232: Transport of opiates by P-glycoprotein expressed in preparations of human placental inside-out vesicles

Abstract

The use of methadone (MT) or buprenorphine (BUP) for treatment of the pregnant opiate-dependent patient improves maternal and neonatal outcome. However, both medications are associated with development of neonatal abstinence syndrome (NAS). The incidence and intensity of NAS should correlate with the concentration of the opiate in the fetal circulation which is regulated by the placenta. The function of human placental P-glycoprotein (P-gp) is the extrusion of its substrates from the feto-placental unit to the maternal circulation. Previously, we demonstrated that MT, BUP, and morphine (MR) inhibit P-gp transport of its prototypic substrate paclitaxel indicating opiate/P-gp interactions. The goal of this investigation was to determine the transport kinetics of [3H]-MT, [3H]-BUP, and [3H]-MR by placental P-gp. Opiate interaction with P-gp was determined by their stimulation of ATP hydrolysis by human P-gp expressing membranes. ATP-dependent transport of [3H]-MT, [3H]-BUP, and [3H]-MR was determined by inside-out brush border membrane vesicles prepared from human placental tissue (n = 60). Specific transport of each opiate was calculated from the difference in its transfer in presence and absence of 600 μM verapamil. The three opiates stimulated ATP hydrolysis by P-gp expressing membranes. The transport of MT and MR by placental P-gp was saturable and revealed the following kinetics parameters: [3H]-MT Kt: 300 ± 100 nM and Vmax 4.3 ± 0.6 pmol/mg protein min; [3H]-MR Kt: 226 ± 200 nM, and Vmax: 3.6 ± 2.4 pmol/mg proteinmin. Verapamil did not inhibit the uptake of [3H]-BUP by inside-out vesicles, indicating that it is not transported by P-gp. The data demonstrate that placental P-gp is involved in regulating the maternal-to-fetal transfer of MT and MR.