Angiotensin II Type 2 Receptor and Receptor Mas Are Colocalized and Functionally Interdependent in Obese Zucker Rat Kidney

The actions of angiotensin II type 2 receptor (AT 2 R) and the receptor Mas (MasR) are complex but show similar pronatriuretic function; particularly, AT 2 R expression and natriuretic function are enhanced in obese/diabetic rat kidney. In light of some reports suggesting a potential positive interaction between these receptors, we tested hypothesis that renal AT 2 R and MasR physically interact and are interdependent to stimulate cell signaling and promote natriuresis in obese rats. We found that infusion of AT 2 R agonist C21 in obese Zucker rats (OZR) increased urine flow and urinary Na excretion which were attenuated by simultaneous infusion of the AT 2 R antagonist PD123319 or the MasR antagonist A-779. Similarly, infusion of MasR agonist Ang-(1–7) in OZR increased urine flow and urinary Na excretion, which were attenuated by simultaneous infusion of A-779 or PD123319. Experiment in isolated renal proximal tubules of OZR revealed that both the agonists C21 and Ang-(1–7) stimulated NO which was blocked by either of the receptor antagonists. Dual labeling of AT 2 R and MasR in OZR kidney sections and human proximal tubule epithelial cells showed that AT 2 R and MasR are colocalized. The AT 2 R also coimmunoprecipitated with MasR in cortical homogenate of OZR. Immunoblotting of cortical homogenate cross-linked with zero-length oxidative (sulfhydryl groups) cross-linker cupric-phenanthroline revealed a shift of AT 2 R and MasR bands upward with overlapping migration for their complexes which were sensitive to the reducing β-mercaptoethanol, suggesting involvement of -SH groups in cross-linking. Collectively, the study reveals that AT 2 R and MasR are colocalized and functionally interdependent in terms of stimulating NO and promoting diuretic/natriuretic response.