We examined the interactions of d,l-laudanosine, a potentially epileptogenic metabolite of the neuromuscular relaxant atracurium besylate, with γ-aminobutyric acid (GABA) and opioid binding sites, all of which have been implicated in seizure activity. Laudanosine was almost ineffective at [ 3H]muscimol binding to high-affinity GABA receptors (IC 50 = 100 μM). However, laudanosine displayed an inhibitory effect at the low-affinity GABA receptors labeled by [ 3H]bicuculline methochloride, with an IC 50 value of 10 μM. At the opioid receptor subtype, laudanosine lowered radiolabeled opioid binding at the μ 1, μ 2, δ, κ 1, and κ 3 receptors with K i values of 2.7, 13, 5.5, 21, and 24 μM, respectively, concentrations seen clinically in blood and approaching those measured in cerebrospinal fluid. Saturation studies of μ 1, μ 2, δ, and κ 3 sites in the presence of laudanosine revealed competitive interactions, with increases in the apparent K d values but without significant changes in the maximal numbers of binding sites. In addition, we investigated whether the in vitro laudanosine-opioid receptor interaction would also be expressed by analgesic physiologic effects. We found that laudanosine elicited a dose-dependent analgesia in mouse tail-flick assay that was attenuated by coadministration of β-funaltrexamine ( μ 1- and μ 2-sselective antagonist) and of naloxonazine ( μ 1 antagonist), but not by nor-binaltophimine ( κ 1-selective antagonist) or naltrindole (δ-selective antagonist), indicating a μ 1 mechanism for analgesia-mediated properly of laudanosine. There is evidence suggesting μ 2 activity as well, but this is due to the ability of laudanosine to elicit analgesia when given intrathecally. We also observed cross-tolerance between laudanosine and morphine, as well as a partial effect of laudanosine on gastrointestinal transit. These results suggest an interaction between laudanosine and the low-affinity GABA receptor, as well as opioid μ 1 and μ 2 receptors.