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P Denis, V Fardin, J P Nordmann, P P Elena, L Laroche, H Saraux, W Rostene; Localization and characterization of substance P binding sites in rat and rabbit eyes.. Invest. Ophthalmol. Vis. Sci. 1991;32(6):1894-1902.
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Specific and high-affinity binding sites for Substance P (SP) were found in eyes from albino rabbits and rats using an in vitro autoradiographic method with 125I-Bolton Hunter SP (BHSP). autoradiograms were generated by apposing 10-20 microns-thick cryostat eye sections to 3H-Hyperfilm or liquid emulsion and quantified by means of image-analysis procedures. Kinetic studies showed that equilibrium was reached after a 75-min incubation at room temperature. In rat retina, specific binding corresponding to approximately 90% of total binding, was reversible, of high affinity (dissociation constant [Kd], 0.13 +/- 0.02 nM). Half-time for dissociation of 125I-BHSP was about 15 min. Unlabeled SP and the two neurokinins (NK) A and B competed in a concentration-dependent manner for retinal sites labeled by 125I-BHSP with the following order of potencies: SP greater than NKA greater than NKB, in agreement with a pharmacologic profile of a SP receptor site. In both species, specific binding was found in the iris sphincter muscle, choroid, and retina. In rats, detectable amounts of SP-binding sites were also expressed in the corneal epithelium and iridial stroma. Quantitative analysis of the autoradiograms revealed that the highest densities of 125I-BHSP binding sites were localized in the iris sphincter muscle in rabbits and the inner retina in rats.
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