Short Communication Long-term Anoxia in Artemia Cysts

Among the many adaptations associated with the rigorous life history of the brine shrimp, Artemia, is the striking resistance of the encysted embryo (cyst) to oxygen lack (anoxia). Dutrieu and Chrestia-Blanchine (1966) reported that these cysts could be incubated in sea water under anoxic conditions for over 5 months without a decrease in viability. Subsequent work showed that anoxic cysts did not carry out a conventional lactate-producing metabolism and that the breakdown of trehalose, a disaccharide required for the energy metabolism of aerobic cysts, was not detected over an g-h period of anoxia (Ewing and Clegg, 1969). Comprehensive studies of the nucleotide pool by Stocco et al. (1972) suggested that utilization of the unusual guanine nucleotide diguanosine tetraphosphate (Gp4G) might provide the free energy presumably required to support the maintenance of anoxic cysts. Most recently, Hand and Gnaiger (1988) used calorimetric methods to show that anoxic energy metabolism, as reflected by heat production, was reduced to less than 2 % of aerobic values during short-term anoxia (10 h). Those authors also calculated that the utilization of Gp4G could account for only about 2 % of anoxic heat production and suggested that the very slow catabolism of trehalose might be a more likely explanation for their results. We examined that suggestion by measuring the concentrations of trehalose and other carbohydrates in cysts incubated under anoxic conditions for 3 months (Clegg and Jackson, 1989). We found no measurable change in trehalose and glycerol content under these conditions, but did observe a statistically significant decrease in the glycogen content of anoxic cysts. In the present paper we report comparable data from additional studies on anoxic cysts incubated for 3 and 6 months, and we measure the free amino acid pool, cyst dry mass and hydration. Cysts of Artemia fransiscanus were collected from the solar salt ponds near Hayward, California, in the summer of 1987 and were processed and dried as previously described (Clegg, 1986). Sea water containing penicillin (2000 units ml") and streptomycin (O.lmgmP) was passed through 0.1/im filters and gassed with 100 % N2 for 3 h. Air-dried cysts (about 80mg) were placed in 5 ml glass tubes and incubated dry at room temperature (about 23 °C) under four changes of 100% N2 for 3 days to remove air trapped in their shells. Anoxic sea