title: Interaction of melatonin with human lymphocytes: evidence for binding sites coupled to potentiation of cyclic AMP stimulated by vasoactive intestinal peptide and activation of cyclic GMP.
abstract: Melatonin binding sites were characterized in human blood lymphocytes. The specific binding 2-[125I]iodo-melatonin ([125I]MEL) to human lymphocytes was dependent on time and temperature, stability, saturation, and reversibility. Moreover, guanine nucleotides decreased the specific binding of [125I]MEL to crude membranes of human lymphocytes, suggesting the coupling of these binding sites to a guanosine nucleotide binding regulatory protein(s). In competition studies, the specific binding of [125I]MEL to lymphocytes was inhibited by increasing concentrations of native melatonin. Scatchard analysis showed that data were compatible with the existence of two classes of binding sites: a high-affinity site with a Kd of 5.20 +/- 0.79 nM and a binding capacity of 50.6 +/- 11.0 fmol/10(7) cells, and a low-affinity site with a Kd of 208.5 +/- 50.2 nM and a binding capacity of 2691 +/- 265 fmol/10(7) cells. However, concentration-dependent binding of [125I]MEL to lymphocytes was saturable and resulted in a linear Scatchard plot, suggesting binding to a single class of binding sites. The Kd for the single site was 1.02 +/- 0.34 nM with a binding capacity of 10.1 +/- 1.6 fmol/10(7) cells. Their affinities closely correlated with the production of cyclic nucleotides, suggesting a physiological role for the melatonin binding sites. Thus, melatonin potentiated the effect of vasoactive intestinal peptide (VIP) on cyclic AMP production (ED50 = 1.9 nM) and stimulated cyclic GMP accumulation (ED50 = 125 nM). Results demonstrate the existence of two binding sites for [125I]MEL in human blood lymphocytes, with a high-affinity binding site coupled to the potentiation of the effect of VIP on cyclic AMP production and a low-affinity binding site coupled to activation of cyclic GMP production.
authors.affiliation: Department of Medical Biochemistry and Molecular Biology, University of Seville School of Medicine, Spain.
authors.forename: M A
authors.initials: MA
authors.lastname: Lopez-Gonzalez
authors.forename: J R
authors.initials: JR
authors.lastname: Calvo
authors.forename: C
authors.initials: C
authors.lastname: Osuna
authors.forename: J M
authors.initials: JM
authors.lastname: Guerrero
mesh_terms: D001667:Binding, Competitive / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects; D000242:Cyclic AMP / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D004357:Drug Synergism; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D008550:Melatonin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D011685:Purine Nucleotides / Q000494:pharmacology; D011742:Pyrimidine Nucleotides / Q000494:pharmacology; D044094:Receptors, Melatonin; D017981:Receptors, Neurotransmitter / Q000187:drug effects / Q000378:metabolism*; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
chemical_list: D011685:Purine Nucleotides; D011742:Pyrimidine Nucleotides; D044094:Receptors, Melatonin; D017981:Receptors, Neurotransmitter; D014660:Vasoactive Intestinal Peptide; C044556:2-iodomelatonin; D000242:Cyclic AMP; D006152:Cyclic GMP; D008550:Melatonin