Adaptations of metabolism for freeze tolerance in the gray tree frog, Hyla versicolor

Abstract

Biochemical adaptations allowing the natural survival of extracellular freezing were examined in the gray tree frog, Hyla versicolor. Laboratory-reared immature adults froze between −1 and −1.5 °C and survived 5 days of freezing at −2 °C as well as repeated rapid bouts of freeze–thaw. Measurements of ice content showed 41.5% of total body water frozen. Glycerol accumulated as the cryoprotectant in sexually mature adult H. versicolor (423 μmol/mL in blood) while both glycerol and glucose accumulated in immature adults (16.3 ± 6.8 and 25.9 ± 11.6 μmol/mL in blood, respectively). Cryoprotectant synthesis was freezing stimulated only and did not occur over long-term cold acclimation at 0 to 1 °C. Cryoprotectant synthesis was correlated with a 203% increase in liver total phosphorylase activity and an increase in phosphorylase a content from 40 to 60%. Activities of 15 other enzymes of intermediary metabolism were determined in liver and leg muscle; activities of most enzymes increased with freezing exposure as did soluble protein content. Survival of freezing depends upon anaerobic mechanisms of energy production in tissues. Frogs frozen at −2 °C accumulated lactate in liver and muscle. Energy charge dropped in both tissues and the creatine phosphate reserves of muscle were depleted.