Fig. 2.

Colocalization of D₁-like dopamine receptors and α_1A-ARs in hRPTCs and human and mouse kidneys. A: hRPTCs were grown on coverslips, serum starved for 1 h, and treated with Fen (1 μM) at the indicated time points. Cells were then fixed, double immunostained for D₁Rs or D₅Rs (pseudocolored green) and α_1A-ARs (pseudocolored red), and visualized via confocal microscopy. Around 30–40 cells were scored, and 5–10 cells were imaged per experiment. The lipid rafts of the plasma membrane were visualized using cholera toxin subunit B (CTxB) conjugated with Alexa fluor 647, and nuclei were visualized using 4′,6-diamidino-2-phenylindole (DAPI). Colocalization between D₁-like dopamine receptors and α_1A-ARs is denoted by yellow punctate areas, whereas colocalization of these receptors with the plasma membrane (PM) is denoted by white in the merged images. B: human kidney sections were antigen retrieved and double immunostained for D₁Rs or D₅Rs (pseudocolored red) and α_1A-ARs (pseudocolored green). Proximal tubules were visualized using brush border-localized CD15 (pseudocolored blue) as a marker. Distal convoluted tubules are indicated with white arrows. DIC, differential interference contrast. C: C57Bl/6J mice were treated with Fen (2 μg·kg⁻¹·min⁻¹ iv) or Veh for 30 min, and kidneys were collected and prepared for immunofluorescence to evaluate the distribution and colocalization of D₁Rs or D₅Rs (pseudocolored red) and α_1A-ARs (pseudocolored green). Proximal tubules were visualized using CD15 (pseudocolored blue) as a marker. The localization of D₅Rs at the brush border is indicated by the white arrows. Magnification: ×600. Scale bars = 10 μm. n = 3.