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Characteristics of 2‐[ 125 I]iodomelatonin binding sites in the pigeon spleen and modulation of binding by guanine nucleotides
Author(s) -
Poon A.M.S.,
Wang X.L.,
Pang S.F.
Publication year - 1993
Publication title -
journal of pineal research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.881
H-Index - 131
eISSN - 1600-079X
pISSN - 0742-3098
DOI - 10.1111/j.1600-079x.1993.tb00499.x
Subject(s) - gtp' , dissociation constant , radioligand , melatonin , guanosine , binding site , biology , medicine , receptor , endocrinology , g protein , radioligand assay , nucleotide , guanine , stereochemistry , biophysics , chemistry , biochemistry , enzyme , gene
2‐[ 125 ]Iodomelatonin binding sites in membrane preparations of pigeon spleen have been characterized. The binding was stable, saturable, reversible, and of high affinity. Rosenthal and Hill analyses showed that the radioligand‐receptor interaction involved a single class of binding sites. Analysis of the binding results of spleens collected during mid‐light revealed an equilibrium dissociation constant (Kd) of 36.6 ± 4.8 pmol/1 (mean ± sem, n = 10) and a maximum density (Bmax) of 2.3 ± 0.2 fmol/mg protein. There was no significant difference in the Kd (46.9 ± 5.0 pmol/1) or the Bmax values (2.4 ± 0.3 fmol/mg protein) for spleens collected during mid‐dark (n = 9), although the mid‐dark serum and pineal melatonin levels were significantly higher ( P < 0.05) than the corresponding mid‐light values. Kinetic analysis showed a Kd of 8.6 ± 2.0 pmol/1 (n ± 4), in agreement with that derived from the saturation studies. Except for inhibition by 2‐iodomelatonin, melatonin, 6‐chloromelatonin, 6‐hydroxymelatonin and N‐acetylserotonin, the other indoles or neurotransmitters tested have little inhibition on the binding. In addition, guanosine 5′‐O‐(3‐thiophosphate) (GTPγS), a nonhydrolysable analog of GTP, was found to inhibit the binding in a dose‐dependent manner. Saturation studies revealed that this is due to a decrease in both the affinity and density of the binding sites. These data suggest that a single type of melatonin receptor is found in the pigeon spleen and that the site is coupled to a guinine nucleotide binding protein (G‐protein). Our findings support a direct pineal melatonin action on the immune system.