['Tramadol is a widely used analgesic, but its mode of action is not well understood. To study the effects of tramadol on norepinephrine transporter (NET) function, we assayed the effect of tramadol on [3H]-norepinephrine ([3H]-NE) uptake and [3H]-desipramine binding to plasma membranes isolated from bovine adrenal medulla. We then characterized [14C]-tramadol binding in cultured bovine adrenal medullary cells. Tramadol inhibited the desipramine-sensitive uptake of [3H]-NE by the cells in a concentration-dependent manner (50% inhibitory concentration = 21.5 +/- 6.0 microM). Saturation analysis revealed that tramadol increased the apparent Michaelis constant of [3H]-NE uptake without changing the maximal velocity, indicating that inhibition occurred via competition for the NET (inhibition constant, K(i) = 13.7 microM). Tramadol inhibited the specific binding of [3H]-desipramine to plasma membranes. Scatchard analysis of [3H]-desipramine binding revealed that tramadol increased the apparent dissociation constant (K(d)) for binding without altering maximal binding, indicating competitive inhibition (K(i) = 11.2 microM). The binding of [14C]-tramadol to the cells was specific and saturable, with a K(d) of 18.1 +/- 2.4 microM. These findings indicate that tramadol competitively inhibits NET function at desipramine-binding sites.'] Does tramadol inhibit norepinephrine transporter function at desipramine-binding sites in cultured bovine adrenal medullary cells?