Kam Yiu-wing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 200-215). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.iii / Publications --- p.iv / Abbreviations --- p.v / Contents --- p.vii / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- G protein-coupled receptors --- p.1 / Chapter 1.1.1 --- Introduction --- p.1 / Chapter 1.1.2 --- G protein-coupled receptors --- p.2 / Chapter 1.1.3 --- Heterotrimeric G-proteins --- p.4 / Chapter 1.1.4 --- Second messenger systems --- p.5 / Chapter 1.1.5 --- Mechanism of GPCR activation --- p.6 / Chapter 1.1.6 --- GPCR cross talk --- p.8 / Chapter 1.2 --- Receptor theory --- p.10 / Chapter 1.2.1 --- Introduction --- p.10 / Chapter 1.2.2 --- Two-state model --- p.11 / Chapter 1.2.3 --- Three-state model --- p.12 / Chapter 1.2.4 --- Extended ternary complex model --- p.12 / Chapter 1.2.5 --- Multiple receptor state --- p.13 / Chapter 1.2.6 --- Constitutively active mutant receptors --- p.14 / Chapter 1.3 --- Agonist trafficking --- p.15 / Chapter 1.3.1 --- Introduction --- p.15 / Chapter 1.3.2 --- Effect of agonist efficacy on receptor coupling --- p.17 / Chapter 1.3.3 --- Effect of receptor expression level on receptor coupling --- p.17 / Chapter 1.3.4 --- Receptor promiscuity --- p.18 / Chapter 1.3.5 --- Agonist-induced conformational changes --- p.19 / Chapter 1.3.5.1 --- Conformational induction --- p.19 / Chapter 1.3.5.2 --- Conformational selection --- p.19 / Chapter 1.3.6 --- Receptor/G-protein system --- p.20 / Chapter 1.3.7 --- Implication of agonist trafficking --- p.21 / Chapter 1.4 --- Receptor switching --- p.22 / Chapter 1.4.1 --- Introduction --- p.22 / Chapter 1.4.2 --- Receptor switching --- p.23 / Chapter Chapter 2 --- Prostacyclin receptors --- p.34 / Chapter 2.1 --- General properties of prostacyclin --- p.34 / Chapter 2.1.1 --- Synthesis of prostacyclin --- p.34 / Chapter 2.1.2 --- Prostacyclin and its mimetics --- p.34 / Chapter 2.1.3 --- Characterization of IP-receptors --- p.36 / Chapter 2.1.3.1 --- Classification of IP-receptors --- p.36 / Chapter 2.1.3.2 --- Distribution of IP-receptors in the peripheral system --- p.36 / Chapter 2.1.3.3 --- Distribution of IP-receptors in the central nervous system --- p.38 / Chapter 2.1.3.4 --- Structure of IP-receptors --- p.39 / Chapter 2.1.4 --- Anti-thrombotic role of prostacyclin --- p.40 / Chapter 2.1.5 --- Cytoprotective role of prostacyclin --- p.41 / Chapter 2.1.6 --- Role of prostacyclin in adipogenesis --- p.42 / Chapter 2.2 --- Coupling capacity of IP-receptors --- p.43 / Chapter 2.2.1 --- Introduction --- p.43 / Chapter 2.2.2 --- Interaction with Gs --- p.44 / Chapter 2.2.3 --- Interaction with Gi --- p.45 / Chapter 2.2.4 --- Interaction with Gq --- p.45 / Chapter 2.2.5 --- Interaction with PPARs --- p.43 / Chapter 2.2.6 --- IP-receptor isoprenylation --- p.49 / Chapter 2.3 --- Regulation of IP-receptors --- p.50 / Chapter 2.3.1 --- Mechanism of signal termination --- p.50 / Chapter 2.3.2 --- Desensitization of IP-receptors --- p.51 / Chapter 2.3.3 --- Internalization of IP-receptors --- p.53 / Chapter Chapter 3 --- Materials and solutions --- p.59 / Chapter 3.1 --- Materials --- p.59 / Chapter 3.2 --- "Culture media, buffers and solutions" --- p.53 / Chapter 3.2.1 --- Culture media --- p.63 / Chapter 3.2.2 --- Buffers --- p.64 / Chapter 3.2.3 --- Solutions --- p.65 / Chapter Chapter 4 --- Methods --- p.67 / Chapter 4.1 --- Cell culture --- p.67 / Chapter 4.2 --- Mammalian cell transfection --- p.68 / Chapter 4.2.1 --- Preparation of plasmid DNA --- p.68 / Chapter 4.2.2 --- Transient transfection of mammalian cells --- p.71 / Chapter 4.2.3 --- β-galactosidase