Habituation in anuran tadpoles and the role of risk uncertainty.

The ability to learn in the context of predation allows prey to respond to threats by adjusting their behavior based on specific information acquired from their current environment. Habituation is a process that allows animals to adapt to environmental changes. Very little is known about habituation in wild animals in general and there are no studies on habituation in anuran tadpoles in particular. Here, we performed three experiments to investigate the behavioral response of predator naïve Pleurodema thaul tadpoles to repeated stimulation with two predation risk cues (injured conspecific and predator fed cues) which a priori provide different information regarding risk. Experiment 1 showed that P. thaul tadpoles habituate the antipredator response when undergo predation risk chemical cues from injured conspecific and that response is long term. Experiment 2 showed that P. thaul tadpoles did not habituate their antipredator response when exposed to cues derived from an event of nymph odonate preying on P. thaul tadpoles (predator fed cues). Experiment 3 specifically evaluated the risk imposed by each of the risk cues used in Experiment 1 and Experiment 2 and showed that the degree of perceived risk in tadpoles appear to be similar in a single experience with any risk stimuli. We suggest that the behavioral habituation of tadpoles in the context of predation could be modulated by the level of uncertainty associated with risk stimuli.

Thaulin-1: The first antimicrobial peptide isolated from the skin of a Patagonian frog Pleurodema thaul (Anura: Leptodactylidae: Leiuperinae) with activity against Escherichia coli.

Patagonia's biodiversity has been explored from many points of view, however, skin secretions of native amphibians have not been evaluated for antimicrobial peptide research until now. In this sense, Pleurodema thaul is the first amphibian specie to be studied from this large region of South America. Analysis of cDNA-encoding peptide in skin samples allowed identification of four new antimicrobial peptides. The predicted mature peptides were synthesized and all of them showed weak or null antimicrobial activity against Klebsiella pneumoniae, Staphylococcus aureus and Escherichia coli with the exception of thaulin-1, a cationic 26-residue linear, amphipathic, Gly- and Leu-rich peptide with moderate antimicrobial activity against E. coli (MIC of 24.7µM). AFM and SPR studies suggested a preferential interaction between these peptides and bacterial membranes. Cytotoxicity assays showed that thaulin peptides had minimal effects at MIC concentrations towards human and animal cells. These are the first peptides described for amphibians of the Pleurodema genus. These findings highlight the potential of the Patagonian region's unexplored biodiversity as a source for new molecule discovery.

Anuran tadpoles learn to recognize injury cues from members of the same prey guild.

Recognition of predation risk from cues released from injured heterospecific could be beneficial when prey belongs to the same prey guild. Here, we performed three experiments. Experiment 1 showed that P. thaul tadpoles reduced their activity levels when exposed to conspecific injury cues, but not when exposed to amphipod injury cues. Experiment 2 tested whether P. thaul tadpoles can learn to recognize predation risk from chemical cues released from injured heterospecifics from the same prey guild (amphipod, Hyalella patagonica). A group of tadpoles were conditioned by exposing them to a specific concentration of amphipod injury cues paired with conspecific injury cues. Two days later, we evaluated changes in the activity of tadpoles when they were exposed to amphipod cues. As a control of learning, we used an unpaired group. Additionally, we used more control groups to fully investigate the learning mechanism. Our results showed that tadpoles can learn to recognize predation risk from injured amphipods and that the mechanism underlying the observed learned response could be associative. Experiment 3 replicated Experiment 2 and also showed that a low concentration of amphipod cues did not sustain that learning.

Lack of functional link in the tadpole morphology induced by predators.

Most studies of predator-induced plasticity have focused on documenting how prey species respond to predators by modifying phenotypic traits and how traits correlate with fitness. We have previously shown that Pleurodema thaul tadpoles exposed to the dragonfly Rhionaeschna variegata responded strongly by showing morphological changes, less activity, and better survival than non-exposed tadpoles. Here, we tested whether there is a functional link between morphological plasticity and increased survival in the presence of predators. Tadpoles that experienced predation risk were smaller, less developed, and much less active than tadpoles without this experience. Burst speed did not correlate significantly with morphological changes and predator-induced deeper tails did not act as a lure to divert predator strikes away from the head. Although we have previously found that tadpoles with predator-induced morphology survive better under a direct predator threat, our results on the functional link between morphology and fitness are not conclusive. Our results suggest that in P. thaul tadpoles (1) burst speed is not important to evade predators, (2) those exposed to predators reduce their activity, and (3) morphological changes do not divert predator attacks away from areas that compromise tadpole survivalEE. Our results show that morphological changes in P. thaul tadpoles do not explain burst speed or lure attraction, although there was a clear reduction of activity, which itself reduces predation. We propose that changes in tadpole activity could be further analyzed from another perspective, with morphological change as an indirect product of behavior mediated by physiological mechanisms.

Chronic stress in Lizards: Studies on the Behavior and Benzodiazepine Receptors in Liolaemus koslowskyi and Cnemidophorus tergolaevigatus.

