Detection of dopamine D2 receptor mRNA and binding sites in monkey amniotic epithelial cells.
Previous results from our laboratory showed that monkey amniotic epithelial cells (MAEC) possess the catecholamine synthesizing enzymes and have the capacity to synthesize and release CA. Recently, we also reported that these cells express dopamine D1 receptor mRNA and binding sites. This study was designed to investigate the presence of dopamine D2 receptors in MAEC. Using RT-PCR, we found that MAEC express dopamine D2 receptor mRNA that is having 98% homology with human dopamine D2 receptors. Radioligand saturation binding studies showed a 3H-YM-09151-2 high-affinity binding site with a K(D) of 0.293+/-0.06 nM and Bmax of 180.69+/-11.61 fmol/mg protein. Competition experiments with a variety of displacing drugs demonstrated that D2 antagonists potently compete with 3H-YM-09151-2 binding, whereas D1 antagonists displayed a weaker competition for the binding sites. The rank order of potency of these compounds in competing with 3H-YM-09151-2 for binding sites was consistent with the pharmacology of the dopamine D2 receptors. All competition curves were better fitted to a one-site model with a Hill coefficient around unity, indicating that 3H-YM-09151-2 is labeling a single population of receptors. These results provide, for the first time, a compelling evidence that MAEC natively express dopamine D2 receptor mRNA and binding sites, and they suggest that monkey amniotic epithelial cells (MAEC) could represent a source of primate dopamine receptors without the need for transformation or cloning procedures using nonprimate cells, as generally happens.
Elwan
M A
MA
Department of Inherited Metabolic Diseases, National Institute of Neuroscience, NCNP, Kodaira, Tokyo, Japan.
Ishii
T
T
Sakuragawa
N
N
D000650:Amnion / Q000166:cytology* / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001549:Benzamides / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D018492:Dopamine Antagonists / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D005260:Female; D006801:Humans; D007700:Kinetics; D008252:Macaca fascicularis; D008969:Molecular Sequence Data; D010920:Placenta / Q000166:cytology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D017448:Receptors, Dopamine D2 / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
D001549:Benzamides; D018492:Dopamine Antagonists; D012333:RNA, Messenger; D017448:Receptors, Dopamine D2; C030265:nemonapride