Determining factors for cryoprotectant accumulation in the freeze-tolerant earthworm, Dendrobaena octaedra.

The freeze-tolerant earthworm Dendrobaena octaedra is found in most of the European forest and tundra, Siberia, North America and Greenland where it over-winters in the top soil and encounters winter frost. In response to freezing this earthworm rapidly synthesises glucose which acts as a cryoprotectant. Frost tolerance varies extensively between geographical populations, and of the populations studied so far, the Finnish worms are most and the Danish worms least frost tolerant. Little is known about the determining factors for glucose synthesis and this study therefore investigated possible roles of acclimation and the cues for synthesis of glucose, in Finnish and Danish worms. The Finnish population had significantly larger glycogen reserves than the Danish during acclimation and in all worms, glucose synthesis was the result of an almost stoichemical reduction in glycogen stores. Maximum glucose levels were reached after the onset of freezing and were significantly higher in Finnish worms where the sugar accounted for as much as 5% of the fresh weight. On average, both the total glycogen phosphorylase activity and the active enzyme pool increased during acclimation in the Finnish but not the Danish populations. However, the increase in this enzyme was only significant during the freezing process. In this study, we show contrary to previous theory that glucose synthesis is initiated before the onset of freezing and that in this species, cryoprotectant synthesis is sensitive to very small temperature changes below 0 degrees C without the presence of ice.