Paraquat embryotoxicity in the Xenopus laevis cleavage phase

The high Paraquat (PQ, 1-1'-dimethyl-4,4'bipyridylium dichloride) embryotoxicity in Xenopus laevis has been shown to be due to its rapid reduction and instantaneous re-oxidation which produces a reactive oxygen species, ROS. Nevertheless, PQ did not show any effects before hatching, stage 32, which showed a resistance, in early X. laevis development, to oxidative damage. Moreover, in view of its genotoxic properties in several experimental models, we studied PQ in the X. laevis cleavage phase that, characterized by a series of rapid mitotic divisions, might be damaged by genotoxic compounds. Embryos were exposed to 20, 40, 60, and 80 mg/l PQ concentrations from stage 2 to stage 9, and then left to develop in control FETAX solution until stage 47. The 80 mg/l PQ concentration gave 19% embryo mortality at the end of the exposure time, and 16.7% larvae mortality at the end of the test; both values were statistically different from the control, 5 and 6.8% respectively. These results confirmed the high resistance in early X. laevis development to PQ oxidative damage. The malformed larva percentages in the PQ exposed groups were higher as regards the control value but did not show any concentration-response; the most frequent malformed larvae found were affected by abnormal tail flexure coupled with abnormal gut coiling. A further experiment was carried out using the same methodology, but exposing embryos only to the 80 mg/l PQ concentration. The surviving blastulae were embedded in Paraplast, then the slides were stained with 4',6-diamidino-2-phenylindole (DAPI) and the nuclei were examined with a confocal microscope. This new preliminary procedure did not reveal any significant presence of micronucleated micromeres in PQ exposed blastulae with respect to the control. Nevertheless, the mechanism by which PQ induced abnormal tail flexure after cleavage exposure remained unknown. PQ seemed to pass through the jelly coats and vitelline membrane, but it expressed teratogenicity between the 2nd and 3rd day. PQ might be accumulated in the embryos during the exposure, and might express teratogenicity later, but it did not seem to induce genotoxicity during the cleavage phase of X. laevis even at very high concentrations.