Triggering of cryoprotectant synthesis by the initiation of ice nucleation in the freeze tolerant frog, Rana sylvatica

The triggering of cryoprotectant synthesis was examined in the freeze tolerant wood frog, Rana sylvatica. A slow decrease in ambient temperature (1°C every 2 days) from 3° to -2.1 °C was used to search for a specific trigger temperature. None was found. Instead it was found that, despite subzero temperature, animals which remained in a supercooled unfrozen state had low blood glucose (1.66±0.44 μmol/ml) while those which had frozen had high blood glucose (181±16 μmol/ml). These results indicate that it is the initiation of ice nucleation, rather than a specific subzero temperature, which triggers cryoprotectant glucose synthesis. This was confirmed by monitoring the freezing curves for individual frogs with sampling of blood and tissues at various times relative to the initiation of nucleation (detected as an instantaneous temperature jump from -3 to -1°C). Animals sampled before nucleation had low blood and liver glucose contents and a low percentage of liver phosphorylase in the a form. Within 4 min of the initiation of freezing, however, blood glucose had jumped to 16 μmol/ml and liver glucose to 39.5 μmol/g wet weight. Glucose in both compartments continued to increase as the time of freezing increased correlated with an in