Galanin receptor 1 gene expression is regulated by cyclic AMP through a CREB-dependent mechanism

The galanin receptor-1 (GalR1) protein belongs to a family of G protein-cou pled receptors for the neuropeptide galanin (GalR1, GalR2 and GalR3) distri buted throughout the central and peripheral nervous system. Activation of g alanin receptors by their ligands results in increased feeding, impaired le arning, enhanced opiate analgesia and decreased opiate place preference. We have shown that opiate withdrawal; which is known to increase levels of cA MP in the locus coeruleus (LC), results in an increase in the number of gal anin binding sites and the level of GalR1 mRNA in the LC. We have isolated a 3.6-kb fragment 5' of the inititiation codon of the mouse GalR1 gene and generated a series of deletion mutations of this fragment driving expressio n of luciferase for use in transient transfection assays in PC12 and Cath.a cell lines. Treatment with forskolin, but not dideoxyforskolin, up-regulat es GalR1 transcription, likely through elevation of cAMP levels. The region between - 1050 and - 700 base pairs upstream of exon one is necessary both for basal activity of the GalR1 promoter and for forskolin-mediated increa ses in transcription. The forskolin effect can be blocked by simultaneous m utation of a CRE-like site and a CRE/DRE-like site, but not mutation of eit her site alone. Gel shift and super-shift experiments demonstrate that the transcription factor CREB can bind to both sites and is likely to be respon sible for the cAMP-mediated increase in GalR1 promoter activity. This study provides a molecular mechanism for the increased GalR1 expression in the L C seen following opiate withdrawal.