The neurotoxic potencies of the aryl phosphoramidate isofenphos ( O-ethyl O-(2-isopropoxycarbonyl)phenylisopropylphosphoramidothioate) and three of its metabolites, isofenphos oxon, des- N-isopropyl isofenphos (DNI), and des- N-isopropyl isofenphos oxon (DNIO), were examined in the American cockroach ( Periplaneta americana). In dissected preparations, the electrical responses of the normally quiescent giant interneurons (GIs) to each chemical consisted of repeated, high-frequency bursts of impulses. The mean times from application (10 ppm) to the ventral nerve cord (VNC) until the induction of GI bursting were 10.8, 32.5, 40, and 85 min for DNIO, isofenphos oxon, DNI, and isofenphos, respectively, as compared with 2.3 min for the cholinesterase inhibitor, paraoxon. Cholinesterase assays on VNCs treated in vitro showed that only paraoxon (I 50 = 3.1 × 10 −7 M) and DNIO (I 50 = 3.1 × 10 −6 M) inhibited in a dose-dependent manner. By contrast, cholinesterase assays on VNCs treated in vivo indicated that all compounds inhibited cholinesterase in a dose-dependent manner; I 50 values were 3.1 × 10 −6 M, 3.6 × 10 −6 M, 2.3 × 10 −5 M, 3.3 × 10 −5 M, and 9.8 × 10 −3 M for paraoxon, DNIO, isofenphos oxon, DNI, and isofenphos, respectively. At inhibitor concentrations required to induce bursting after 15 min, in vivo cholinesterase activity ranged from 27 to 42% of control activity. In vivo assays performed after pretreatment with the mixed-function oxidase inhibitor β-diethylaminoethyl diphenylpropylacetate (SKF 525A) showed that cholinesterase inhibition by each metabolite was reduced by at least one order of magnitude compared with untreated preparations. The results are consistent with the hypothesis that the neurotoxic action of isofenphos involves oxidative bioactivation and subsequent cholinesterase inhibition by its metabolites, as well as an induction of high-frequency bursting in identifiable GIs.