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Molecular cloning and characterisation of GABA-B receptors from Xenopus laevis.

James, R.; (2008) Molecular cloning and characterisation of GABA-B receptors from Xenopus laevis. Doctoral thesis , University of London. Green open access

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Gamma-amino butyric acid (GABA) is the principal inhibitory neurotransmitter in the adult central nervous system (CNS) and signals via ionotropic GABAA receptors and metabotropic GABAB receptors. GABAB receptors are obligate heterodimers comprised of GABAB(i) and GABAB(2) subunits, members of the Family 3 G-protein coupled receptors (GPCRs). GABABL is an orphan Family 3 GPCR of unknown function, most closely related to GABAB receptors. To investigate the functions of these genes during vertebrate development, Xenopus laevis GABAB(1), GABAB(2) and GABABL cDNAs were isolated, and their spatiotemporal expression patterns during embryogenesis analysed by RT-PCR and in situ hybridization. Maternal GABAB(2) transcripts were detected by RT-PCR in blastulae, whereas GABAB(1) and GABABL transcripts were not detected until gastrulation and neurulation respectively. In situ hybridization revealed that GABAB(i{rcub} and GABAB{lcub}2) transcripts were co-expressed in most brain regions, although areas of unique GABAB(i{rcub} expression also existed, and GABABL transcripts were located primarily in the brain and otic vesicle of the tailbud embryo. Co-expression of GABAB(1) and GABAB(2) transcripts suggests a role for metabotropic GABA receptor signalling in the developing brain of Xenopus embryos. However, overexpression of GABAB(1) and GABAB(2) transcripts together or in isolation, during embryonic development did not generate a distinct morphological phenotype. In contrast, embryos overexpressing GABABL during embryonic development exhibited a significant body truncation phenotype. Animal cap assays indicated that GABABL overexpression interferes with mesodermal convergent extension, whilst RT-PCR shows that the expression of mesoderm-specific markers is not affected, demonstrating morphogenetic but not biochemical activity of GABABL. Whilst the temporal expression pattern of GABABL does not support an endogenous role in the regulation of convergent extension in Xenopus , these experiments demonstrate that GABABL is a functional protein that acts in a manner reminiscent of a GPCR by disrupting intracellular signalling cascades.

Type: Thesis (Doctoral)
Title: Molecular cloning and characterisation of GABA-B receptors from Xenopus laevis.
Identifier: PQ ETD:591606
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest. Third party copyright material has been removed from the ethesis
UCL classification: UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Structural and Molecular Biology
URI: https://discovery.ucl.ac.uk/id/eprint/1444304
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