Microplastics- and copper-induced changes in neurogenesis and DNA methyltransferases in the early life stages of zebrafish

Abstract

In this study, zebrafish embryos were exposed to microplastics (MPs, 2 mg/L) and copper (Cu, 60 and 125 μg/L), alone or combined, for 14 days, and the development of motor neurons was assessed through gene expression and immunohistochemistry. DNA methyltransferases (DNMTs) genes expression was also evaluated. The results showed a downregulation of neuronal proliferation (sox2, pcna), neurogenesis (neuroD, olig2), and motor neurons development (islet) related genes, implying potential deficits in the neurogenesis of the exposed zebrafish early life stages. Downregulation of the maintenance and de novo DNMTs expression was also found, indicating that the DNA methylation patterns could be modulated by MPs and Cu. A high relative volume of proliferating cell nuclear antigen (PCNA)-positive cells was found in the fish retina from the MPs exposed group, suggesting that MPs increased the rate of cellular division. In contrast, a significant decrease of PCNA-positive cells, and therefore a lower cell proliferation, was found in the retina and brain of zebrafish exposed to Cu and Cu + MPs, which could lead to cognitive and behavioral functions impairment. No alterations were found in the relative volume of ISL1&2-positive cells. This study contributes to the knowledge of the mechanisms by which MPs and Cu cause neurotoxicity, fundamental for a comprehensive and realistic ecological risk assessment in aquatic populations.

Keywords: Danio rerio; Gene expression; Heavy metals; Methylation; Plastic microspheres; Stereological analysis.