Adenosine A1Receptor mRNA in Microdissected Rat Nephron Segments

Abstract

AbstractAdenosine plays several roles in the kidney mediated by the specific receptors A1, A2, and possibly A3. We studied the localization of adenosine A1receptor mRNA in rat nephron segments using reverse transcription and polymerase chain reaction (RT-PCR). The nephron segments of male Sprague-Dawley rats (6 to 8 weeks old) were microdissected. Total RNA was prepared by the acid-guanidinium–phenol–chloroform method and used in the following RT-PCR assay. Because the PCR primers spanned no intron, samples reacted in the absence of RT were used as controls for amplification of genomic DNA. The PCR products were size-fractionated by electrophoresis, visualized with ethidium bromide staining, and confirmed by Southern blot analysis. PCR products were detected in all of the nephron segments examined. No signals were detected in samples reacted in the absence of RT. Strong signals were detected in glomeruli, medullary collecting duct, cortical thick ascending limb, and medullary thick ascending limb, while weak signals were found in proximal convoluted and straight tubules. Previously, the presence of A1receptors has been demonstrated in glomeruli, collecting duct, and thick ascending limb in the rat kidney by autoradiography and binding studies. In addition to these segments, we further detected A1receptor mRNA in proximal convoluted and straight tubules. Thus, A1receptor mRNA seems to be broadly expressed along the nephron.