In vitro systems have been proposed, but not yet demonstrated, as a method to assess the neurotoxicity of compounds in an efficient and rapid manner. Although such tests are desired both for pharmaceuticals and environmental agents, such a battery has yet to be developed that is based on known processes of nervous system dysfunction. In this review article, characteristics and potential limitations associated with in vitro methods are discussed. Many of these features have been identified from a larger body of work examining the neurotoxicity of environmental agents and the mechanisms underlying activity of known neurotoxicants. These issues include relevant drug concentrations, factors that limit or alter drug accessibility to the nervous system, and the need for assays to reflect biologically meaningful end points. This commentary briefly surveys in vitro systems of increasing biological complexity currently available for toxicity testing, from single cell types to systems that preserve some aspects of tissue structure and function. A small number of studies to evaluate drugs for cytotoxicity and biological responses in vitro are presented as representative of the current state of the field and to provide a reference and direction for additional development of methods to assess a compound's potential for neurotoxicity.