Differential Developmental Neurotoxicity and Tissue Uptake of the Per- and Polyfluoroalkyl Substance Alternatives, GenX and PFBS

Abstract

Per- and polyfluoroalkyl substances (PFAS) are a class of synthetic chemicals with several applications. Multiple adverse health effects are reported for longer carbon chain (≤C8) PFAS. Shorter carbon chain PFAS, [e.g., hexafluoropropylene oxide dimer acid (HFPO-DA; GenX) and perfluorobutanesulfonic acid (PFBS)] were introduced as alternatives. Past studies indicate that longer-chain PFAS are neurotoxic targeting the dopamine pathway, but it is not known if shorter-chain PFAS act similarly. This study aimed to evaluate developmental neurotoxicity and tissue uptake of GenX and PFBS using the zebrafish (Danio rerio). First, acute toxicity was assessed by measuring LC50 at 120 h postfertilization (hpf). Body burden was determined after embryonic exposure (1-72 hpf) to sublethal concentrations of GenX or PFBS by LC-ESI-MS/MS. Locomotor activity using a visual motor response assay at 120 hpf and dopamine levels at 72 hpf was assessed after embryonic exposure. PFBS was more acutely toxic and bioaccumulative than GenX. GenX and PFBS caused hyperactivity at 120 hpf, but stronger behavioral alterations were observed for PFBS. An increase in whole organism dopamine occurred at 40 ppb of GenX, while a decrease was observed at 400 ppb of PFBS. Differences detected in dopamine for these two PFAS indicate differential mechanisms of developmental neurotoxicity.

Keywords: HFPO−DA; behavior; dopamine; perfluoroalkyl substances; perfluorobutanesulfonic acid; polyfluoroalkyl substances; toxicokinetics; zebrafish.

Figure 1.

Body burden at 72 hpf and concentration of dosing solution at exposure initiation for GenX and PFBS. (A) Body burden of whole pooled eleuthero-embryos at 72 hpf at the cessation of chemical exposure (N = 4 with 70 subsamples pooled per treatment per replicate). (B) Concentration of dosing solutions at the initiation of chemical exposure (at 1 hpf, N = 4). Error bars are the standard deviation.

Figure 2.

Body burden at 120 hpf and concentrations of dosing solutions at the initiation of exposure, cessation of exposure, and time of behavior analysis. (A) Body burden of whole pooled larvae at 120 hpf aligning with time of behavior analysis 48 h after chemical exposure ceased (N = 3–4 with 70 larvae pooled per treatment per replicate). (B) Concentration of GenX dosing solution at the initiation of exposure (1 hpf), at the end of chemical exposure (72 hpf), and 48 h after the chemical exposure ceased (at 120 hpf when behavior analysis was completed, N = 2–4). (C) Concentration of PFBS dosing solution at the initiation of exposure (1 hpf), at the end of chemical exposure (72 hpf), and 48 h after the chemical exposure ceased (at 120 hpf when behavior analysis was completed, N = 3–4). Error bars are standard deviation.

Figure 3.

Visual motor response behavior assay in zebrafish larvae at 120 hpf following an embryonic exposure (1–72 hpf) to GenX (A, C, E) or PFBS (B, D, F). N = 7 biological replicates for GenX with 15–19 subsamples per treatment per replicate to total 129–132 fish per treatment group. N = 6 for PFBS exposure with 18–19 subsamples per treatment per replicate to total 111–113 fish per treatment group. Error bars represent standard deviation. *p < 0.05 compared to the control treatment group within each phase. (D1: first dark phase; L1: first light phase; D2: second dark phase; L2: second light phase; D3: third dark).

Figure 4.

Average whole-body dopamine content in eleutheroembryos exposed to (A) GenX or (B) PFBS from 1 to 72 hpf. N = 5 (for PFBS) and N = 10 (for GenX) with pools of 35−70 fish per treatment group per biological replicate. Error bars are standard deviation. *p < 0.05.