Acute toxic effects of 8-epidiosbulbin E, a 19-norclerodane diterpene from yam Dioscorea antaly, on medaka Oryzias latipes embryos.

The fractionation of an aqueous extract of yam Dioscorea antaly from Madagascar led to the isolation of terpenoids and flavonoids. Compounds were identified on the basis of modern mass spectrometry and two-dimensional nuclear magnetic resonance (2D-NMR). Toxicological effects of the most abundant isolated compound, 8-epidiosbulbin E were studied on medaka Oryzias latipes embryo-larval development. The lethal concentration (killing 50%; LC(50) ) to embryos treated 24 h before hatching and for 3 days after hatching was estimated to be 0·56 mg ml(-1) (P< 0·05). No mortality was observed with O. latipes larvae exposed after hatching until day 4. Anatomo-pathological studies of embryos exposed to 0·56 mg ml(-1) showed development anomalies of the central nervous system, liver, muscle and intestine. The present data thus extend the model of O. latipes embryos as a useful animal model to analyse the effects of food toxins.

Proteomic and phosphoproteomic analysis of cellular responses in medaka fish (Oryzias latipes) following oral gavage with microcystin-LR.

Chronic and subchronic toxicity resulting from exposure to microcystins (MCs) receives increasing attention due to the risk of bioaccumulation of these toxins by aquatic animals, including fish. The mechanisms of action of MCs that target the liver, involve modifications of protein phosphorylation resulting from phosphatases 1 and 2A inhibition. Therefore, studying phosphoprotein modifications by using a specific phosphoprotein stain Pro-Q Diamond in fish liver contaminated with MC-leucine-arginine (MC-LR), the most toxic MC, should help dissecting disturbed signaling and metabolic networks. We have recently used this technology to identify several proteins that are modulated either in expression or phosphorylation in the liver of medaka following short-term exposure to MC-LR by balneation. In the present study, we have decided to use an alternative way of introducing the toxin into fish; that is by gavage (force-feeding). This was first achieved using tritiated MC-LR and allowed us to quantify the quantity of toxin incorporated into fish and to demonstrate that the toxin is mainly accumulated in liver. Afterwards a proteomics study limited to liver cytosolic proteins of contaminated animals showed that several proteins were up or down regulated either in quantity or in phosphorylation or both. Some of them had been previously detected as modified in balneation experiments but new molecules were identified as involved in signal transduction pathways activated by the toxin. In addition, in the conditions used (5 microg toxin/g body weight) anatomopathological studies supported a process of apoptonecrosis established after 24h, which was suggested to proceed by the evolution of some of the proteins after 2h contamination.