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Abstract:The apelin peptide was recently discovered and demonstrated to be the endogenous ligand for the G protein-coupled receptor, APJ. A search of the GenBank databases retrieved a rat expressed sequence tag partially encoding the preproapelin sequence. The GenBank search also revealed a human sequence on chromosome Xq25-26.1, containing the gene encoding preproapelin. We have used the rat sequence to screen a rat brain cDNA library to obtain a cDNA encoding the full-length open reading frame of rat preproapelin. This cDNA encoded a protein of 77 amino acids, sharing an identity of 82% with human preproapelin. Northern and in situ hybridization analyses revealed both human and rat apelin and APJ to be expressed in the brain and periphery. Both sequence and mRNA expression distribution analyses revealed similarities between apelin and angiotensin II, suggesting they that share related physiological roles. A synthetic apelin peptide was injected intravenously into male Wistar rats, resulting in immediate lowering of both systolic and diastolic blood pressure, which persisted for several minutes. Intraperitoneal apelin injections induced an increase in drinking behavior within the first 30 min after injection, with a return to baseline within 1 h.

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We report the isolation of a cDNA clone named GPR54, which encodes a novel G protein-coupled receptor (GPCR). A PCR search of rat brain cDNA retrieved a clone partially encoding a GPCR. In a library screening this clone was used to isolate a cDNA with an open reading frame (ORF) encoding a receptor of 396 amino acids long which shared significant identities in the transmembrane regions with rat galanin receptors GalR1 (45%), GalR3 (45%) and GalR2 (44%). Northern blot and in situ hybridization analyses revealed that GPR54 is expressed in brain regions (pons, midbrain, thalamus, hypothalamus, hippocampus, amygdala, cortex, frontal cortex, and striatum) as well as peripheral regions (liver and intestine). In COS cell expression of GPR54 no specific binding was observed for 125 I-galanin. A recent BLAST search with the rat GPR54 ORF nucleotide sequence recovered the human orthologue of GPR54 in a 3.5 Mb contig localized to chromosome 19p13.3.z 1999 Federation of European Biochemical Societies.

Although the perturbation of either the dopaminergic system or brain-derived neurotrophic factor (BDNF) levels has been linked to important neurological and neuropsychiatric disorders, there is no known signaling pathway linking these two major players. We found that the exclusive stimulation of the dopamine D1-D2 receptor heteromer, which we identified in striatal neurons and adult rat brain by using confocal FRET, led to the activation of a signaling cascade that links dopamine signaling to BDNF production and neuronal growth through a cascade of four steps: (i) mobilization of intracellular calcium through Gq, phospholipase C, and inositol trisphosphate, (ii) rapid activation of cytosolic and nuclear calcium/calmodulin-dependent kinase II␣, (iii) increased BDNF expression, and (iv) accelerated morphological maturation and differentiation of striatal neurons, marked by increased microtubule-associated protein 2 production. These effects, although robust in striatal neurons from D5 ؊/؊ mice, were absent in neurons from D1 ؊/؊ mice. We also demonstrated that this signaling cascade was activated in adult rat brain, although with regional specificity, being largely limited to the nucleus accumbens. This dopaminergic pathway regulating neuronal growth and maturation through BDNF may have considerable significance in disorders such as drug addiction, schizophrenia, and depression.brain-derived neurotrophic factor activation ͉ calcium signaling pathway ͉ calcium/calmodulin-dependent kinase II ͉ neuronal maturation ͉ GPCR oligomerization D opamine promotes neuronal differentiation, maintenance, and survival (1-4) by modulating the transcription of different genes. Little however, is known regarding the molecular events that govern these dopamine-mediated effects. Evidence has emerged indicating a positive relationship between functions mediated by dopamine and brain-derived neurotrophic factor (BDNF) and its receptor TrkB (2-7). However, a direct mechanism bridging dopamine signaling to BDNF has not yet been described. Classically, dopamine exerts its actions through D1-like (D1, D5) and D2-like (D2, D3, D4) receptors, which regulate activation or inhibition of cAMP accumulation, through Gs/olf or Gi/o proteins, respectively (8). Other signaling cascades have also been reported (9, 10), including phosphatidylinositol turnover in brain through D1-like receptor activation (11,12), but no such activation was observed when the cloned D1 receptor was expressed (13-15). These observations led us to the discovery of the dopamine D1-D2 receptor heterooligomer, which is able to mobilize intracellular calcium (15-18). However, the signaling cascade and the physiological functions of the dopamine D1-D2 receptor heteromer in brain are unknown.Because calcium is involved in the activation of BDNF signaling (19), we hypothesized that this pathway may be central to dopamine activation of BDNF and subsequent neuronal maturation and differentiation.In this context, we describe a signaling pathway that links dopamine action through the...