FUNCTIONAL EXPRESSION OF HUMAN D-3 DOPAMINE-RECEPTORS IN DIFFERENTIATED NEUROBLASTOMA-X-GLIOMA NG108-15 CELLS

This study describes the depression of calcium currents caused by acti vation of human D-3 dopamine receptors which have been stably expresse d in the neuroblastoma x glioma NG108-15 cell line. Transfected cells, which had been differentiated with prostaglandin E(1) and isobutylmet hylxanthine, exclusively expressed D-3 receptor mRNA, which was demons trated by reverse transcription polymerase chain reaction techniques. Transfected cells had high affinity binding sites for iodosulpiride, w ith a K-d of 0.8 nM and receptor density of 240 fmol mg(-1) protein. C alcium currents were recorded using nystatin-perforated patch clamp te chniques. In contrast to untransfected cells that had been differentia ted, high-threshold calcium currents in differentiated hD(3)-NG108-15 cells were depressed by application of dopamine and quinpirole. These responses were abolished by the dopamine receptor antagonist S-(-)sulp iride (1 mu M), demonstrating;that they were caused by the activation of the transfected dopamine receptors. Coupling of human D-3 receptors to calcium currents was sensitive to the action of pertussis toxin, s uggesting the involvement of G-proteins of the G(i) and/or G(o) subtyp e. These results demonstrate that human D-3 receptors represent a func tional class of dopamine receptor.