Dual phases of functional change in norepinephrine transporter in cultured bovine adrenal medullary cells by long-term treatment with clozapine.

The effects of long-term treatment with clozapine, a prototype of atypical antipsychotic drugs, on the functional activity, synthesis and mRNA of norepinephrine (NE) transporter were examined in bovine adrenal medullary cells in culture. Treatment of cells with clozapine at 0.1–3.0 µm concentrations produced dual phases of changes in [3H]NE uptake, i.e. the first phase showed a decrease in [3H]NE uptake at 2–48 h, and the following phase showed an increase in uptake at 72–168 h. Treatment with clozapine for 6 h decreased Vmax to 40% of the control without changing the Km value for [3H]NE uptake. However, treatment with clozapine for 96 h increased Vmax by 56% over the control without a change in Km. Scatchard plot analysis of [3H]desipramine (DMI) binding to membranes isolated from cells treated with clozapine for 6 h revealed a decrease in Bmax without any change in Kd; in contrast, treatment with clozapine for 96 h caused an increase in Bmax without any change in Kd. Both actinomycin D and cycloheximide, which are inhibitors of protein synthesis, suppressed the clozapine (96 h)-induced increase in [3H]NE uptake. Treatment of cells with clozapine for 12–96 h increased the level of NE transporter mRNA in a concentration-dependent manner (0.3–3.0 µm). These findings suggest that treatment of cells with clozapine results in the down-regulation and subsequent up-regulation of NE transporter. The latter change may be caused by the synthesis of new proteins of NE transporter via an increase in its mRNA.