Intracellular Sodium Modulates the Expression of Angiotensin II Subtype 2 Receptor in PC12W Cells

Abstract

Abstract —Although the angiotensin II subtype 2 receptor (AT 2 -R) is expressed abundantly in the adrenal medulla, its physiological significance has not yet been determined. To obtain fundamental knowledge of the regulation of AT 2 -R expression in the adrenal medulla, we investigated the effects of modulating several ion channels on AT 2 -R expression in PC12W cells. Experiments were performed after 24 hours of serum depletion under subconfluent conditions. After 48 hours of treatment with various agonists or antagonists, the receptor density and mRNA level of AT 2 -Rs were quantified by 125 I-[Sar 1 ,Ile 8 ]angiotensin II binding and Northern blot analysis. Ouabain (10 to 100 nmol/L) and insulin (10 to 100 nmol/L) dose-dependently increased receptor density and mRNA level. Analysis of the binding characteristics revealed that the ouabain-dependent increase in AT 2 -R levels was due to an increase in binding capacity without a change in the K d value. These increases were blocked by lowering the Na + concentration in the medium. A low concentration of the sodium ionophore monensin (10 nmol/L), the K + -channel blocker quinidine (10 μmol/L), and the ATP-sensitive K + -channel blockers tolbutamide (100 μmol/L) and glybenclamide (10 μmol/L) also significantly increased receptor density, but the ATP-sensitive K + -channel agonist cromakalim (100 μmol/L) decreased receptor density significantly ( P <0.01). Nifedipine (10 μmol/L) decreased basal receptor density and completely blocked the increase in receptor density caused by these agents. The increase in receptor density caused by an increase in intracellular Na + was accompanied by an increase in mRNA level, whereas the ATP-sensitive K + -channel blockers did not change mRNA level. Nifedipine slightly decreased mRNA level. These results suggest that AT 2 -R expression is sensitively regulated by intracellular cation levels. The change in intracellular Na + level transcriptionally regulates AT 2 -R expression, whereas the K + -channel blocker–dependent upregulation appears to be at least in part posttranslational.