In the continued presence of cholinergic ligands, the acetylcholine receptor-channel complex (AChR) in postsynaptic membranes undergoes a sequence of conformational changes. On addition of the ligand, the receptor rapidly changes from a closed channel to an open channel conformation, then slowly changes to a nonconducting state termed desensitization. The lifetime of the open channel conformation and the rate of desensitization are both dependent on the magnitude of the membrane potential, suggesting that the ligand-induced conformational changes in AChR may involve the movement of electrical charges within the membrane. Measurements of charge redistribution in AChR-containing membranes following ligand binding have not been reported. Recently, measurements of changes in the membrane partition coefficient of hydrophobic ions have been used to detect electrostatic changes in both biological and model membranes. We report here that cholinergic ligands induce changes in the partition coefficient of the hydrophobic cation tetraphenylphosphonium (TPP) into AChR-enriched membranes. The extent and time course of these changes in TPP partition coefficient are accounted for in a kinetic model. We conclude that TPP movement is a monitor of a molecular event which may be associated with the slow component of AChR desensitization.