Fig. 7

Knockdown study to determine the essentiality of ppar genes for VSR hyperactivity in PFOS-exposed zebrafish. (A) CRISPR/Cas9-based gene editing experimental design were embryos that experience gene knockdown (set 1 or set 2 crispants) relative to negative control crispants (NC) that did not experience gene knockdown and were subsequently exposed to 7.86μ⁢M PFOS at 1 dpf. The chemical was removed at 4 dpf and behavior was assessed at 5 dpf. VSR within 3 s after light–dark transition (VSR2) in (B) pparg-knockdown, (C) pparaa/ab-knockdown, or (D,E) pparda/db-knockdown larvae exposed to 7.86μ⁢M PFOS are shown. Data are represented as box and violin plots and comprise 1 value per larva. Significance was determined by Tukey-adjusted estimated marginal means following a linear mixed effects model. Significant differences (𝑝<0.05) between groups are indicated by different letters. Individual box plots comprise a box that describes the IQR, a bold line that represents the median, and whiskers that indicate the calculated minimum (25th percentile –1.5× IQR) and the calculated maximum (75th percentile +1.5× IQR). Dots represent outliers beyond the calculated minima or maxima. Violins around the box plots describe the kernel probability density of the underlying data. Replicate numbers were 24–41 for pparg-knockdown (B), 31–44 for pparaa/ab-knockdown (C), 10–47 for initial pparda/db-knockdown, and 44–75 for pparda/db-knockdown with revised guide sets. Summary data can be found in Excel Tables S20–S23. Note: CRISPR, Clustered Regularly Interspaced Short Palindromic Repeats; DNT, developmental neurotoxicity; dpf, days postfertilization; IQR, interquartile range; PFOS, perfluorooctanesulfonic acid; ppar, peroxisome proliferator-activated receptor; VSR, visual startle response.