RUI: Aquaporins and Osmoregulation in a Freeze-Tolerant Amphibian

尽管人们梦想着能够冷冻整个身体，然后让它们复活，但目前成功的生物医学冷冻保存仅限于血液或胚胎等相对较小的结构。然而，有些动物，包括一些两栖动物和爬行动物，能够在北方的冬天存活下来，部分原因是它们能够忍受冰冻。这项研究将研究其中一种物种——柯普灰树蛙——耐寒性的生理机制。这种青蛙在抗冻脊椎动物中是不寻常的，因为它在体液中积累甘油作为冷冻保护剂；甘油也常用于生物医学防腐溶液中。需要解决的具体问题是，当树蛙适应寒冷和实际冻结时，甘油是如何进出树蛙的组织的，以及甘油的运动如何与组织中水的分布有关，这对于防止细胞内结冰至关重要，否则会导致致命的伤害。特别是，实验将研究水通道蛋白在完成甘油和水的运动中的作用。水通道蛋白是一种已知的蛋白质，可以形成通道，允许水，有时也允许甘油等小分子穿过细胞膜。据推测，它们对抗冻能力至关重要。目的是加强对这一迷人的自然现象的理解，并使用耐寒性的自然模型来阐明可应用于生物医学冷冻保存的原理。在这个过程中，这项工作将为本科生提供从比较生物学原理到整个动物和分子生物学的多层次分析培训。这项训练将鼓励学生在动物生理学研究中运用综合思维和多种方法。

Although people dream of being able to freeze whole bodies and later return them to life, currently successful biomedical cryopreservation is restricted to relatively small structures like blood or embryos. However, there are animals, including several amphibians and reptiles, that survive northern winters in part by an ability to tolerate being frozen. This research will examine the physiological mechanisms responsible for freeze tolerance in one of those species, Cope's gray treefrog. This frog is unusual among freeze-tolerant vertebrates in that it accumulates glycerol in its body fluids as a cryoprotective agent; glycerol is also commonly used in biomedical preservative solutions. Specific questions to be addressed are how glycerol moves in and out of tissues of the treefrogs as they acclimate to cold and actually freeze, and how the glycerol movement is related to the distribution of water among tissues, which is critical to preventing ice formation inside cells where it would lead to lethal damage. In particular, experiments will investigate the role of aquaporins in accomplishing that glycerol and water movement. Aquaporins are proteins known to form passageways that allow water and, sometimes, other small molecules like glycerol to cross cell membranes, and it is hypothesize that they are critical to freeze tolerance. The objectives are to enhance the understanding of this fascinating natural phenomenon, and to use a natural model of freeze tolerance to elucidate principles that can be applied to biomedical cryopreservation. In the process, this work will provide training to undergraduate students at multiple levels of analysis, from principles of comparative biology to whole animals and molecular biology. This training will encourage integrative thinking and application of diverse methods to the study of animal physiology.