Expression of Dopamine Receptor mRNA in the Rat Area Postrema

Pain control during cavity preparation is very important for conservative treatment. We have reported that the dopamine D2- and D5- receptors, which are related to the analgesia , are localized in the area postrema. However it is not known whether the mRNA of these receptors is expressed in the area postrema. Objectives: The purpose of this study was to examine the expression of dopamine receptors mRNA in rat area postrema by in situ hybridization. Methods: Area postrema was obtained from adult male Sprague-Dawley rats(200-250g) undergoing brain surgery. Tissue sample was embedded in Tissue -Tek compound, then quickly frozen with Spot Freeze S( FC-302 )and stored in a freezer (-20°C) until required. For sectioning, the samples were cut at 10µm thickness using a cryostat microtome and mounted on silane-coated slide. RT-PCR product (cDNA) of each dopamine receptor mRNA from D1- to D5- receptor was cloned into the EcoR I site of pCR¨… TOPO ¨(Invitrogen) between the SP6 and T7 promoters. Non-isotopic riboprobes were prepared by in vitro transcription of linear plasmid templates using digoxigenin (DIG)-UTP and SP6 or T7 RNA polymerases (Roche). Sections were processed for in situ hybridization overnight using sense (control) and antisense DIG-labelled riboprobes. Specific hybrids were detected using anti-DIG antibody conjugated to alkaline phosphatase (AP) and the substrate NBT/BCIP (Roche). Results: Dopamine D2- and D5- receptors mRNA express in area postrema. On the other hand , the expressions of dopamine D1-,D3-,D4-receptors mRNA were not detected in this area.Conclusion:These results suggest that the pain-induced analgesia and the autonomic control mediated by D2- and D5- receptors work in the area postrema of rats.