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Title: In vitro and in vivo radioligand binding studies for the N-methyl-D-aspartate receptor and the 5-hydroxytryptamine transporter
Author: Shirakawa, Kiyoharu
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1996
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The thesis contains in vitro and in vivo radioligand binding studies for the N-methyl-D-aspartate (NMDA) receptor, the 5-HT1A receptor and the 5-HT transporter. A novel NMDA antagonist FR115427 ((+)-1-methyl-1-phenyl-1,2,3,4-tetrahydroisoquinoline hydrochloride; (+)FR) was characterised using [3H]MK-801 binding to rat brain membranes. As with (+)MK-801, the affinity of (+)FR was increased when 10 μM L-glutamate was added in the assay buffer. (+)FR inhibited [3H]MK-801 binding to rat cortical synaptosomal membranes in the presence of 10 μM L-glutamate with a Ki value of 40 nM although (+)FR was 12-fold less potent than (+)MK-801. These results indicate that (+)FR is a non-competitive NMDA antagonist. [3H](+)FR synthesised by tritiation of (+)Cl-FR (mono-chlorinated aromatic precursor of (+)FR) was successfully purified by an open column method. In vivo distribution of intravenous injection of [3H](+)FR in rats was analysed. Intravenous injection of [3H](+)FR resulted in a rapid accumulation of radioactivity in brain, and the brain regional distribution of radioactivity appeared to reflect the localisation of NMDA receptors. The possibility of using derivatives of citalopram (selective 5-HT uptake inhibitor) as a photoaffinity or SPET ligand for the 5-HT transporter was evaluated using 5-azido-citalopram (5-AC) (photoaffinity ligand) and 5-iodo-citalopram (5-IC) (SPET ligand). 5-AC had high affinity (Ki = 1.65 nM) for [3H]citalopram binding sites being only 1.8-fold less potent than citalopram itself. In the presence of 5-AC, repeated U.V. irradiation (15W) of rat cortical membranes produced a significant 20% reduction in the Bmax value for [3H]citalopram binding site. Therefore radiolabelled 5-AC may provide a tool for isolation and characterisation of the 5-HT transporter.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available