Abstract

In many invertebrates, environmental hypoxia involves the accumulation of anionic endproducts such as succinate, propionate, and acetate (Grieshaber et al 1994 Rev Physiol Biochem Pharmacol 125: 44) during anaerobic energy metabolism. Especially in freshwater or semi-terrestrial invertebrates, these metabolites interfere with the anion:cation-balance and the comparatively  low osmotic concentration. Increasing amounts of anion equivalents are compensated by a decrease of chloride in the leech Hirudo medicinalis (Hildebrandt JP and Zerbst-Boroffka I 1992 J Exp Zool 263: 374). The objective of this study was to test whether the same adaptive mechanisms are present in the oligochaete Lumbricus terrestris. The use of several methods allowed us to analyze all osmolytes in individual hemolymph samples during normoxia and environmental hypoxia.
In normoxic animals, sodium (81,0 ± 7 mmol x l-1) accounted for almost half of the osmotic concentration (169 ± 10 mosmol x kg-1). The anionic fraction contained primarily chloride (48 ± 5 mmol x l^-1), hydrogencarbonate (13,4 ± 4,0 mmol x l^-1) and organic acids (11,6 ± 5,0 mmol x l^-1; 22,9 ± 9,8 meq x l^-1, mainly malate (6,9 ± 2,6 mmol x l^-1)). During the first three hours of severe hypoxia (nitrogen), the increase in succinate was counterbalanced by an equimolar degradation of malate and increasing sodium concentrations. The subsequent accumulation of organic acids (succinate, propionate and acetate; after 48 h 11,2 ± 1,9 mmol x l^-1, 5,7 ± 1,2 mmol x l^-1 and 3,2 ± 0,5 mmol x l^-1, respectively) mainly was counterbalanced by a sharp drop in hydrogencarbonate (from about 13 mmol x -1 to 1 mmol x l^-1 after 48 h). L. terrestris accumulated much less lactate (1,9 ± 1,0 mmol x l^-1) under hypocapnic hypoxia than H. medicinalis (about 13 mmol x l^-1 after 48h, Hildebrandt JP 1992 J Exp Zool 261: 379).  Osmolality and chloride concentrations remained constant during the first 24 hours, but after 48 h decreased to 151,6 ± 4,5 mosmol x kg^-1 and 40,8 ± 3,2 mmol x l^-1, respectively. Concentrations of potassium and calcium stayed constant throughout the entire experiment (about 5 mmol x l^-1 and 2 mmol x l^-1, respectively).
In accordance with other clitellates, malate serves as an initial energy reserve under hypoxic conditions in L. terrestris. However, the accumulation of organic acids is much less pronounced than in other anoxia-tolerant annelids. In contrast to H. medicinalis, the decline of hydrogencarbonate is sufficient to counterbalance the accumulation of the anionic endproducts propionat and acetat under prolonged hypocapnic hypoxia. Further studies will test whether this mechanism is involved also under normocapnic hypoxia.