Figure 2

Behavior analysis of scn1lab^s552 homozygous mutant larvae during long-term exposure of AEDs. To examine the effect of long-term exposure of AEDs on scn1lab^s552 swim behavior was analyzed using the DanioVision running EthoVision XT software (DanioVision, Noldus Information Technology). As previously described (Sourbron et al., 2017b), 6 dpf scn1lab^s552 homozygous mutants were identified based on pigmentation and arrayed in a 96-well plate. Larvae were treated with 2% DMSO (control) or compound using drug concentrations previously published (Zhang et al., 2015; Sourbron et al., 2016, 2017a). After 22-h incubation at 28°C with a 14:10 h light/dark photoperiod, and 30 min of chamber habituation, the swim behavior of 7 dpf larvae was analyzed for 10 min under dark conditions. Both the (A) total distance moved and (B) velocity were analyzed by normalizing the activity of AED treated larvae to vehicle treated controls (previously described as method A, Sourbron et al., 2017b). Graphs represents mean (±SD) normalized to control treated larvae from three independent experiments using the average from 12 larvae per treatment each time. Statistical significance was determined by one-way ANOVA followed by Dunnet's multiple comparison test. No statistical significance changes in swim behavior of scn1lab^s552 homozygous mutants when exposed to known AEDs or putative AEDs. Recommended treatments (blue), contraindicated treatments (red) and recent experimental treatments (yellow) for DS are shown. Acetaminophen is also shown (gray). These behavioral results, which also fail to distinguish between any of the experimental situations tested, are entirely consistent with the data independently obtained using the iZAP system and fail to support the validation of this low micromolar, 22-h exposure assay as an effective means to identify drugs with antiepileptic activity in scn1lab^s552 homozygous mutants.