Dose-dependent alkaloid sequestration and N-methylation of decahydroquinoline in poison frogs

Sequestration of chemical defenses from dietary sources is dependent on the availability of compounds in the environment and the mechanism of sequestration. Previous experiments have shown that sequestration efficiency varies among alkaloids in poison frogs, but little is known about the underlying mechanism. The aim of this study was to quantify the extent to which alkaloid sequestration and modification are dependent on alkaloid availability and/or sequestration mechanism. To do this, we administered different doses of histrionicotoxin (HTX) 235A and decahydroquinoline (DHQ) to captive‐bred Adelphobates galactonotus and measured alkaloid quantity in muscle, kidney, liver, and feces. HTX 235A and DHQ were detected in all organs, whereas only DHQ was present in trace amounts in feces. For both liver and skin, the quantity of alkaloid accumulated increased at higher doses for both alkaloids. Accumulation efficiency in the skin increased at higher doses for HTX 235A but remained constant for DHQ. In contrast, the efficiency of HTX 235A accumulation in the liver was inversely related to dose and a similar, albeit statistically nonsignificant, pattern was observed for DHQ. We identified and quantified the N‐methylation of DHQ in A. galactonotus, which represents a previously unknown example of alkaloid modification in poison frogs. Our study suggests that variation in alkaloid composition among individuals and species can result from differences in sequestration efficiency related to the type and amount of alkaloids available in the environment

Differences and Similarities among Parotoid Macrogland Secretions in South American Toads: A Preliminary Biochemical Delineation

Amphibians are known by cutaneous glands, spread over the skin, containing toxins (proteins, peptides, biogenic amines, steroidal bufadienolides, and alkaloids) used as chemical defense against predators and microbial infection. Toads are characterized by the presence of parotoid macroglands. The common toads have lately been divided into two genera: Bufo (Europe, Asia, and Africa) and Rhinella (South America). Basal Rhaebo genus is exclusively of Central America and Amazon region. Although Rhinella and Rhaebo are related, species may share differences due to the diversity of environments that they live in. In this work, we have performed a biochemical characterization of the components of the poison of eight Rhinella species and one Rhaebo by means of RP-HPLC with either UV or MS detection and by SDS-PAGE, in order to verify whether phylogenetic and biological differences, such as habitat, diet, and defensive strategies, between them may also be reflected in poison composition. Although some components were common among the secretions, we were able to identify exclusive molecules to some species. The fact that closely related animals living in different habitats secrete different molecules into the skin is an indication that biological features, and not only evolution, seem to directly influence the skin secretion composition

Morphological and physiological specialization for digging in amphisbaenians, an ancient lineage of fossorial vertebrates

Amphisbaenians are legless reptiles that differ significantly from other vertebrate lineages. Most species dig underground galleries of similar diameter to that of the animal. We studied the muscle physiology and morphological attributes of digging effort in the Brazilian amphisbaenid Leposternon microcephalum (Squamata; Amphisbaenia), which burrows by compressing soil against the upper wall of the tunnel by means of upward strokes of the head. The individuals tested

Parental investment by skin feeding in a caecilian amphibian

Although the initial growth and development of most multicellular animals depends on the provision of yolk, there are many varied contrivances by which animals provide additional or alternative investment in their offspring(1). Providing offspring with additional nutrition should be favoured by natural selection when the consequent increased fitness of the young offsets any corresponding reduction in fecundity(2). Alternative forms of nutrition may allow parents to delay and potentially redirect their investment. Here we report a remarkable form of parental care and mechanism of parent-offspring nutrient transfer in a caecilian amphibian. Boulengerula taitanus is a direct-developing, oviparous caecilian(3), the skin of which is transformed in brooding females to provide a rich supply of nutrients for the developing offspring. Young animals are equipped with a specialized dentition, which they use to peel and eat the outer layer of their mother's modified skin. This new form of parental care provides a plausible intermediate stage in the evolution of viviparity in caecilians. At independence, offspring of viviparous and of oviparous dermatotrophic caecilians are relatively large despite being provided with relatively little yolk. The specialized dentition of skin-feeding (dermatophagous) caecilians may constitute a pre-adaptation to the fetal feeding on the oviduct lining of viviparous caecilians.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62957/1/nature04403.pd

Pitviper infrared receptor organ.

<p>(A) Lateral view of <i>Bothriopsis taeniata</i> head. (B) High magnification showing infrared (IR) receptor organ pit opening and papillae (arrow) emerging from the anterior corner of the orbit in <i>Bothrops asper</i>. (C) Schematic representation of the IR receptor organ showing the location of the papillae (pa) in relation to pit membrane (pm) and domed epithelium (de) of the bottom of the inner chamber (after <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090622#pone.0090622-Goris1" target="_blank">[2]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090622#pone.0090622-Terashima1" target="_blank">[7]</a>). (D) View of the anterior orbital adnexa showing the IR receptor organ pore encircled by papillae in <i>Bothrops asper</i>. The arrow points to the more elongate papillae in the anterior corner of the orbit.</p

Osteogenesis Imperfecta: clinical, biochemical and molecular findings

Mutations in COL1A1 and COL1A2 genes, encoding the 1 and 2 chain of type I collagen, respectively, are responsible for the vast majority of cases of Osteogenesis Imperfecta (95% of patients with a definite clinical diagnosis). We have investigated twenty two OI patients, representing an heterogeneous phenotypic range, at the biochemical and molecular level. A causal mutation in either type I collagen gene was identified in twenty of them: no recurrent mutation was found in unrelated subjects; fifteen out of twenty mutations had not been reported previously. In two patients we could not find any causative mutation in either type I collagen gene, after extensive genomic DNA sequencing. Failure of COL1A1/COL1A2 mutation screening may be due, in a few cases, to further clinical heterogeneity, i.e. additional non collagenous disease loci are presumably involved in OI types beyond the traditional Sillence’s classification

