No abstract

Melanin-concentrating hormone (MCH) is involved in the regulation of feeding and energy homeostasis. Recently, a 353-amino acid splice variant form of the human orphan receptor SLC-1 (1) (hereafter referred to as MCH 1 ) was identified as an MCH receptor. This report describes the cloning and functional characterization of a novel second human MCH receptor, which we designate MCH 2 , initially identified in a genomic survey sequence as being homologous to MCH 1 receptors. Using this sequence, a full-length cDNA was generated with an open reading frame of 1023 base pairs, encoding a polypeptide of 340 amino acids, with 38% identity to MCH 1 and with many of the structural features conserved in G protein-coupled receptors. This newly discovered receptor belongs to class 1 (rhodopsin-like) of the G protein-coupled receptor superfamily. HEK293 cells transfected with MCH 2 receptors responded to nanomolar concentrations of MCH with an increase in intracellular Ca 2؉ levels and increased cellular extrusion of protons. In addition, fluorescently labeled MCH bound with nanomolar affinity to these cells. The tissue localization of MCH 2 receptor mRNA, as determined by quantitative reverse transcription-polymerase chain reaction, was similar to that of MCH 1 in that both receptors are expressed predominantly in the brain. The discovery of a novel MCH receptor represents a new potential drug target and will allow the further elucidation of MCH-mediated responses. Melanin-concentrating hormone (MCH)1 is a cyclic neuropeptide that was first discovered in teleost fish, in which it acts as a skin color-regulating hormone (2). In rodents its tissue distribution in the perikarya of the lateral hypothalamus and the zona incerta suggests that MCH may be involved in a variety of behavioral responses (3). Similar tissue distributions have been reported in both bird (4) and monkey (5). Reports implicating MCH in the regulation of feeding behavior show that increased food intake occurs after direct administration of MCH into the brain (6) and that MCH is up-regulated after fasting and in obese leptin-deficient mice (7). There are also reports that suggest MCH may be involved in aggressive behavior, anxiety, and reproductive function (8, 9).Recently, several groups independently identified a 353-amino acid splice variant of the orphan G protein-coupled receptor (GPCR) SLC-1 (1) as an MCH receptor (10 -14). In view of the findings of the current study, we propose that this form of SLC-1 be hereafter referred to as MCH 1 . Southern blot and related studies have indicated the absence of additional MCH receptor subtypes that closely resemble MCH 1 at the DNA level (3, 10, 15). However, because degeneracy in receptor-ligand pairings throughout the GPCR superfamily is common, we reasoned that other MCH receptors with low homology to the MCH 1 receptor may exist. This suggestion is supported by reports of pharmacological differences between the MCH 1 receptor and MCH binding sites in various cell lines and tissues (10).Sequencing of the human...

No abstract

Despite only modest homology between h5-HT1B and h5-HT1D receptor amino acid sequences, these receptors display a remarkably similar pharmacology. To date there are few compounds which discriminate between these receptor subtypes and those with some degree of selectivity, such as ketanserin, have greater affinity for other 5-HT receptor subtypes. We now report on two compounds, SB-216641 (N-[3-(2-dimethylamino) ethoxy-4-methoxyphenyl]-2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)-(1,1'-biphenyl)-4-carboxamide) and BRL-15572 3-[4-(3-chlorophenyl) piperazin-1-yl]-1,1-diphenyl-2-propanol), which display high affinity and selectivity for h5-HT1B and h5-HT1D receptors, respectively. In receptor binding studies on human receptors expressed in CHO cells, SB-216641 has high affinity (pKi = 9.0) for h5-HT1B receptors and has 25-fold lower affinity at h5-HT1D receptors. In contrast, BRL-15572 has 60-fold higher affinity for h5-HT1D (pKi = 7.9) than 5-HT1B receptors. Similar affinities for these compounds were determined on native tissue 5-HT1B receptors in guinea-pig striatum. Functional activities of SB-216641 and BRL-15572 were measured in a [35S]GTPgammaS binding assay and in a cAMP accumulation assay on recombinant h5-HT1B and h5-HT1D receptors. Both compounds were partial agonists in these high receptor expression systems, with potencies and selectivities which correlated with their receptor binding affinities. In the cAMP accumulation assay, results from pK(B) measurements on the compounds again correlated with receptor binding affinities (SB-216641, pK(B) = 9.3 and 7.3; BRL-15572, pK(B) = <6 and 7.1, for h5-HT1B and h5-HT1D receptors respectively). These compounds will be useful pharmacological agents to characterise 5-HT1B and 5-HT1D receptor mediated responses.

No abstract