To determine the appropriate time points during ethanol treatment at which to assess the biochemical and behavioral effects of compromising ADH function, we observed the kinetics of the onset of intoxication in wild-type animals. We placed individual worms (n=6) in a copper ring on a plate with 500 mM exogenous ethanol, and recorded their locomotion continuously for 15 minutes, starting immediately after the animals had been placed on the plate. Animals rapidly decrease speed in the first few minutes of ethanol exposure, and reach a plateau at 6 minutes, suggesting that the animals accumulate enough ethanol in this time to become maximally impaired (Figure 1a). When we examined the data in 1-minute bins, we found that the neighboring bins were not different after 6 minutes (P > 0.05, Figure 1b). We chose 10 minutes exposure as an initial time point for biochemical and behavioral testing for convenience and consistency with previous experimental designs (Davies et al., 2003, 2004; Kapfhamer et al., 2008).
