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Title: The generation and analysis of transgenic mice over-expressing the 5-hydroxytryptamine (5-HT) transporter
Author: Loder, Merewyn Katharine
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2005
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To explore the regulation and function of the serotonin transporter (5-HTT), transgenic (Tg) mice that over express the human 5-HTT were made. A 500Kb modified yeast artificial chromosome &YAC35D8) containing the human 5-HTT gene and its 5’ regulatory sequence was injected into fertilized mouse oocytes, and Tg offspring were identified by PCR. Analysis by in situ hybridisation showed that h5-HTT mRNA is overexpressed in the cortex, hippocampus and mesencephalic raphe nuclei of these animals. RNase protection and RT-PCR showed that the human 5-HTT is expressed in these regions at levels comparable to, or greater than, that of the endogenous 5-HTT whilst the mouse 5-HTT RNA’ remained unaltered. [3H]Citalopram, a high affinity 5-HTT ligand was used to measure the protein expression. [3H]-citalopram binding showed a higher KD value (P<0.02) and a 2.3 – 3.5 fold increase in binding site density (P < 0.01) in cortical membranes from Tg mice compared to controls. In addition, the Hill coefficient was less than 1 (consistent with the presence of two binding sites corresponding to the human and mouse 5-HTT). Using HPLC, tissue extracts from the brainstem, hippocampus, midbrain, hypothalamus cerebellum and basal ganglia were analysed for 5-HT, DOPAC and 5H1AA. The transgenic mice show a drop in 5-HT levels in all areas except the cerebellum (-17 to – 40%) and a slight increase in 5H1AA levels (6-17%); though this is only significant in the cortex (42% p<0.05). Thus the 5H1AA to 5-HT ratio is decreased by 27-76% in transgenic animals (p<0.005).
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available