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Characterisation of prostacyclin receptors in adult rat dorsal root ganglion cells.

Rowlands Dewi Kenneth. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 113-121). / Abstract --- p.i / Acknowledgements --- p.iii / Publications --- p.iv / Abbreviations --- p.v / Contents --- p.vii / Chapter Chapter 1 --- Prostaglandins --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Prostanoid biosynthesis and metabolism --- p.1 / Chapter 1.3 --- Prostaglandin receptors --- p.3 / Chapter 1.3.1 --- DP-receptors --- p.3 / Chapter 1.3.2 --- EP1-receptors --- p.4 / Chapter 1.3.3 --- EP2-receptors --- p.4 / Chapter 1.3.4 --- EP3-receptors --- p.5 / Chapter 1.3.5 --- EP4-receptors --- p.6 / Chapter 1.3.6 --- FP-receptors --- p.7 / Chapter 1.3.7 --- IP-receptors --- p.8 / Chapter 1.3.8 --- TP-receptors --- p.11 / Chapter 1.4 --- Agonists and antagonists --- p.11 / Chapter Chapter 2 --- Role of prostacyclin in pain modulation --- p.14 / Chapter 2.1 --- Pain --- p.14 / Chapter 2.2 --- Prostaglandins and pain --- p.15 / Chapter 2.3 --- Prostacyclin and pain --- p.16 / Chapter 2.3.1 --- [3H]-Iloprost binding sites --- p.16 / Chapter 2.3.2 --- IP-receptor mRNA --- p.17 / Chapter 2.3.3 --- IP-receptor knockout mice --- p.17 / Chapter 2.3.4 --- Direct nociceptive action of prostacyclin --- p.18 / Chapter 2.4 --- Treatment of prostanoid-induced pain --- p.19 / Chapter Chapter 3 --- Dorsal root ganglion cells --- p.21 / Chapter 3.1 --- In vitro model of pain --- p.21 / Chapter 3.2 --- Characteristics of cultured DRG cells --- p.22 / Chapter 3.2.1 --- Size and distribution --- p.22 / Chapter 3.2.2 --- Biochemical and physiological characteristics --- p.22 / Chapter 3.2.2.1 --- Gapsaicin-sensitive neurones --- p.23 / Chapter 3.2.2.2 --- Neuropeptide content --- p.23 / Chapter 3.2.2.3 --- Elevation of [Ca2+]i --- p.24 / Chapter 3.3 --- Effect of nerve growth factor --- p.24 / Chapter Chapter 4 --- Materials and solutions --- p.26 / Chapter 4.1 --- Materials --- p.26 / Chapter 4.2 --- Solutions --- p.30 / Chapter 4.2.1 --- Culture medium --- p.30 / Chapter 4.2.2 --- Buffers --- p.31 / Chapter 4.2.3 --- Solutions --- p.32 / Chapter Chapter 5 --- Development of dorsal root ganglion cell preparation --- p.33 / Chapter 5.1 --- Introduction --- p.33 / Chapter 5.2 --- Methods --- p.34 / Chapter 5.2.1 --- Dissection of dorsal root ganglia --- p.34 / Chapter 5.2.2 --- Preparation of a single-cell suspension --- p.34 / Chapter 5.2.2.1 --- Effect of trimming dorsal root ganglia --- p.34 / Chapter 5.2.2.2 --- Enzymatic dissociation --- p.35 / Chapter 5.2.2.3 --- Mechanical dissociation --- p.36 / Chapter 5.2.3 --- Neuronal cell enrichment --- p.36 / Chapter 5.2.3.1 --- Differential adhesion --- p.36 / Chapter 5.2.3.2 --- BSA gradient --- p.37 / Chapter 5.2.3.3 --- Combination of BSA gradient and differential adhesion --- p.37 / Chapter 5.2.4 --- Cell counting --- p.37 / Chapter 5.2.5 --- Culture conditions --- p.38 / Chapter 5.2.6 --- Size distribution of DRG cells --- p.39 / Chapter 5.2.7 --- Immunocytochemistry --- p.39 / Chapter 5.3 --- Results and discussion --- p.40 / Chapter 5.3.1 --- Preparation of single-cell suspension --- p.40 / Chapter 5.3.2 --- Neuronal cell enrichment --- p.42 / Chapter 5.3.3 --- Size distribution of DRG cells --- p.32 / Chapter 5.3.4 --- Effects of mitotic inhibitors and NGF --- p.45 / Chapter 5.3.5 --- Immunocytochemistry --- p.48 / Chapter 5.4 --- Conclusions --- p.48 / Chapter Chapter 6 --- Methods --- p.53 / Chapter 6.1 --- Dorsal root ganglion cell preparation --- p.53 / Chapter 6.1.1 --- Preparation of tissue culture plates and coverslips --- p.54 / Chapter 6.1.2 --- Preparation of Pasteur pipettes --- p.54 / Chapter 