Publication Date

Summer 2021

Advisor(s) - Committee Chair

Jarrett Johnson (Director), Kevin Bilyk, Clara do Amaral, and Noah Ashley

Degree Program

Department of Biology

Degree Type

Master of Science

Abstract

Freeze-tolerant gray treefrogs survive winter by producing natural cryoprotectants and accommodating ice formation within extracellular spaces. While frozen, gray treefrogs endure hyperglycemia, dehydration, and anoxia due to the halt of all bodily functions. Upon thawing, the frogs’ anoxic cells receive a rapid influx of oxygen, which can cause oxidative damage to vital macromolecules including DNA. Previous studies have suggested freeze-tolerant frogs avoid oxidative damage after freeze-thaw events by elevating antioxidant activity, but recent work has shown upregulated DNA repair encoding genes in post-freeze frogs. The objective of this thesis is to assess the cellular costs of freezing by measuring oxidative DNA damage in gray treefrogs (Hyla versicolor) as they thaw. Experimental frogs were frozen for 24 hours at -3°C and dissected after 2, 6, or 24 hours of thawing. Both liver and skeletal muscle tissues were excised and examined due to previously reported differences in antioxidant and cryoprotectant capacity. An ELISA was used to detect concentrations of oxidatively damaged guanine in each tissue sample. It was found that oxidative DNA damage within liver tissue did not increase from baseline during the freeze-thaw event. In muscle tissue, damaged DNA concentrations were elevated after 2 hours of thawing but slowly decreased with time. By 24 hours post-freeze, damage levels in muscle tissue returned to baseline, suggesting a full recovery. The absence of freezing-induced damage in the liver and short-lasting damage in the muscle shows that gray treefrogs are well adapted to mitigate freeze-thaw injury. These results reaffirm that freeze-tolerance is an effective overwintering strategy that is comprised of a complex series of evolutionary adaptations, which deserve further investigation.

Disciplines

Biology | Cellular and Molecular Physiology | Zoology

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