Use of 2-[125I]iodomelatonin to characterize melatonin binding sites in chicken retina.
2-[125I]Iodomelatonin binds with high affinity to a site possessing the pharmacological characteristics of a melatonin receptor in chicken retinal membranes. The specific binding of 2-[125I]iodomelatonin is stable, saturable, and reversible. Saturation experiments indicated that 2-[125I]iodomelatonin labeled a single class of sites with an affinity constant (Kd) of 434 +/- 56 pM and a total number of binding sites (Bmax) of 74.0 +/- 13.6 fmol/mg of protein. The affinity constant obtained from kinetic analysis was in close agreement with that obtained in saturation experiments. Competition experiments showed a monophasic reduction of 2-[125I]iodomelatonin binding with a pharmacological order of indole amine affinities characteristic of a melatonin receptor: 2-iodomelatonin greater than 6-chloromelatonin greater than or equal to melatonin greater than or equal to 6,7-dichloro-2-methylmelatonin greater than 6-hydroxymelatonin greater than or equal to 6-methoxymelatonin much greater than N-acetyltryptamine greater than N-acetyl-5-hydroxytryptamine greater than 5-methoxytryptamine greater than 5-hydroxytryptamine (inactive). The affinities of these melatonin analogs in competing for 2-[125I]iodomelatonin binding sites were correlated closely with their potencies for inhibition of the calcium-dependent release of [3H]dopamine from chicken and rabbit retinas, indicating association of the binding site with a functional response regulated by melatonin. The results indicate that 2-[125I]iodomelatonin is a selective, high-affinity radioligand for the identification and characterization of melatonin receptor sites.
D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D002645:Chickens; D004298:Dopamine / Q000378:metabolism; D007457:Iodine Radioisotopes; D007700:Kinetics; D008550:Melatonin / Q000378:metabolism*; D044094:Receptors, Melatonin; D017981:Receptors, Neurotransmitter / Q000378:metabolism*; D012160:Retina / Q000378:metabolism*
D007457:Iodine Radioisotopes; D044094:Receptors, Melatonin; D017981:Receptors, Neurotransmitter; D008550:Melatonin; D004298:Dopamine