Bufotenine is able to block rabies virus infection in BHK-21 cells

Abstract Background Rabies is a fatal zoonotic neglected disease that occurs in more than 150 countries, and kills more than 55.000 people every year. It is caused by an enveloped single stranded RNA virus that affects the central nervous system, through an infection initiated by the muscular nicotinic acetylcholine receptor, according to many authors. Alkaloids, such as acetylcholine, are widespread molecules in nature. They are present in numerous biological fluids, including the skin secretion of many amphibians, in which they act (together with proteins, peptides and steroids) as protection agents against predators and/or microorganisms. Among those amphibians that are rich in alkaloids, there is the genus Rhinella. Methods Bufotenine was isolated from Rhinela jimi skin secretion after a liquid-liquid partition (H2O:CH2Cl2) and reversed phase high-performance liquid chromatography analyses (RP-HPLC). Bufotenine was also extracted from seeds of Anadenanthera colubrina in acetone solution and purified by RP-HPLC, as well. Structural characterization was performed by mass spectrometry and nuclear magnetic resonance analyses. Cytotoxic tests of bufotenine were performed over baby hamster kidney (BHK-21) cells using MTT test. For the antiviral activity, Rabies virus strain Pasteur vaccine (PV) was used on fluorescence inhibition test and fluorescent foci inhibition test, with both simultaneous and time course treatment of the cells with the virus and bufotenine. Results In the present work we describe the effects of bufotenine, obtained either from toads or plants, that can inhibit the penetration of rabies virus in mammalian cells through an apparent competitive mechanism by the nicotinic acetylcholine receptor. Moreover, this inhibition was dose- and time-dependent, pointing out to a specific mechanism of action. Conclusions This work do not present or propose bufotenine as a drug for the treatment of rabies due to the hallucinogen and psychotropic effects of the molecule. However, continued studies in the elucidation of the antiviral mechanism of this molecule, may lead to the choice or development of a tryptamine analogue presenting potential clinical use

Consuming viscous prey: a novel protein-secreting delivery system in neotropical snail-eating snakes

Background: Efficient venom delivery systems are known to occur only in varanoid lizards and advanced colubroidean snakes among squamate reptiles. Although components of these venomous systems might have been present in a common ancestor, the two lineages independently evolved strikingly different venom gland systems. In snakes, venom is produced exclusively by serous glands in the upper jaw. Within the colubroidean radiation, lower jaw seromucous infralabial glands are known only in two distinct lineages-the basal pareatids and the more advanced Neotropical dipsadines known as goo-eating snakes. Goo-eaters are a highly diversified, ecologically specialized clade that feeds exclusively on invertebrates (e. g., gastropod molluscs and annelids). Their evolutionary success has been attributed to their peculiar feeding strategies, which remain surprisingly poorly understood. More specifically, it has long been thought that the more derived Dipsadini genera Dipsas and Sibynomorphus use glandular toxins secreted by their infralabial glands to extract snails from their shells.Results: Here, we report the presence in the tribe Dipsadini of a novel lower jaw protein-secreting delivery system effected by a gland that is not functionally related to adjacent teeth, but rather opens loosely on the oral epithelium near the tip of the mandible, suggesting that its secretion is not injected into the prey as a form of envenomation but rather helps control the mucus and assists in the ingestion of their highly viscous preys. A similar protein-secreting system is also present in the goo-eating genus Geophis and may share the same adaptive purpose as that hypothesized for Dipsadini. Our phylogenetic hypothesis suggests that the acquisition of a seromucous infralabial gland represents a uniquely derived trait of the goo-eating clade that evolved independently twice within the group as a functionally complex protein-secreting delivery system.Conclusions: The acquisition by snail-eating snakes of such a complex protein-secreting system suggests that the secretion from the hypertrophied infralabial glands of goo-eating snakes may have a fundamental role in mucus control and prey transport rather than envenomation of prey. Evolution of a functional secretory system that combines a solution for mucus control and transport of viscous preys is here thought to underlie the successful radiation of goo-eating snakes

Functional assessment of toad parotoid macroglands: A study based on poison replacement after mechanical compression

Toads have a pair of parotoid macroglands behind the eyes that secrete poison used in passive defence against predators. These macroglands are composed of juxtaposed alveoli, each one bearing a syncytial gland, all connected to the exterior by ducts. When the parotoids are bitten, the poison is expelled on the predator oral mucosa in the form of jets, causing several pharmacological actions. After poison release, the empty secretory syncytia immediately collapse in the interior of their respective alveoli and gradually start refilling. After parotoid manual compression, simulating a predator's bite, we studied, by means of morphological methods, the replacement of the poison inside the alveoli. The results showed that after compression, a considerable number of alveoli remained intact. In the alveoli that were effectively affected the recovery occurs in different levels, from total to punctual and often restrict to some areas of the syncytia. The severely affected alveoli seem not recover their original functional state. The fact that only a part of the parotoid alveoli is compressed during an attack seems to be crucial for toad survival, since the amphibian, after being bitten by a predator, do not lose all its poison stock, remaining protected in case of new attacks.The authors thank FAPESP (Process # 2008/57898-0) and CNPq-INCTTox (Process # 573790/2008-6) for the financial support. MMA (Process #312660/2013-8), CJ (Proc. #306529/2010-6) and MTR (Process #300541/2010-4) received grants from CNPq. SISBIO (Federal Brazilian Government) provided permission (# 15964-1) to MMA for collection of the animals