Behavioral and physiological adaptive responses of animals facing chronic exposure to a single stressor may allow them to overcome its negative effects for future exposures to similar stressful situations. At chemical level, the GABAA /benzodiazepine complex is considered one of the main receptor systems involved in the modulation of stress-induced responses. Here, we describe the behavioral responses of two different lizard species, Liolaemus koslowskyi and Cnemidophorus tergolaevigatus exposed to three potential chronic stressful treatments: (a) high temperature, (b) forced swimming, and (c) simulated predator. Additionally, we aimed to determine in those lizards whether the central-type benzodiazepine receptor (CBR; an allosteric modulator site of the GABAA receptor) is related to adaptive responses to those stressful stimulations. Our results revealed that the simulated predator was the stress condition that showed the largest difference in behavioral responses between the two species, resembling previously described strategies in nature. The basal affinity of CBRs (obtained from undisturbed animals) showed differences between both species, and the simulated predator was the only stressor that altered the affinity of CBRs. L. koslowskyi CBRs showed a decreased receptor affinity, whereas C. tergolaevigatus showed an increased receptor affinity in comparison to their respective control groups. We show for the first time the effects of different types of stressors upon behavioral responses and CBR biochemical parameters in two lizard species. Our findings suggest a potential GABA/benzodiazepine role in the ability of lizards to cope with a repeated exposure to a stressful (e.g., predator) condition.

Prenatal and postnatal ethanol experiences modulate consumption of the drug in rat pups, without impairment in the granular cell layer of the main olfactory bulb.

The effect of moderate exposure to ethanol during late gestation was studied in terms of its interaction with moderate exposure during nursing from an intoxicated dam. A further issue was whether behavioral effects of ethanol, especially the enhanced ethanol intake known to occur after moderate ethanol prenatally or during nursing, depend upon teratological effects that may include death of neurons in the main olfactory bulb (MOB). During gestational days 17-20 rats were given 0, 1 or 2g/kg ethanol doses intragastrically (i.g.). After parturition these dams were given a dose of 2.5g/kg ethanol i.g. each day and allowed to perform regular nursing activities. During postnatal days (PDs) 15 and 16, ethanol intake of pups was assessed along with aspects of their general activity. In a second experiment pups given the same prenatal treatment as above were tested for blood ethanol concentration (BEC) in response to an ethanol challenge on PD6. A third experiment (Experiment 2b) assessed stereologically the number of cells in the granular cell layer of the MOB on PD7, as a function of analogous pre- and postnatal ethanol exposures. Results revealed that ethanol intake during the third postnatal week was increased by prenatal as well as postnatal ethanol exposure, with a few interesting qualifications. For instance, pups given 1g/kg prenatally did not have increased ethanol intake unless they also had experienced ethanol during nursing. There were no effects of ethanol on either BECs or conventional teratology (cell number). This increases the viability of an explanation of the effects of prenatal and early postnatal ethanol on later ethanol intake in terms of learning and memory.

Ethanol exposure during late gestation and nursing in the rat: effects upon maternal care, ethanol metabolism and infantile milk intake.

Ethanol experiences, during late gestation as well as during nursing, modify the behavioral dynamics of the dam/pup dyad, and leads to heightened ethanol intake in the offspring. This study focuses on: a) behavioral and metabolic changes in intoxicated dams with previous exposure to ethanol during pregnancy and b) infantile consumption of milk when the dam is either under the effects of ethanol or sober. Pregnant rats received water, 1.0 or 2.0 g/kg ethanol, and were administered with water or ethanol during the postpartum period. Intoxication during nursing disrupted the capability of the dam to retrieve the pups and to adopt a crouching posture. These disruptions were attenuated when dams had exposure to ethanol during pregnancy. Ethanol experiences during gestation did not affect pharmacokinetic processes during nursing, whereas progressive postpartum ethanol experience resulted in metabolic tolerance. Pups suckling from intoxicated dams, with previous ethanol experiences, ingested more milk than did infants suckling from ethanol-intoxicated dams without such experience. Ethanol gestational experience results in subsequent resistance to the drug's disruptions in maternal care. Consequently, better maternal care by an intoxicated dam with ethanol experience during gestation facilitates access of pups to milk which could be contaminated with ethanol.

Fetal learning about ethanol and later ethanol responsiveness: evidence against "safe" amounts of prenatal exposure.

Near-term fetuses of different mammalian species, including humans, exhibit functional sensory and learning capabilities. The neurobiological literature indicates that the unborn organism processes sensory stimuli present in the amniotic fluid, retains this information for considerable amounts of time, and is also capable of associating such stimuli with biologically relevant events. This research has stimulated studies aimed at the analysis of fetal and neonatal learning about ethanol, a topic that constitutes the core of the present review. Ethanol has characteristic sensory (olfactory, taste, and trigeminal) attributes and can exert pharmacologic reinforcing effects. The studies under examination support the hypothesis that low to moderate levels of maternal ethanol intoxication during late pregnancy set the opportunity for fetal learning about ethanol. These levels of prenatal ethanol exposure do not generate evident morphologic or neurobehavioral alterations in the offspring, but they exert a significant impact upon later ethanol-seeking and intake behaviors. Supported by preclinical and clinical findings, this review contributes to strengthening the case for the ability of prenatal ethanol exposure to have effects on the postnatal organism.