Molecular cloning and pharmacological characterization of rat melatonin MT1 and MT2 receptors.

In order to interpret the effects of melatonin ligands in rats, we need to determine their activity at the receptor subtype level in the corresponding species. Thus, the rat melatonin rMT1 receptor was cloned using DNA fragments for exon 1 and 2 amplified from rat genomic DNA followed by screening of a rat genomic library for the full length exon sequences. The rat rMT2 receptor subtype was cloned in a similar manner with the exception of exon 1 which was identified by screening a rat genomic library with exon 1 of the human hMT2 receptor. The coding region of these receptors translates proteins of 353 and 364 amino acids, respectively, for rMT1 and rMT2. A 55% homology was observed between both rat isoforms. The entire contiguous rat MT1 and MT2 receptor coding sequences were cloned, stably expressed in CHO cells and characterized in binding assay using 2-[125I]-Iodomelatonin. The dissociation constants (Kd) for rMT1 and rMT2 were 42 and 130 pM, respectively. Chemically diverse compounds previously characterized at human MT1 and MT2 receptors were evaluated at rMT1 and rMT2 receptors, for their binding affinity and functionality in [35S]-GTPγS binding assay. Some, but not all, compounds shared a similar binding affinity and functionality at both rat and human corresponding subtypes. A different pharmacological profile of the MT1 subtype has also been observed previously between human and ovine species. These in vitro results obtained with the rat melatonin receptors are thus of importance to understand the physiological roles of each subtype in animal models.