Two cDNAs encoding novel isoforms of Xenopus laevis melatonin receptors were cloned using PCR primers specific for the X. laevis-melanophore Mel1c melatonin receptor described in a recent publication. The novel isoforms were highly homologous to the described frog Mel1c cDNA, although the C-terminal tail of both was shorter by 65 amino acid residues. Nucleotide sequences of these novel isoforms, called Mel1c(alpha) and Mel1c(beta), differed from each other by only 35 nucleotides and six amino acid residues. Studies on several animals of various Xenopus species indicate that Mel1c(alpha) and Mel1c(beta) receptors may correspond to allelic variants of the same locus. Studies on cells transfected with both receptor cDNAs showed the expression of high-affinity 2-[125I]iodomelatonin binding sites. Agonist stimulation of Mel1c(alpha) receptor was associated with the inhibition of cAMP accumulation stimulated by forskolin (IC50 approximately 10(-10) M) in HeLa, Ltk-, and human embryonic kidney 293 (HEK 293) cells. Mel1c(beta) receptor modulated cAMP in HeLa and HEK 293 cells but not in Ltk- cells. Both receptors inhibited, in a dose-dependent manner, cGMP accumulation in all three cell lines incubated with a phosphodiesterase inhibitor. This effect was localized upstream of soluble guanylyl cyclase and was blocked by pertussis toxin treatment. However, IC50 values (approximately 10(-10) M for Mel1c(beta) and 10(-9) to 10(-7) M for Mel1c(alpha)) and maximal inhibition levels showed that Mel1c(alpha) receptors are much less efficiently coupled to the cGMP pathway. Coupling differences may be explained by the fact that five of the six amino acid substitutions between Mel1c(alpha) and Mel1c(beta) receptors are located within cytoplasmic regions potentially involved in signal transduction. The existence of coupling differences is in agreement with the observation that expression of both receptors is evolutionally conserved in native tissue. In conclusion, two novel, potentially allelic, isoforms of Xenopus Mel1c melatonin receptors display identical ligand-binding characteristics, but different potencies in modulating cAMP and cGMP levels through G(i)/G(o)-dependent pathways. Furthermore, to our knowledge, this study provides the first data on the modulation of intracellular cGMP levels by cloned melatonin receptors.