title: High affinity binding of [3H]propionyl-[Met(O2)11]substance P(7-11), a tritiated septide-like peptide, in Chinese hamster ovary cells expressing human neurokinin-1 receptors and in rat submandibular glands.
abstract: Propionyl-[Met(O2)11]substance P(7-11) [ALIE-124 or propionyl-[Met(O2)11]SP(7-11)] has been designed as a septide-like ligand adequate for tritiation and, therefore, adequate for binding studies. In Chinese hamster ovary (CHO) cells expressing human tachykinin neurokinin (NK)-1 receptors, ALIE-124 displaced [3H][Pro9]substance P (SP) from its binding site at micromolar concentrations. However, ALIE-124 stimulated phosphatidylinositol hydrolysis, as previously shown for septide-like peptides. With [3H]ALIE-124 (95 Ci/mmol), we have been able to reveal a high affinity binding site in CHO cells (Kd = 6.6 +/- 1.0 nM), with a low maximal binding capacity. [3H]ALIE-124 specific maximal binding represented only 15-20% of that observed with [3H][Pro9]SP in CHO cells. Septide-like peptides, including septide and NKA, were potent competitors (in the nanomolar range) of [3H]ALIE-124 specific binding site. Interestingly, SP and [Pro9]SP were also potent competitors, with 10-fold greater potency for sites labeled with [3H]ALIE-124 than for sites labeled with [3H][Pro9]SP. The NK-1 antagonist RP 67580 also showed a higher potency for [3H]ALIE-124 than for [3H][Pro9]SP-specific binding sites. NKB and [Lys5,methyl-Leu9,Nle10]NKA(4-10) displaced [3H]ALIE-124 binding but with lower potency, whereas senktide had no affinity. The existence of [3H]ALIE-124 specific binding sites was also demonstrated in rat submandibular gland. In this tissue, [3H]ALIE-124 specific maximal binding was higher, reaching 40-50% of that achieved with [3H][Pro9]SP.
authors.affiliation: Laboratoire de Chimie Organique Biologique, URA CNRS 493, Université Pierre et Marie Curie, Paris, France.
authors.forename: S
authors.initials: S
authors.lastname: Sagan
authors.forename: J C
authors.initials: JC
authors.lastname: Beaujouan
authors.forename: Y
authors.initials: Y
authors.lastname: Torrens
authors.forename: M
authors.initials: M
authors.lastname: Saffroy
authors.forename: G
authors.initials: G
authors.lastname: Chassaing
authors.forename: J
authors.initials: J
authors.lastname: Glowinski
authors.forename: S
authors.initials: S
authors.lastname: Lavielle
mesh_terms: D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D008297:Male; D010446:Peptide Fragments / Q000378:metabolism*; D011761:Pyrrolidonecarboxylic Acid / Q000031:analogs & derivatives; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D018040:Receptors, Neurokinin-1 / Q000378:metabolism*; D013363:Submandibular Gland / Q000378:metabolism*; D013373:Substance P / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D018040:Receptors, Neurokinin-1; C107105:propionyl-(Met(O2)11)substance P(7-11); D013373:Substance P; C054725:septide; D011761:Pyrrolidonecarboxylic Acid