Aldosterone regulation of sodium channel gamma-subunit mRNA in cortical collecting duct cells.
Specific regulatory mechanisms of aldosterone-stimulated Na+ reabsorption through the apical amiloride-sensitive channel are unknown. In this study, we examined the effects of aldosterone on Na+ channel gamma-subunit mRNA levels in cultured rabbit cortical collecting duct cells. With the use of reverse transcriptase-polymerase chain reaction (RT-PCR) with RNA isolated from aldosterone-treated cells and degenerate primers, a 446-base pair (bp) PCR product was amplified and further characterized by nested PCR and sequencing. The nested PCR yielded a predicted 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 determined by quantitative PCR after a 24-h aldosterone treatment. The results demonstrate that Na+ channel gamma-subunit mRNA levels were significantly higher (2.6 +/- 0.42) in aldosterone-treated cultures vs. the controls. This increase, however, is less than the aldosterone-induced increase (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 responsible, at least in part, for the aldosterone-induced Na+ current in the kidney.
Denault
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DL
Department of Physiology, Dartmouth Medical School, Lebanon, New Hampshire 03756, USA.
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Náray-Fejes-Tóth
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D000450:Aldosterone / Q000494:pharmacology / Q000502:physiology*; D000732:Androstanols / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004553:Electric Conductivity; D004594:Electrophysiology; D007672:Kidney Cortex; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015335:Molecular Probes; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D015222:Sodium Channels / Q000235:genetics*; D014158:Transcription, Genetic
D000732:Androstanols; D015335:Molecular Probes; D012333:RNA, Messenger; D015222:Sodium Channels; D000450:Aldosterone; C049154:11,17-dihydroxy-6-methyl-17-(1-propynyl)androsta-1,4,6-triene-3-one