Joint Action Toxicity of Arsenic (As) and Lead (Pb) Mixtures in Developing Zebrafish

Arsenic (As) and lead (Pb) are environmental pollutants found in common sites and linked to similar adverse health effects. Multiple studies have investigated the toxicity of each metal individually or in complex mixtures. Studies defining the joint interaction of a binary exposure to As and Pb, especially during the earliest stages of development, are limited and lack confirmation of the predicted mixture interaction. We hypothesized that a mixture of As (iAsIII) and Pb will have a concentration addition (CA) interaction informed by common pathways of toxicity of the two metals. To test this hypothesis, developing zebrafish (1-120 h post fertilization; hpf) were first exposed to a wide range of concentrations of As or Pb separately to determine 120 hpf lethal concentrations. These data were then used in the CA and independent action (IA) models to predict the type of mixture interaction from a co-exposure to As and Pb. Three titration mixture experiments were completed to test prediction of observed As and Pb mixture interaction by keeping the Pb concentration constant and varying As concentrations in each experiment. The prediction accuracy of the two models was then calculated using the prediction deviation ratio (PDR) and Chi-square test and regression modeling applied to determine type of interaction. Individual metal exposures determined As and Pb concentrations at which 25% (39.0 ppm Pb, 40.2 ppm As), 50% (73.8 ppm Pb, 55.4 ppm As), 75% (99.9 ppm Pb, 66.6 ppm As), and 100% (121.7 ppm Pb, 77.3 ppm As) lethality was observed at 120 hpf. These data were used to graph the predicted mixture interaction using the CA and IA models. The titration experiments provided experimental observational data to assess the prediction. PDR values showed the CA model approached 1, whereas all PDR values for the IA model had large deviations from predicted data. In addition, the Chi-square test showed most observed results were significantly different from the predictions, except in the first experiment (Pb LC₂₅ held constant) with the CA model. Regression modeling for the IA model showed primarily a synergistic response among all exposure scenarios, whereas the CA model indicated additive response at lower exposure concentrations and synergism at higher exposure concentrations. The CA model was a better predictor of the Pb and As binary mixture interaction compared to the IA model and was able to delineate types of mixture interactions among different binary exposure scenarios.