authors declare that they have no competing interests.


Fig. 1
Fig. 1
Mucus glands in a digital pad of Hyla cinerea. a 3D view of the whole digital tip. Only the left half of the approximately bi-laterally symmetric dorsal mucus glands is shown. b Frontal, c lateral, and d ventral view of the glands. The dotted lines indicate the proximal border of gland segmentation. x, y, z: longitudinal, lateral, and vertical spatial coordinates
Fig. 2
Fig. 2
Spatial distribution of the digital mucus gland pores in Hyla cinerea. a 3D view of the dorsal (crosses; only the left half of the dorsal mucus pores is shown) and ventral (circles) pores on the digital pad cutis. b Spatial variation of the dorsal and c ventral mucus pore density, approximated by the nearest-neighbour-distance NND [colour-coded in µm], across the pad surface in dorsal view
Fig. 3
Fig. 3
Brightfield micrographs of cryo-histochemically stained mucus glands in the digital pads of Hyla cinerea. Per staining, an overview of a whole representative transverse section through the gland space is shown. Insets: magnified views of a dorsal (top) and a ventral (bottom) mucus gland; black arrowheads: positive staining, white arrowheads: equivocal staining, Scale bar = 25 µm; CRO Crossmons’s light green trichrome including Mayer’s haematoxylin and Alcian blue, OIO Oil Red O, PAS Periodic acid-Schiff, PAS-D Periodic acid-Schiff-Diastase, AB Alcian blue (pH =1, pH =2.5), NIN Ninhydrin-Schiff, COO Coomassie blue, BRO Mercuric bromophenol blue (pH =3.8, pH =4.6). CH Chromatophore, CO Collagen, DE Dermis, DP Digital phalanx, DU Mucus gland duct, ED Epidermis, MG mucus glands. OIO- and NIN-stained sections were imaged using differential interference contrast
Fig. 4
Fig. 4
ATR-IR spectra of frog mucus. a Comparison of ventral digital (solid line) and abdominal (dotted line; absorbance scaled by ×3) mucus in Hyla cinerea. Vibrational peaks attributed to amide, N-H stretch, aromatic C-H stretch, and aliphatic C-H stretches were obtained. b Comparison of abdominal mucus spectra of the arboreal frog species H. cinerea (black) and Osteopilus septentrionalis (red; absorbance scaled by ×0.1) as well as the terrestrial species Pyxicephalus adspersus (blue; absorbance scaled by ×0.5). The spectra are offset along the ordinate axis in steps of 0.002 for clarity
Fig. 5
Fig. 5
SFG spectra of frog mucus. a Comparison of ventral digital (crosses) and abdominal (circles) mucus in Hyla cinerea in (a) PPP and (c) SSP polarisations. CH3 vibrations include methyl symmetric, methyl Fermi, and methyl asymmetric vibrations. b Comparison of the ventral abdominal mucuses of the arboreal frog species H. cinerea (black) and Osteopilus septentrionalis (red) as well as the terrestrial species Pyxicephalus adspersus (blue) and Ceratophrys cranwelli (magenta) in (b) PPP and (d) SSP polarisations. The solid lines represent the fitting curves using the Lorentzian equation (Eq. 2). The spectra are offset along the ordinate axis in steps of 100 (PPP) and 50 (SSP) for clarity

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