BIOCHEMISTRY BELOW 0-DEGREES-C - NATURES FROZEN VERTEBRATES

Although alien to man, the ability to endure the freezing of extra-cel lular body fluids during the winter has developed in several species o f terrestrially hibernating frogs and turtles as well as in many speci es of insects and other invertebrates. Wood frogs, for example, can en dure freezing for at least 2 weeks with no breathing, no heart beat or blood circulation, and with up to 65% of their total body water as ic e. Our studies are providing a comprehensive view of the requirements for natural freezing survival and of the physical and metabolic protec tion that must be offered for effective cryopreservation of vertebrate organs. Molecular mechanisms of natural freeze tolerance in lower ver tebrates include: 1) control over ice crystal growth in plasma by ice nucleating proteins, 2) the accumulation of low molecular weight cryop rotectants to minimize intracellular dehydration and stabilize macromo lecular components, and 3) good ischemia tolerance by all organs that may include metabolic arrest mechanisms to reduce organ energy require ments while frozen. Cryomicroscopy of tissue slices and magnetic reson ance imaging (MRI) of whole animals is revealing the natural mode of i ce propagation through an organism. MRI has also revealed that thawing is non-uniform; core organs (with high cryoprotectant levels) melt fi rst, facilitating the early resumption of heart beat and blood circula tion, Studies of the production and actions of the natural cryoprotect ant, glucose, in frogs have shown its importance in maintaining a crit ical minimum cell volume in frozen organs and new work on the metaboli c effects of whole body dehydration in 3 species of frogs has indicate d that adaptations supporting freeze tolerance grew out of mechanisms that deal with desiccation resistance in amphibians, Studies of the re gulation of cryoprotectant glucose synthesis by wood frog liver have s hown the role of protein kinases and of alpha and beta adrenergic rece ptors in regulating the glycemic response, and of changes in membrane glucose transporter proteins to facilitate cryoprotectant distribution .