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Molecular charaterisation of serotonin 5-HT2 receptors
Başlık:
Molecular charaterisation of serotonin 5-HT2 receptors
Yazar:
Wu, Chun, author.
ISBN:
9780438043602
Yazar Ek Girişi:
Fiziksel Tanımlama:
1 electronic resource (303 pages)
Genel Not:
Source: Dissertation Abstracts International, Volume: 76-08C.
Özet:
Serotonin (5-hydroxytryptamine; 5-HT) mediates many functions of the central nervous system by its interaction with specific neuronal receptors. Through physiological, pharmacological and molecular cloning studies, at least 14 different subtypes of the 5-HT receptors have been identified. The localisation of these receptors in the central and the peripheral nervous system is critical for determining the functions and mechanism of action of each receptor and may have important implications for development of receptor specific psychiatric drugs. The progress in the research on 5-HT receptors has been restricted by the lack of reagents that are highly specific for each receptor. For instance, the use of ligands or certain receptor specific drugs has been limited by their reactivity with more than one type of receptor. The present research was undertaken to develop monoclonal antibodies that are highly specific for each, member of the 5-HT2 receptor family that includes 5-HT2AR, 5-HT2BR and 5-HT2CR. The 5-HT2A, 5-HT2B and 5-HT2C receptor genes were amplified from human brain and retinal libraries and cloned into baculovirus expression vectors. The expressed recombinant proteins were purified from insect cells and used to develop monoclonal antibodies to the receptors. Through extensive characterisation involving the use of recombinant proteins, cell lines that ectopically express the receptors, and various tissues, monoclonal antibodies that are highly specific for each receptor were selected. These monoclonal antibodies were subsequently used for studying the expression and distribution of each 5-HT2 receptor within mouse embryos and rat brain tissues. From the immunostaining studies, a comprehensive description of the distribution of the three receptors within these tissues was obtained. In general, there was differential staining in both mouse embryos and rat brain, varying from regions with no staining to regions of intense reactivity with antibodies to the three 5-HT2 receptors. Interestingly, studies using adult human brain tissues showed that 5-HT2AR staining was elevated in activated astrocytes in the brains of patients with Alzheimer's disease, Huntington's disease or hypertensive encephalopathy. Double staining of tissue sections confirmed the colocalisation of glial fibrillary acidic protein GFAP and 5-HT2A receptor. Whether the occurence of 5-HT2AR protein mediated activity is beneficial or deleterious requires further investigation. An RNase protection assay comprising a probe set for 5-HT2A, 5-HT2B and 5-HT2C receptors was developed to aid the simultaneous analysis and quantification of the three receptors within each RNA sample from rat tissue. Using this assay it was observed that 5-HT2CR expression was much stronger than the other two receptors, with the expression of 5-HT2BR being the weakest in adult rat brain. In general 5-HT2AR was expressed at higher levels in the cortex and striatum than in other areas of the brain, whereas the strongest expression of 5-HT2CR was in the hypothalamus. Interestingly the 5-HT2 receptors were differentially expressed during rat embryogenesis. 5-HT2AR levels gradually increased from embryonic day (ED) 11 to ED21 whereas the expression of 5-HT2BR decreased from ED9 to ED11 and then remained relatively constant through ED21. 5-HT2cR was initially expressed at residual levels between ED9 and ED12 but dramatically increased to peak levels at ED13 and then decreased by ED17. It is anticipated that the BV expression vectors, the mAbs, and the RNA probes developed in this study will be useful tools for studying the anatomical distribution, regulation and function of the serotonin receptors.
Notlar:
School code: 1543
Tüzel Kişi Ek Girişi:
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Yer Numarası | Demirbaş Numarası | Shelf Location | Lokasyon / Statüsü / İade Tarihi |
---|---|---|---|
XX(684297.1) | 684297-1001 | Proquest E-Tez Koleksiyonu | Arıyor... |
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