Characterization of [ 3H]mazindol binding sites in cultured monkey amniotic epithelial cells

Our previous studies showed that monkey amniotic epithelial cells (MAEC) synthesize and release catecholamines and possess D1 and D2 dopamine (DA) receptors (Elwan, M.A., Ishii, T., Ono, F. and Sakuragawa, N., Evidence for the presence of dopamine D1 receptor mRNA and binding sites in monkey amniotic epithelial cells. Neurosci. Lett., 262 (1999) 9–12; Elwan, M.A., Ishii, T. and Sakuragawa, N., Detection of dopamine D2 receptor mRNA and binding sites in monkey amniotic epithelial cells. J. Neurosci. Res., 56 (1999) 316–322; Elwan, M.A., Thangavel, R., Ono, F. and Sakuragawa, N., Synthesis and release of catecholamines by cultured monkey amniotic epithelial cells. J. Neurosci. Res., 53 (1998) 107–113). In the present study we tested the presence of DA transporter (DAT) in MAEC using radioligand binding experiments. Saturation studies showed that [ 3H]mazindol binds to a high affinity site with K D and B max values of 7.85±1.25 nM and 123.22±18.34 fmol/mg protein, respectively. Competition studies indicated that selective DAT inhibitors are potent displacers of [ 3H]mazindol binding, compared to inhibitors of other types of transporters. The rank order of potency of the competing drugs is consistent with the pharmacology of DAT. These results provide, for the first time, clear evidence that MAEC natively possess DAT binding sites and suggest that MAEC may provide a potential primate cell model to study DA release and uptake processes and to explore new drugs active at this site.