We studied desensitization of nicotinic acetylcholine (ACh) receptor channels in the clonal BC3H-1 cell line. We measured the current response to rapid perfusion of outside-out patches with 1 microM to 5 mM ACh, carbamylcholine and suberyldicholine. After binding to the receptors and opening the ion channels, all agonists induce a rapid, concentration-dependent decay of channel activity. The time constant of the current decay ranged from several seconds at low agonist concentrations to about 50 ms at saturating concentrations. The decay rate at saturating concentrations was independent of voltage. The ratio of steady-state to peak current ranged from 0.5 at low agonist concentrations to 0.02 or less at high concentrations. The rate of recovery from desensitization after removal of agonist was also measured. For ACh, the recovery time constant was 320 ms; recovery from desensitization by carbamylcholine was twice as fast. A linear kinetic results. The data are consistent with a cyclic model, although, it is not possible to uniquely determine all of the rate constants in this scheme. The results are compared with competitive binding and single channel studies of desensitization in BC3H-1 cells.