THE EFFECT OF DEVELOPMENTAL ATRAZINE EXPOSURE ON KEY NEUROENDOCRINE AND NEUROTRANSMISSION PATHWAYS

Atrazine is an herbicide used to control broadleaf and grassy weeds on agricultural fields in the US and other global regions, but this herbicide has been banned from use in the European Union since 2003, based mainly on risk of contamination of surface and groundwater. Atrazine is categorized as an endocrine disrupting chemical (EDC) but the specific mechanism that leads to this disruption is not yet clearly defined. This study investigated the hypothesis that the main target of neuroendocrine atrazine toxicity is the kisspeptin system, which then leads to multiple adverse health outcomes that are seen affecting multiple endocrine axes. Furthermore, an embryonic atrazine exposure was expected to result in adverse effects on neurohormones and associated gene expression along the endocrine axes into adulthood. Using the zebrafish model, concentrations of estradiol, dopamine, kisspeptin, and luteinizing hormone were measured in atrazine exposed larvae as well as male and female adult brains using ELISA. Adult wild type zebrafish were bred to obtain embryos, collected at 1 hour post fertilization (1 hpf), and randomly assigned to 0, 0.3, 3, or 30 ppb (µg/L) atrazine treatment, surrounding the current US EPA regulatory level in drinking water of 3 ppb. Exposure ceased at the end of embryogenesis (72 hpf) and fish placed in filtered aquaria water for continued development. Zebrafish were either collected for larval evaluation [72, 120, 144, or 168 hpf] or grown to different timepoints in adulthood for collection of brains [6 months post fertilization (mpf), 2 years post fertilization (ypf), or 2.5 ypf]. Gene expression of neuroendocrine molecular targets was examined to determine if an embryonic atrazine exposure perturbed neuroendocrine development using qPCR. Behavior analysis was conducted on larvae and in adults to assess downstream functional changes related to dopaminergic signaling. Significant findings in gene expression, neurohormone and protein concentration, and behavior were observed and a CRISPR-Cas9 knockdown model was designed to further investigate the kisspeptin system as a viable target of atrazine toxicity in connection with the multitude of adverse effects. Studies to further examine perturbations along the pathways associated with these biomarkers are necessary to elucidate the mechanism of atrazine and further characterize the role of the kisspeptin system in atrazine toxicity.