Organ transplant therapy has been increasingly utilized in recent years for the clinical treatment of diseased organs. Expanded use of this mode of treatment will come only when mechanisms for the long term storage of donor organs are found. Cryopreservation is an obvious answer yet, to date, freezing of whole organs has been largely unsuccessful; the technology that permits cryopreservation of isolated research will continue using mammalian organs, the problems and solutions to organ cryopreservation can be attacked directly using a vertebrate model animal with a natural tolerance of freezing. The wood frog, Rana sylvatica, readily survives for weeks in a frozen state with up to 65 % of total body water as extracellular ice; freeze tolerance is the key to winter survival for this terrestrially- hibernating species. Studies in my laboratory investigate the molecular mechanisms and biochemical regulation supporting natural freeze tolerance in R. sylvatica. The proposed research provides an organ-specific analysis of the stresses of freezing, the actions of cryoprotectants, and metabolic responses to the frozen state in the wood frog. Isolated cell systems (hepatocytes, synaptosomes) from frogs and laboratory rats are employed for model studies to determine of the effects of freezing and the actions of cryoprotectants (both natural and the agents commonly used in medical cryopreservation) in preserving both structural and metabolic integrity during freezing. Particular emphasis is placed on metabolic actions of cryoprotectants; these have received little prior attention but appear to be key to the choice of glucose as the natural cryoprotectant in the wood frog. The involvement of metabolic depression as a mechanism of ischemia tolerance for long term freezing survival is evaluated and the metabolic regulation controlling cryoprotectant synthesis in liver and permitting the extreme hyperglycemia of the frozen state is assessed.

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National Institute of General Medical Sciences (NIGMS)
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Surgery, Anesthesiology and Trauma Study Section (SAT)
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Carleton University
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Hemmings, S J; Storey, K B (2001) Characterization of sarcolemma and sarcoplasmic reticulum isolated from skeletal muscle of the freeze tolerant wood frog, Rana sylvatica: the beta(2)-adrenergic receptor and calcium transport systems in control, frozen and thawed states. Cell Biochem Funct 19:143-52
Holden, C P; Storey, K B (2000) Purification and characterization of protein kinase A from liver of the freeze-tolerant wood frog: role in glycogenolysis during freezing. Cryobiology 40:323-31
Hermes-Lima, M; Storey, K B (1998) Role of antioxidant defenses in the tolerance of severe dehydration by anurans. The case of the leopard frog Rana pipiens. Mol Cell Biochem 189:79-89
Holden, C P; Storey, K B (1997) Second messenger and cAMP-dependent protein kinase responses to dehydration and anoxia stresses in frogs. J Comp Physiol B 167:305-12
Cai, Q; Storey, K B (1997) Upregulation of a novel gene by freezing exposure in the freeze-tolerant wood frog (Rana sylvatica). Gene 198:305-12
Willmore, W G; Storey, K B (1997) Antioxidant systems and anoxia tolerance in a freshwater turtle Trachemys scripta elegans. Mol Cell Biochem 170:177-85
Cai, Q; Greenway, S C; Storey, K B (1997) Differential regulation of the mitochondrial ADP/ATP translocase gene in wood frogs under freezing stress. Biochim Biophys Acta 1353:69-78
Cai, Q; Storey, K B (1997) A novel RNA species from the turtle mitochondrial genome: induction and regulation of transcription and processing under anoxic and freezing stresses. Genome 40:534-43
Mehrani, H; Storey, K B (1997) Protein kinase C from bat brain: the enzyme from a hibernating mammal. Neurochem Int 31:139-50
Cai, Q; Storey, K B (1997) Freezing-induced genes in wood frog (Rana sylvatica): fibrinogen upregulation by freezing and dehydration. Am J Physiol 272:R1480-92

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