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Title: The pharmacology, signalling and expression of the lipid-sensing receptor GPR92
Author: Dorning, Ashley J.
ISNI:       0000 0004 2745 9700
Awarding Body: University of Aberdeen
Current Institution: University of Aberdeen
Date of Award: 2013
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G protein-coupled receptors (GPCRs) are 7 transmembrane domain proteins capable of initiating cellular responses following ligand binding. GPR92 is expressed in the central nervous system (CNS) and is demonstrated to be involved in pain signalling via neurons of the spinal cord and dorsal root ganglion (DRG). Activated by the endogenous lipid lysophosphatidic acid (LPA), GPR92 is now considered the 5th LPA receptor. There remains controversy regarding GPR92 pharmacology however, as studies show another endogenous lipid, farnesyl pyrophosphate (FPP), also activates GPR92 with similar potency and efficacy to LPA. LPA-induced activation of GPR92 results in increased intracellular calcium (Ca2+) and cAMP. Furthermore, GPR92 mediates neurite retraction via Rho kinase and activates the cAMP responsive element-binding (CREB) protein. This transcription factor is important in synaptic plasticity and regulates the expression of many neuronal genes including brain derived neurotrophic factor (BDNF), yet the role of GPR92 in the brain has not been explored. Here, I describe the pharmacology and signalling of GPR92, with preliminary data describing its expression. I utilise a cell line lacking endogenous LPA responsiveness (B103 rat neuroblastoma) in which I stably express GPR92. Using intracellular Ca2+ changes and CREB phosphorylation as read-outs, I find FPP to be the most potent and efficacious ligand. The similarly structured lipid geranylgeranyl pyrophosphate (GGPP) produced comparable results to FPP. GPR92-mediated CREB phosphorylation has been described in a mouse model of pain. The ligands which induce this response however, have not been assessed. I describe for the first time, ligand-induced CREB phosphorylation via GPR92, and found that FPP causes the most robust CREB response. LPA and GGPP also induced GPR92-mediated CREB phsphorylation. Furthermore, I find that GPR92 mediates a novel Gq-dependent, Ca2+ independent, signalling pathway resulting in CREB phosphorylation. In addition, I examine GPR92 expression in the CNS using reverse-transcriptase polymerase chain reaction (RT-PCR) and in situ hybridisation. GPR92 is expressed throughout the brain, with particularly high expression in 7 the brain stem, DRG, and hippocampus. In situ hybridisation revealed a distinct expression pattern confirming high levels of GPR92 mRNA in the hippocampal region. GPR92 mRNA is also observed in the cortex, habenula, thalamus and hypothalamus. I also report the expression of the FPP synthesising enzyme FPP synthase (FPPS). Relative expression levels of FPPS between CNS regions are similar to GPR92. Preliminary data suggests FPP is capable of mobilising Ca2+ in hippocampal neuronal and non-neuronal cells. These data suggest an important role for GPR92 in the brain where it, and the enzyme involved in generating one of its endogenous ligands, are highly expressed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Cell receptors