Dopamine D1 and D5 receptors differentially regulate oxidative stress through paraoxonase 2 in kidney cells

Background. The renal dopaminergic system plays an important role in the pathogenesis of hypertension. Dopamine D-1-like receptors (D1R and D5R) decrease reactive oxygen species (ROS) production via inhibition of pro-oxidant enzymes such as NADPH oxidase. Paraoxonase 2 (PON2) is also involved in the inhibition of NADPH oxidase activity. Therefore, we tested the hypothesis that D1R and D5R inhibit ROS production by increasing the expression of PON2, including those in membrane microdomains. Methods and results. PON2 colocalized with D1R and D5R in mouse renal proximal tubules (RPTs), human RPT (hRPT) cells, and HEK293 cells heterologously expressing human D1R (HEK-hD(1)R ) or D5R (HEK-hD(5)R). Fenoldopam, an agonist for both D1R and D5R, increased PON2 co-immunoprecipitation with D1R and D5R in HEK-hD(1)R and HEK-hD(5)R cells, respectively. Silencing PON2 increased ROS production and NADPH oxidase activity, and impaired the inhibitory effect of fenoldopam. Fenoldopam increased PON2 protein in both lipid rafts (LRs) and non-LRs in HEK-hD(1)R cells, but only in non-LRs in HEK-hD(5)R and hRPT cells. Long-term (hrs) fenoldopam stimulation increased PON2 protein in a time-dependent manner in HEK-hD(5)R, but not in HEK-hD(1)R cells. Because the effects of fenoldopam on non-LR and total PON2 expressions were similar in HEK-hD(5)R and hRPT cells, additional studies were performed to determine the relationship between D5R and PON2. Renal PON2 protein was decreased in D-5(- /-) mice. In hRPT cells, silencing D5R decreased PON2 expression and increased ROS production. Conclusions. We conclude that D-1-like receptors inhibit ROS production by altering PON2 distribution in membrane microdomains in the short-term, and by increasing PON2 expression in the long-term.