Immunolabeling of Central Serotonin 5‐HT1Dβ Receptors in the Rat, Mouse, and Guinea Pig with a Specific Anti‐Peptide Antiserum

: A synthetic peptide (25 amino acids) corresponding to a specific portion of the third intracytoplasmic loop of the rat serotonin 5‐HT1B/1Dβ receptor was coupled to keyhole limpet hemocyanin and injected monthly into rabbits. Anti‐peptide antibodies were detected by enzyme‐linked immunosorbent assay and characterized by immunoprecipitation of the 5‐HT1B/1Dβ receptor in CHAPS‐solubilized extracts from rat striatal membranes. Up to 60% of solubilized striatal serotonin‐O‐carboxymethylglycyl[125I]iodotyrosinamide ([125I]GTI; a selective 5‐HT1B/1D radioligand) binding sites were immunoprecipitated and subsequently pharmacologically identified as 5‐HT1B receptors. The remaining 40% of [125I]GTI binding sites were shown to be 5‐HT1D receptors. In addition, these antibodies were successfully used in immunofluorescence experiments to detect the 5‐HT1B/1Dβ, but not the 5‐HT1D/1Dα, receptor in transiently transfected LLC‐PK1 cells. Immunoautoradiographic experiments performed with brain sections from the rat, mouse, and guinea pig showed that the substantia nigra and globus pallidus contained the highest densities of 5‐HT1Dβ receptor‐like immunoreactivity. Comparison of the regional distribution of immunolabeling with that of the specific binding of [125I]GTI in the brain of these species further confirmed that the anti‐peptide antibodies selectively recognized only the 5‐HT1Dβ component of [125I]GTI specific receptor binding sites.