ALDOSTERONE REGULATION OF SODIUM-CHANNEL GAMMA-SUBUNIT MESSENGER-RNA IN CORTICAL COLLECTING DUCT CELLS

Specific regulatory mechanisms of aldosterone-stimulated Na+ reabsorpt ion through the apical amiloride-sensitive channel are unknown. In thi s study, we examined the effects of aldosterone on Na+ channel gamma-s ubunit mRNA levels in cultured rabbit cortical collecting duct cells. With the use of reverse transcriptase-polymerase chain reaction (RT-PC R) with RNA isolated from aldosterone-treated cells and degenerate pri mers, a 446-base pair (bp) PCR product was amplified and further chara cterized by nested PCR and sequencing. The nested PCR yielded a predic ted 164-bp product. Sequencing of the 446-bp PCR product revealed 83% nucleotide and 91% amino acid identity to the rat colonic Na+ channel gamma-subunit. The relative abundance of Na+ channel mRNA was determin ed by quantitative PCR after a 24-h aldosterone treatment. The results demonstrate that Na+ channel gamma-subunit mRNA levels were significa ntly higher (2.6 +/- 0.42) in aldosterone-treated cultures vs. the con trols. This increase, however, is less than the aldosterone-induced in crease (3.2 +/- 2.0) in the amiloride-sensitive short-circuit current. These results indicate that Na+ channel gamma-subunit mRNA levels are increased by aldosterone and that this increase is likely to be respo nsible, at least in part, for the aldosterone-induced Na+ current in t he kidney.