Electrophysiological techniques for the assessment of in vitro neurotoxicology have several advantages over other currently-used methods (for example, morphological techniques), including the ability to detect damage at a very early stage. Novel recording techniques based on microelectrode arrays are available, and could improve recording power. In this study, we investigated how a three-dimensional microelectrode array detects the electrophysiological endpoints of neurotoxicity. We conclude that electrophysiology sensitively reveals neurotoxic actions, and that three-dimensional microelectrode arrays could be proposed for use in neurotoxicology as recording tools that allow easy and sensitive multisite recording, from both rodent and human brain tissue.