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Skin ice nucleators and glycerol in the freezing-tolerant frog Litoria ewingii

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Abstract

The brown tree frog (Litoria ewingii) is the only known Southern Hemisphere vertebrate that can survive full-body freezing. Freezing challenges living organisms in many ways, with ice formation in the body producing a suite of physical and metabolic stresses which can damage cells and tissues. The present study looked at two mechanisms that address some of these stresses: cryoprotectants and ice nucleating agents (INAs). Skin secretions from L. ewingii were sampled along with microhabitat substrate and tested for the presence of INAs, which help control ice formation in the body. L. ewingii plasma was tested for seasonal and freezing-induced changes in both glucose and glycerol, which may have a cryoprotective role in freezing-tolerant frogs. Glycerol levels increased on freezing and decreased on thawing, while glucose levels did not change on freezing but increased on thawing. This suggests that glycerol may be acting as a cryoprotectant, although levels are low compared to other frogs. A clear seasonal change was seen in INA activity, with greater activity in winter than in summer. While potent INAs came from the microhabitat substrate, this work has shown for the first time that skin secretions also contain active INAs.

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Abbreviations

INA:

Ice nucleating agent

k.c.p.:

Keyhole cardiac puncture

SVL:

Snout-vent length

T b :

Body temperature

T c :

Crystallization temperature

T circ :

Circulator temperature

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Acknowledgments

Our thanks to Ken Miller for help with image preparation and to the Phillips family for access to their land. This work was supported by a University of Otago Research Grant to D.A.W. and a departmental grant to K.R.H. We are also grateful to the two anonymous reviewers whose comments were of great help in improving this manuscript.

Ethical standards

Housing and experimental protocols and procedures for the care and use of Litoria ewingii complied with the guidelines of the Animal Welfare Act 1999, the National Animal Ethics Advisory Committee Good Practice Guide and the University of Otago Code of Ethical Conduct. All procedures were approved by the University of Otago Animal Ethics Committee.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Zoology, University of Otago, Dunedin, 9054, New Zealand

    Kalinka M. J. Rexer-Huber, Phillip J. Bishop & David A. Wharton

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  1. Kalinka M. J. Rexer-Huber
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  2. Phillip J. Bishop
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  3. David A. Wharton
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Correspondence to David A. Wharton.

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Communicated by I.D. Hume.

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Rexer-Huber, K.M.J., Bishop, P.J. & Wharton, D.A. Skin ice nucleators and glycerol in the freezing-tolerant frog Litoria ewingii . J Comp Physiol B 181, 781–792 (2011). https://doi.org/10.1007/s00360-011-0561-7

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