The cooperative roles of the dopamine receptors, D(1)R and D(5)R, on the regulation of renal sodium transport

Determining the individual roles of the two dopamine D(1)-like receptors (D(1)R and D(5)R) on sodium transport in the human renal proximal tubule has been complicated by their structural and functional similarity. Here we used a novel D(5)R-selective antagonist (LE-PM436) and D(1)R or D(5)R-specific gene silencing to determine second messenger coupling pathways and heterologous receptor interaction between the two receptors. D(1)R and D(5)R co-localized in renal proximal tubule cells and physically interact, as determined by co-immunoprecipitation and FRET microscopy. Stimulation of renal proximal tubule cells with fenoldopam (D(1)R/D(5)R agonist) led to both adenylyl cyclase and phospholipase C (PLC) activation using real-time FRET biosensors ICUE3 and CYPHR, respectively. Fenoldopam increased cAMP accumulation and PLC activity and inhibited both NHE3 and NaKATPase activities. LE-PM436 and D(5)R siRNA blocked the fenoldopam-stimulated PLC pathway but not cAMP accumulation, while D(1)R siRNA blocked both fenoldopam-stimulated cAMP accumulation and PLC signaling. Either D(1)R or D(5)R siRNA, or LE-PM436 blocked the fenoldopam dependent inhibition of sodium transport. Further studies using the cAMP-selective D(1)R/D(5)R agonist SKF83822 and PLC-selective D(1)R/D(5)R agonist SKF83959 confirmed the cooperative influence of the two pathways on sodium transport. Thus, D(1)R and D(5)R interact in the inhibition of NHE3 and NaKATPase activity, the D(1)R primarily by cAMP, while the D(1)R/D(5)R heteromer modulates the D(1)R effect through a PLC pathway.