%0 Journal Article %1 Raymond1989 %A Raymond, J. R. %A Fargin, A. %A Lohse, M. J. %A Regan, J. W. %A Senogles, S. E. %A Lefkowitz, R. J. %A Caron, M. G. %D 1989 %J Mol Pharmacol %K & 8-Hydroxy-2-(di-n-propylamino)tetralin Affinity Azides/*metabolism Cell D2 Dopamine Dopamine/analysis Humans Iodine Labels/*metabolism Line Precipitin Radioisotopes/diagnostic Serotonin/*analysis/immunology Spiperone/analogs Tests Tetrahydronaphthalenes/metabolism derivatives/*metabolism use Receptor %N 1 %P 15-21 %T Identification of the ligand-binding subunit of the human 5-hydroxytryptamine1A receptor with N-(p-azido-m-125I iodophenethyl)spiperone, a high affinity radioiodinated photoaffinity probe %U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=2526292 %V 36 %X The ligand-binding subunit of the human 5-hydroxytryptamine1A (5-HT1A) receptor transiently expressed in COS-7 cells and of the native human 5-HT1A receptor derived from hippocampus and frontal cortex were identified by photoaffinity labeling with N-(p-azido-m-125Iiodophenethyl)spiperone ( 125IN3-NAPS), previously characterized as a high affinity radioiodinated D2-dopamine receptor probe. The identity of the ligand-binding subunit was confirmed by immunoprecipitation with an antipeptide rabbit antiserum, JWR21, raised against a synthetic peptide derived from the predicted amino acid sequence of the putative third intracellular loop of the human 5-HT1A receptor. In transiently transfected COS-7 cells expressing 14 +/- 3 pmol/mg of protein human 5-HT1A receptors, a single broad 75-kDa band was photoaffinity labeled by 125IN3-NAPS. This band displayed the expected pharmacology of the 5-HT1A receptor, as evidenced by the ability of a series of competing ligands to block 125IN3-NAPS photoincorporation. Moreover, antiserum JWR21 specifically and quantitatively immunoprecipitated the 75-kDa photoaffinity-labeled band from a soluble extract of the transfected COS-7 cell membranes, further confirming its identity. Finally, utilizing a combination of photoaffinity labeling and immunoprecipitation, the native ligand-binding subunit of 62-64 kDa was identified in human hippocampus and frontal cortex. The availability of the high specific activity, high affinity, photoaffinity ligand 125IN3-NAPS and of a potent immunoprecipitating antiserum (JWR21) should greatly facilitate the biochemical characterization of the human 5-HT1A receptor.

@article{Raymond1989, abstract = {The ligand-binding subunit of the human 5-hydroxytryptamine1A (5-HT1A) receptor transiently expressed in COS-7 cells and of the native human 5-HT1A receptor derived from hippocampus and frontal cortex were identified by photoaffinity labeling with N-(p-azido-m-[125I]iodophenethyl)spiperone [( 125I]N3-NAPS), previously characterized as a high affinity radioiodinated D2-dopamine receptor probe. The identity of the ligand-binding subunit was confirmed by immunoprecipitation with an antipeptide rabbit antiserum, JWR21, raised against a synthetic peptide derived from the predicted amino acid sequence of the putative third intracellular loop of the human 5-HT1A receptor. In transiently transfected COS-7 cells expressing 14 +/- 3 pmol/mg of protein human 5-HT1A receptors, a single broad 75-kDa band was photoaffinity labeled by [125I]N3-NAPS. This band displayed the expected pharmacology of the 5-HT1A receptor, as evidenced by the ability of a series of competing ligands to block [125I]N3-NAPS photoincorporation. Moreover, antiserum JWR21 specifically and quantitatively immunoprecipitated the 75-kDa photoaffinity-labeled band from a soluble extract of the transfected COS-7 cell membranes, further confirming its identity. Finally, utilizing a combination of photoaffinity labeling and immunoprecipitation, the native ligand-binding subunit of 62-64 kDa was identified in human hippocampus and frontal cortex. The availability of the high specific activity, high affinity, photoaffinity ligand [125I]N3-NAPS and of a potent immunoprecipitating antiserum (JWR21) should greatly facilitate the biochemical characterization of the human 5-HT1A receptor.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {Raymond, J. R. and Fargin, A. and Lohse, M. J. and Regan, J. W. and Senogles, S. E. and Lefkowitz, R. J. and Caron, M. G.}, biburl = {https://www.bibsonomy.org/bibtex/2067a2bbf52b10a3c7980784ad97798d9/pharmawuerz}, endnotereftype = {Journal Article}, interhash = {d3f2d5265a89117bb67d17f66b750cbf}, intrahash = {067a2bbf52b10a3c7980784ad97798d9}, issn = {0026-895X (Print) 0026-895X (Linking)}, journal = {Mol Pharmacol}, keywords = {& 8-Hydroxy-2-(di-n-propylamino)tetralin Affinity Azides/*metabolism Cell D2 Dopamine Dopamine/analysis Humans Iodine Labels/*metabolism Line Precipitin Radioisotopes/diagnostic Serotonin/*analysis/immunology Spiperone/analogs Tests Tetrahydronaphthalenes/metabolism derivatives/*metabolism use Receptor}, month = Jul, note = {Raymond, J R Fargin, A Lohse, M J Regan, J W Senogles, S E Lefkowitz, R J Caron, M G HL-16037/HL/NHLBI NIH HHS/United States NS-19576/NS/NINDS NIH HHS/United States S07-RR-05405/RR/NCRR NIH HHS/United States Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. United states Molecular pharmacology Mol Pharmacol. 1989 Jul;36(1):15-21.}, number = 1, pages = {15-21}, shorttitle = {Identification of the ligand-binding subunit of the human 5-hydroxytryptamine1A receptor with N-(p-azido-m-[125I] iodophenethyl)spiperone, a high affinity radioiodinated photoaffinity probe}, timestamp = {2010-12-14T18:20:08.000+0100}, title = {Identification of the ligand-binding subunit of the human 5-hydroxytryptamine1A receptor with N-(p-azido-m-[125I] iodophenethyl)spiperone, a high affinity radioiodinated photoaffinity probe}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=2526292}, volume = 36, year = 1989 }