Tetrabromobisphenol A-induced depolarization of rat cerebellar granule cells: ex vivo and in vitro studies Dominik Diamandakis a , Elzbieta Zieminska a , Marcin Siwiec b , Krzysztof Tokarski b , Elzbieta Salinska a , Jacek Lenart a , Grzegorz Hess b , Jerzy W. Lazarewicz a, * a Department of Neurochemistry, Mossakowski Medical Research Centre, Polish Academy of Sciences, Pawinskiego 5, 02-106, Warsaw, Poland b Department of Physiology, Institute of Pharmacology, Polish Academy of Sciences, Smetna 12, 31-343, Krakow, Poland highlights  TBBPA depolarized CGC both in rat cerebellar slices and in primary culture.  Simultaneous application of MK-801, CNQX and TTX prevented TBBPA depolarization.  Prevention of intracellular Ca 2þ mobilization by TBBPA reduced CGC depolarization.  TBBPA did not change NMDA receptor radioligand binding to rat cortical membranes.  TBBPA-induced glutamate release triggered depolarization mediated by NMDAR & AMPAR. article info Article history: Received 30 October 2018 Received in revised form 5 February 2019 Accepted 7 February 2019 Available online 8 February 2019 Handling Editor: A. Gies Keywords: Neurotoxicity NMDA receptor Oxonol VI Plasma membrane potential Radioligand binding Whole-cell current clamp recording abstract The brominated flame retardant tetrabromobisphenol A (TBBPA) is toxic to cultured brain neurons, and glutamate receptors partially mediate this effect; consequently, the depolarizing effect of TBBPA on neurons is to be expected, but it is yet to be actually demonstrated. The aim of this study was to detect TBBPA-evoked depolarization and identify the underlying mechanisms. The plasma membrane potential of rat cerebellar granule cells (CGC) in cerebellar slices or in primary cultures was measured using whole- cell current clamp recordings, or the fluorescent probe oxonol VI, respectively. The contribution of NMDA and AMPA receptors, voltage-gated sodium channels and intracellular calcium mobilization was tested using their selective antagonists or inhibitors. Direct interactions of TBBPA with NMDARs were tested by measuring the specific binding of radiolabeled NMDAR ligands to isolated rat cortical membrane fraction. TBBPA (25 mM) strongly depolarized CGC in cerebellar slices, and at  7.5 mM concentration-dependently depolarized primary CGC cultures. Depolarization of the primary CGC by 25 mM TBBPA was partly reduced when MK-801 was applied alone or in combination with either TTX or CNQX, or where bastadin 12 was applied in combination with ryanodine, whereas depolarization was completely prevented when MK-801, CNQX and TTX where combined. TBBPA had no effect on the specific binding of NMDAR radio- ligands to isolated cortical membranes. These results demonstrate the depolarizing effect of TBBPA on CGC, which is mainly mediated by ionotropic glutamate receptors, while voltage-gated sodium channels are also involved. We found no evidence for the direct activation of NMDARs by TBBPA. © 2019 Published by Elsevier Ltd. 1. Introduction Brominated flame retardants (BFRs) reduce the flammability of products such as electronics, furniture, textiles and building ma- terials. The BFR 2,2 0 ,6,6 0 ,-tetrabromo-4,4 0 -isopropylidene diphenol (tetrabromobisphenol A, TBBPA) has been produced in mass quantities and utilized worldwide. TBBPA has been found in soil, water and air samples collected from industrial areas, in the blood of workers exposed to BFRs, and even in the blood and breast milk * Corresponding author. E-mail addresses: dominik.diamandakis@gmail.com (D. Diamandakis), elziem@ imdik.pan.pl (E. Zieminska), siwiec@if-pan.krakow.pl (M. Siwiec), ktok@if-pan. krakow.pl (K. Tokarski), e.salinska@imdik.pan.pl (E. Salinska), jlenart@imdik.pan. pl (J. Lenart), hess@if-pan.krakow.pl (G. Hess), jerzyl@imdik.pan.pl (J.W. Lazarewicz). Contents lists available at ScienceDirect Chemosphere journal homepage: www.elsevier.com/locate/chemosphere https://doi.org/10.1016/j.chemosphere.2019.02.032 0045-6535/© 2019 Published by Elsevier Ltd. Chemosphere 223 (2019) 64e73