Localization of P2Y1 purinoceptor transcripts in the rat penis and urinary bladder.

ABSTRACT

PURPOSE: The aim of this study was to determine the expression and localization of the P2Y1 purinoceptor mRNA in rat penis and urinary bladder using reverse transcription polymerase chain reaction (RT-PCR), northern blotting and in situ hybridization (ISH). MATERIALS AND METHODS: RT-PCR First strand cDNA was prepared from rat penis and urinary bladder dome total RNA and used for PCR with primers designed to amplify fragments of the P2Y1 purinoceptor cDNA sequence. Northern blotting PCR products were subcloned into the pGEM-5Zf(+) plasmid vector, sequenced and random primer labeled using 32p. Labeled probe was hybridized. ISH Digoxigenin labeled cRNA probes were synthesized by in vitro transcription. RESULTS: P2Y1 purinoceptor mRNA was detected by RT-PCR analysis in both rat penis and urinary bladder. RNA blotting using a P2Y1 purinoceptor cDNA probe revealed a single transcript of 4.2kb in both tissues. This band was the same size as that expressed by the heart, which contains high levels of P2Y1 purinoceptor (Burnstock, G. Physiological and pathological roles of purines an update. Drug. Dev. Res., 28 195, 1993). By ISH, P2Y1 purinoceptor mRNA was localized in detrusor smooth muscle cells and blood vessels in urinary bladder. In penis, positive signals were detected in endothelial cells which line the lacunar space and blood vessels. No hybridization was seen in corpus cavernosum smooth muscle cells and urethra. CONCLUSION: These results indicate that mRNAs for P2Y1 purinoceptor are expressed in detrusor smooth muscle cells and blood vessels of rat urinary bladder. However, in penis, this receptor is expressed in endothelial cells which lines the lacunar space and blood vessels, but not expressed in corpus cavernosum smooth muscle cells and urethra.