The present study investigated the effects of carbofuran on the energy metabolism (levels of glycogen, total proteins, total lipids, triglycerides, and lipoperoxidation), Na+/K+ATPase activity, and reproductive parameters (formation of couples, ovigerous females, and mean number of eggs) in the freshwater amphipods Hyalella pleoacuta and Hyalella curvispina. These crustaceans live in limnetic environments of the plateau (H. pleoacuta) and coastal plain (H. curvispina) of the state of Rio Grande do Sul in southern Brazil. The animals were collected in the winter of 2006 in the Vale das Trutas (28 degrees 47'00''S-49 degrees 50'53''W) in the Municipality of São José dos Ausentes, and in Gentil Lagoon (29 degrees 56'30''S, 50 degrees 07'50''W) in the Municipality of Tramandaí. In the laboratory, the amphipods were kept submerged in aquariums under controlled conditions of photoperiod (12 h light: 12 h dark), temperature (23 degrees C+/-1), and constant oxygenation. Animals were exposed to carbofuran at a dose of 5 or 50 microg/L for a period of 7 days. At the end of this period, the animals were immediately frozen for determination of the biochemical parameters, lipoperoxidation levels (TBARS), and enzyme Na+/K+ATPase activity. During each day of culture, several reproductive parameters were observed. Statistical analysis (ANOVA) revealed that carbofuran induces significant decreases in glycogen, proteins, lipids, triglycerides, and Na+/K+ATPase, as well as a significant increase in lipoperoxidation levels. Studies of all the biochemical parameters seem to be quite promising, in order to assess and predict the effects of toxicants on non-target organisms. The results also suggest that the reproductive parameters (formation of couples, ovigerous females and mean number of eggs) may provide sensitive criteria for assessing ecotoxicological effects. Furthermore, H. pleoacuta and H. curvispina are suitable organisms for use in toxicity tests, and we suggest that they are sensitive species that could be used in monitoring studies.