RT-PCR microlocalization of mRNA for guanylyl cyclase-coupled ANF receptor in rat kidney.
Microlocalization of mRNA coding for the guanylyl cyclase-coupled atrial natriuretic factor (ANF) receptor was carried out in the rat kidney. We used a combination of reverse transcription and polymerase chain reaction (RT-PCR) in individual microdissected renal tubule segments, glomeruli, and vasa recta bundles. Relative quantitation of the resulting amplified cDNA utilized densitometry of autoradiograms from Southern blots probed with a specific 32P-labeled probe. Among renal tubule segments, the largest signal was found in the terminal inner medullary collecting duct (IMCD). Slightly smaller signals were found in the initial IMCD and in loop of Henle segments from the inner medulla. Readily detectable signals were also seen in the following segments (in descending order): cortical collecting duct, proximal convoluted tubule, medullary thick ascending limb, cortical thick ascending limb, distal convoluted tubule, and outer medullary collecting duct. Large signals were also detected in glomeruli and in vasa recta bundles from the inner stripe of the outer medulla. Based on these results, we conclude that 1) renal microlocalization of specific mRNAs coding for hormone receptors is feasible through application of the RT-PCR procedure in microdissected renal tubules and vascular elements, and 2) the gene for the guanylyl cyclase-coupled ANF receptor is broadly expressed along the nephron, raising the possibility that multiple sites of ANF action are present.
Terada
Y
Y
Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, Clinical Neuroscience Branch, National Institutes of Health, Bethesda, Maryland 20892.
Moriyama
T
T
Martin
B M
BM
Knepper
M A
MA
Garcia-Perez
A
A
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D006162:Guanylate Cyclase / Q000378:metabolism*; D007668:Kidney / Q000737:chemistry*; D007679:Kidney Medulla / Q000737:chemistry; D007685:Kidney Tubules, Collecting / Q000737:chemistry; D008138:Loop of Henle / Q000737:chemistry; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D017461:Receptors, Atrial Natriuretic Factor; D011956:Receptors, Cell Surface / Q000235:genetics*; D014018:Tissue Distribution; D014158:Transcription, Genetic
D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D012194:RNA-Directed DNA Polymerase; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor