This grant application presents the cloning of a candidate for the human imidiazolene receptor (IR) cDNA and gene. The brainstem IR has been thought to function in the central neuronal modulation of blood pressure by clonidine, but the probable role of IR in other brain region is currently under investigation. The investigator's laboratory has demonstrated consistent and robust elevations in platelet IR radioligand binding in five earlier studies of depressed patients, followed by a normalization of radioligand binding after chronic treatments with earlier desipramine or fluoxetine. The investigator and other investigators also observed alterations in IR in depressed suicide victims brain tissue. Hence it is of interest to identify the signaling mechanisms of IR and the manner in which IR are normally regulated, or possibly dysregulated in depression. The following studies are proposed in order to characterize a cloned IR-like cDNA: a)The first aim is to establish whether the investigator's IR- like cDNA encodes the pharmacological properties expected of the IR, subtype of imidzoline receptors. Preliminary transfection data have indicated that the expressed protein possesses high affinity for IR1>IR2 ligands. b) To test the investigator's hypothesis that IR are coupled to a phosphatidylcholine-selective phospholipase C (PC-PLC) signal transduction pathway, it will be determined whether transfected IR-like cDN will express ligand-specific coupling to the PC-PLC, and other second messenger pathways. c) To determine the regionalization of brain IR (and to test whether an endogenous IR candidate neurotransmitter, agmatine, shares a similar distribution in rats and humans), the regional distribution of IR-like mRNA will also be determined by situ hybridization. d) A final aim of this grant will be to identify possible regulatory DNA sequences in the upstream gene promoter region of the human and rat IR gene. These studies will clarify the molecular nature of the IR anit's signal transduction pathways. This basic grant is thematically linked to a clinical grant (Halaris, PI) which has also been submitted for consideration. The clinical grant will explore whether brain IR1 protein, IR1 mRNA and agmatine are elevated in brains of depressed suicide victims, whether imidazoline receptors underlie a blunted growth hormone response to clonidine as observed in depressed patients, and whether platelet IR1 levels and/or plasma agmatine concentrations are sensitive to the severity of depression. The findings from the clinical grant will best be understood within the context of these studies in the basic grant, and vice versa.
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