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Alarm cue properties and euthanasia impact on tadpole antipredator behaviour in Rana temporaria and Bufo bufo

Konrad Lipkowski,D. Abondano Almeida,D. Wenzel,L. M. Schulte

Published 2025 in Animal Behaviour

ABSTRACT

Chemical cues are crucial for aquatic organisms to assess predation risks, impacting their antipredator mechanisms. Tadpoles, recognized as valuable model organisms, use conspeci fi c alarm cues to modulate their behaviours. However, ethical considerations in chemical communication research often necessitate the use of chemical anaesthetics, potentially disrupting ecologically relevant processes and complicating result interpretation. Furthermore, our understanding of alarm cue characteristics across most anuran species and their potential interactions with chemical anaesthetics remains limited or unexplored. In this study, we examined euthanasia and extract processing techniques on tadpole behaviour in the common toad, Bufo bufo, and common frog, Rana temporaria, to achieve three objectives: investigate alarm cue properties, assess the impact of euthanasia methods on alarm cue effectiveness, and explore anaesthetic interference mechanisms using tricaine and L -arginine. Tadpoles were exposed to various chemical stimuli, including alarm cues extracted via mechanical, hypothermal and chemical euthanasia methods, as well as differently processed crude extracts (formerly frozen, boiled, aged or protein-digested). Additionally, tad-poles were exposed to varying concentrations of L -arginine and tricaine, both individually and in combination. Our results revealed consistent species-speci fi c responses, with R. temporaria tadpoles exhibiting reduced activity in response to alarm cues, while B. bufo tadpoles displayed no behavioural changes. Moreover, we observed signi fi cant differences among methods of euthanasia. Tadpoles of R. temporaria responded to processed extracts and L -arginine, indicating the presence of amino acids but not proteinaceous compounds in alarm cues. Finally, divergent species ’ responses to L -arginine and tricaine suggest interference within individuals rather than alarm cues. This study enhances our understandingof alarm cue characteristics in tadpoles and underscores the potential species-speci fi c impact of chemical anaesthetics on behavioural responses in aquatic organisms. It underscores the necessity for further research into interference mechanisms and alternative euthanasia methods while emphasizing the importance of comparative studies in comprehending chemical communication in aquatic ecosystems

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