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Screening and isolation of a natural dopamine D₁ receptor antagonist using cell-based assays

Yu, Sungryul, Park, Jeong Soo, Paredes, Verenice, Song, Myoung-Chong, Baek, Nam-In, Lee, Sang-Ik, Lim, Jong-Soon, Cho, Nam-Young, Yoon, Jaeseung, Baek, Kwanghee
Journal of biotechnology 2010 v.145 no.3 pp. 304-309
Chinese hamsters, Gleditsia sinensis, agonists, animal ovaries, antagonists, biotechnology, dopamine, dose response, fractionation, humans, medicinal plants, plant extracts, screening
To develop a cell-based assay to screen for human dopamine D₁ receptor agonists or antagonists from medicinal plant extracts, a stable Chinese hamster ovary (CHO) cell line (CHO-D1R) expressing the human dopamine D₁ receptor was established using an expression vector containing a scaffold attachment region (SAR) element. CHO-D1R cells showed specific binding to [³H]-SCH23390 with high affinity (K d =1.47±0.17nM) and dose-dependent responses for the dopamine-mediated stimulation of cAMP concentrations (EC₅₀ =20.6±1.44nM). The screening of medicinal plant extracts using cell-based cAMP assays revealed that an extract of Gleditsia sinensis Lam., which is known to be rich in saponin, had strong antagonist activity for the D₁ receptor. From the activity-guided fractionation and chemical structural analysis of the G. sinensis extract, a compound called gleditsioside F was isolated and was identified to have antagonist activity for the D₁ receptor. Gleditsioside F showed very effective D₁ antagonist activity by inhibiting ligand binding to the D₁ receptor as well as by inhibiting dopamine-mediated increases in cAMP concentration.