A High-Throughput, High-Content Analysis of Dopaminergic Neurodegeneration in Caenorhabditis elegans Exposed to Per- and Polyfluoroalkyl Substances.

Dopaminergic neurodegeneration is a hallmark of Parkinson's disease (PD), and environmental contaminants have been implicated in disrupting dopaminergic pathways. However, practical in vivo workflows for rapid, standardized, and accessible assessment of dopaminergic neurotoxicity remain limited. In this study, we built on our laboratory's established high-throughput framework and implemented a high-content imaging workflow to quantify DA neurodegeneration in Caenorhabditis elegans following exposure to representative per- and polyfluoroalkyl substances (PFAS). We evaluated the neurotoxic effects of perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorohexanesulfonic acid (PFHxS), perfluorohexanoic acid (PFHxA), and three PFAS mixtures with environmentally relevant component ratios. Functional relevance was assessed using dopamine-dependent behavioral endpoints, including basal slowing response (BSR) and area-restricted search (ARS). PFOS exhibited the greatest potency, followed by PFHxS, PFHxA, and PFOA, based on morphological degeneration and benchmark concentration modeling. Structural neuronal damage was significantly associated with behavioral impairment. Under mixture conditions, neurotoxicity was more strongly associated with PFOS molar fraction than with total PFAS concentration (ΣPFAS), suggesting a composition-dependent toxicity profile. Collectively, these findings establish a scalable in vivo framework for assessing PFAS-induced dopaminergic neurotoxicity and support the potential use of this platform for screening environmental pollutants with dopaminergic neurotoxic potential.