A human relevant mixture of persistent organic pollutants (POPs) and perfluorooctane sulfonic acid (PFOS) differentially affect glutamate induced excitotoxic responses in chicken cerebellum granule neurons (CGNs) in vitro

Evelien Verbruggen,Mussie Ghezu Hadera,Ragnhild Elisabeth Paulsen,Marie Kerhoas,Karin Elisabeth Zimmer,Ajay Yadav,Eva Henriëtte Willemijn Huiberts,Erik Ropstad,Hanne Friis Berntsen,Steven Verhaegen

doi:10.1016/j.reprotox.2021.01.008

Evelien Verbruggen, Mussie Ghezu Hadera + Show 8 more

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https://doi.org/10.1016/j.reprotox.2021.01.008

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Abstract

Primary cultures of cerebellar granule neurons (CGNs) derived from chicken embryos were used to explore the effects on developmental neurotoxicity by a complex defined mixture of persistent organic pollutants (POPs). Its chemical composition and concentrations were based on blood levels in the Norwegian/Scandinavian population. Perfluorooctane sulfonic acid (PFOS) alone, its most abundant compound was also evaluated. Different stages of CGNs maturation, between day in vitro (DIV) 1, 3, and 5 were exposed to the POP mixture, or PFOS alone. Their combination with glutamate, an excitatory endogenous neurotransmitter important in neurodevelopment, also known to cause excitotoxicity was evaluated. Outcomes with the mixture at 500x blood levels were compared to PFOS at its corresponding concentration of 20 μM. The POP mixture reduced tetrazolium salt (MTT) conversion at earlier stages of maturation, compared to PFOS alone. Glutamate-induced excitotoxicity was enhanced above the level of that induced by glutamate alone, especially in mature CGNs at DIV5. Glutathione (GSH) concentrations seemed to set the level of sensitivity for the toxic insults from exposures to the pollutants. The role of N-methyl-D-aspartate receptor (NMDA-R) mediated calcium influx in pollutant exposures was investigated using the non-competitive and competitive receptor antagonists MK-801 and CGP 39551. Observations indicate a calcium-independent, but still NMDA-R dependent mechanism in the absence of glutamate, and a calcium- and NMDA-R dependent one in the presence of glutamate. The outcomes for the POP mixture cannot be explained by PFOS alone, indicating that other chemicals in the mixture contribute its overall effect.

Highlights

Exposures in humans to complex mixtures of contaminants are sus pected to cause adverse health outcomes
For the persistent organic pollutants (POPs) mixture mixture reduced tetrazolium salt (MTT) activity was significantly reduced by 28 % (P < 0.0001) at DIV3, and by 35 % (P < 0.0001) at DIV5, when compared to solvent control (Fig. 1B, C)
In the presence of glutamate, Ca2+ influxes were reduced by NMDAR blockers (Fig. 4). In this situation we found MK-801 was very effective in rescuing cells from reduction of MTT induced by either perfluorooctane sulfonic acid (PFOS) or POP mixture (Fig. 6A, B)

Summary

Introduction

Exposures in humans to complex mixtures of contaminants are sus pected to cause adverse health outcomes. (III) In teractions (synergism and antagonism): applied if the combined effect of two or more chemicals is either stronger (synergistic, potentiating, supra-additive) or weaker (antagonistic, inhibitive, sub-additive, infra-additive) than would be expected on the basis of dose/concentra tion addition or response addition In this case the extent of interactions may vary according to the relative dose levels (e.g. too low to have such an effect); route of exposure (e.g. absorption too low or different meta bolism); duration of exposure (including bio-persistence of compo nents); and biological target [9,10,11]

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