CHARACTERIZATION AND MAPPING OF MELATONIN RECEPTORS IN THE BRAIN OF 3 MAMMALIAN-SPECIES - RABBIT, HORSE AND SHEEP - A COMPARATIVE INVITRO BINDING STUDY

Melatonin receptors were characterized in the brains of three mammals (rabbit, horse and sheep) by an in vitro binding technique, using 2-[I-125]iodomelatonin as labelled ligand. Although binding sites for melatonin have been described recently in several vertebrate species (including the sheep), the rabbit and the horse have not been the subject of investigation so far. Apart from characterization, the present report describes receptor distribution in a number of brain regions, thus allowing for direct interspecies comparison under the same methodological conditions. 2-[I-125]iodomelatonin labelled high-affinity binding sites in crude membrane preparations from these species. A series of kinetic and saturation experiments revealed that the binding was rapid, stable, saturable, reversible, of high affinity (K(d) in the low picomolar range) and low capacity (B(max) between 1 and 20 fmol/mg protein). The competition studies showed that the relative order of potency of a variety of indoles for inhibition of 2-[I-125]iodomelatonin binding was as follows: 2-iodomelatonin > 6-chloromelatonin > melatonin > > > 5-methoxytryptophol > 5-methoxytryptamine, and that it was similar in the different brain regions. Prazosin, which has been reported as an extremely potent melatonin analog in the hamster brain, possessed no potency in all preparations from different regions in the three species under investigation. The regional distribution of the receptor showed insignificant species differences. Highest density was always recorded in the median eminence/pars tuberalis (ME/PT) area. Other regions (SCN, POA and certain cortical areas), showed lower, but significant, receptor content. Saturation and competition studies revealed that these binding sites were also of high affinity, low capacity and high specificity. These results demonstrate that melatonin receptor properties and distribution are similar in these species; the apparent receptor density, however, was highest in the sheep ME/PT. Most of the rabbit brain regions exhibited higher apparent B(max) values, compared to the corresponding ones in the other two species.