6.2 --- Measurement of adenylate cyclase activity --- p.55 / Chapter 6.2.1 --- Introduction --- p.55 / Chapter 6.2.2 --- Preparation of columns --- p.55 / Chapter 6.2.3 --- Measurement of [3H]-cyclic AMP production --- p.56 / Chapter 6.2.4 --- Data analysis --- p.57 / Chapter 6.3 --- Measurement of phospholipase C activity --- p.58 / Chapter 6.3.1 --- Introduction --- p.58 / Chapter 6.3.2 --- Preparation of columns --- p.58 / Chapter 6.3.3 --- Measurement of [3H]-inositol phosphate production --- p.59 / Chapter 6.3.4 --- Data analysis --- p.60 / Chapter 6.4 --- Measurement of [Ca2+]i --- p.60 / Chapter 6.4.1 --- Introduction --- p.60 / Chapter 6.4.2 --- Preparations of cells --- p.61 / Chapter 6.4.3 --- Measurement of Fura-2 fluorescence --- p.62 / Chapter 6.5 --- Measurement of neuropeptides --- p.62 / Chapter 6.5.1 --- Introduction --- p.62 / Chapter 6.5.2 --- Preparation of cells --- p.63 / Chapter 6.5.3 --- CGRP assay --- p.64 / Chapter 6.5.4 --- Substance P assay --- p.64 / Chapter 6.5.5 --- Purification of samples using Sep-Pak cartridges --- p.65 / Chapter Chapter 7 --- Characterisation of prostacyclin receptors on adult rat dorsal root ganglion cells --- p.66 / Chapter 7.1 --- Stimulation of adenylate cyclase --- p.66 / Chapter 7.1.1 --- Introduction --- p.66 / Chapter 7.1.2 --- Agonist concentration-response curves --- p.67 / Chapter 7.1.3 --- Cross-desensitisation experiments --- p.72 / Chapter 7.1.4 --- Evidence for EP3-receptors --- p.77 / Chapter 7.1.5 --- G-protein coupling of the IP-receptor --- p.77 / Chapter 7.1.6 --- Discussion --- p.78 / Chapter 7.1.7 --- Conclusions --- p.82 / Chapter 7.2 --- Stimulation of phospholipase C --- p.82 / Chapter 7.2.1 --- Introduction --- p.82 / Chapter 7.2.2 --- Agonist concentration-response curves --- p.83 / Chapter 7.2.3 --- G-protein coupling --- p.83 / Chapter 7.2.4 --- Discussion and Conclusions --- p.84 / Chapter 7.3 --- Stimulation of changes in [Ca2+]i --- p.87 / Chapter 7.3.1 --- Introduction --- p.87 / Chapter 7.3.2 --- Preliminary results --- p.87 / Chapter 7.3.3 --- Discussion and conclusions --- p.89 / Chapter Chapter 8 --- Neuropeptide release by adult rat dorsal root ganglion cells --- p.90 / Chapter 8.1 --- Introduction --- p.90 / Chapter 8.2 --- Methods and Results --- p.91 / Chapter 8.3 --- Discussion --- p.91 / Chapter 8.4 --- Conclusions --- p.92 / Chapter Chapter 9 --- Regulation of prostacyclin receptors on adult rat DRG cells --- p.93 / Chapter 9.1 --- Introduction --- p.93 / Chapter 9.2 --- Contribution of non-neuronal cells --- p.93 / Chapter 9.3 --- Effect of DRG cell density --- p.94 / Chapter 9.4 --- Effect of indomethacin --- p.99 / Chapter 9.5 --- Contribution of endogenously-produced non-prostanoid ligands --- p.100 / Chapter 9.6 --- Effect of PKC activation --- p.102 / Chapter 9.7 --- Discussion --- p.104 / Chapter 9.8 --- Conclusions --- p.106 / Chapter Chapter 10 --- General Discussion and Conclusions --- p.107 / Chapter 10.1 --- Development of DRG cell preparation --- p.107 / Chapter 10.2 --- Effect of prostanoid mimetics on intracellular messengers --- p.108 / Chapter 10.3 --- Regulation of prostacyclin receptors --- p.109 / Chapter 10.4 --- Role of prostacyclin in pain modulation --- p.111 / References --- p.113

Identiferoai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_323028
Date January 2000
ContributorsRowlands, Dewi Kenneth., Chinese University of Hong Kong Graduate School. Division of Pharmacology.
Source SetsThe Chinese University of Hong Kong
LanguageEnglish
Detected LanguageEnglish
TypeText, bibliography
Formatprint, xi, 121 leaves : ill. ; 30 cm.
RightsUse of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/)

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