assay --- p.73 / Chapter 4.2.3.1 --- Introduction --- p.73 / Chapter 4.2.3.2 --- Preparation of cell lysate --- p.73 / Chapter 4.2.3.3 --- Micro β-galactosidase assay --- p.73 / Chapter 4.3 --- Measurement of adenylate cyclase activity --- p.74 / Chapter 4.3.1 --- [3H]-adenine prelabelling method --- p.74 / Chapter 4.3.1.1 --- Preparation of columns --- p.74 / Chapter 4.3.1.2 --- Incubation of cells --- p.75 / Chapter 4.3.1.3 --- Measurement of [3H]-cyclic AMP production --- p.75 / Chapter 4.3.1.4 --- Data analysis --- p.76 / Chapter 4.3.2 --- cAMP [125I]-scintillation proximity assay (SPA) --- p.77 / Chapter 4.3.2.1 --- Introduction --- p.77 / Chapter 4.3.2.2 --- Cell lysis method --- p.77 / Chapter 4.3.2.3 --- cAMP [I25I]-scintillation proximity assay --- p.78 / Chapter 4.4 --- Measurement of phospholipase C activity --- p.78 / Chapter 4.4.1 --- Introduction --- p.78 / Chapter 4.4.1.1 --- Preparation of columns --- p.78 / Chapter 4.4.1.2 --- [3H]-inositol phosphate assay --- p.79 / Chapter 4.4.1.3 --- Measurement of [3H]-inositol phosphate production --- p.79 / Chapter 4.4.1.4 --- Data analysis --- p.80 / Chapter 4.4.2 --- "D-myo-Inositol 1,4,5-trisphosphate (IP3) [3H] assay system" --- p.81 / Chapter 4.4.2.1 --- Introduction --- p.81 / Chapter 4.4.2.2 --- Sample preparation --- p.81 / Chapter 4.4.2.3 --- "D-myo-Inositol 1,4,5-trisphosphate (IP3) [3H] assay system" --- p.82 / Chapter 4.5 --- Measurement of changes in intracellular Ca2+ concentration --- p.82 / Chapter 4.5.1 --- Introduction --- p.82 / Chapter 4.5.2 --- Cell preparation --- p.83 / Chapter 4.5.3 --- Measurement of Fura-2 fluorescence --- p.83 / Chapter 4.6 --- Radioligand binding --- p.84 / Chapter 4.6.1 --- Introduction --- p.84 / Chapter 4.6.2 --- [3H]-iloprost ligand binding --- p.85 / Chapter 4.6.3 --- Data analysis --- p.86 / Chapter 4.7 --- Cytotoxicity test using trypan blue exclusion test --- p.86 / Chapter Chapter 5 --- Multiple coupling capacity of prostacyclin receptors in CHO cells --- p.88 / Chapter 5.1 --- Introduction --- p.88 / Chapter 5.2 --- Method --- p.88 / Chapter 5.3 --- Results and Discussion --- p.89 / Chapter 5.3.1 --- IP agonist log concentration-response curves for [3H]-cAMP and [3H]-inositol phosphate production in mIP-CHO cells --- p.89 / Chapter 5.3.2 --- Effect of varying Gαs cDNA concentration on cicaprost-stimulated [3H]-cAMP and [3H]-inositol phosphate production in mlP-CHO cells --- p.90 / Chapter 5.3.3 --- Effect of varying Gαq cDNA concentration on cicaprost-stimulated [3H]-cAMP and [3H]-inositol phosphate production in mlP-CHO cells --- p.92 / Chapter 5.4 --- Conclusion --- p.95 / Chapter Chapter 6 --- Multiple coupling capacity of prostacyclin receptors in neuroblastoma cells --- p.113 / Chapter 6.1 --- Introduction --- p.113 / Chapter 6.2 --- Method --- p.114 / Chapter 6.3 --- Results and Discussion --- p.114 / Chapter 6.3.1 --- Characterization of prostanoid receptors in SK-N-SH cells --- p.114 / Chapter 6.3.2 --- Property of IP-receptor signaling in SK-N-SH cells --- p.116 / Chapter 6.3.3 --- Effect of Gαq subunits on PLC activation in SK-N-SH cells --- p.117 / Chapter 6.3.4 --- Coupling capacity of IP-receptorin rat/mouse neuroblastoma (NG108-15) cells --- p.119 / Chapter 6.4 --- Conclusion --- p.123 / Chapter Chapter 7 --- Agonist trafficking --- p.133 / Chapter 7.1 --- Introduction --- p.133 / Chapter 7.2 --- Method --- p.134 / Chapter 7.3 --- Results and Discussion --- p.134 / Chapter 7.3.1 --- Simultaneous measurement of AC and PLC activation in metabolically-labelled mIP-CHO cells --- p.134 / Chapter 7.3.1.1 --- Effect of IBMX on PLC activation --- p.135 / Chapter 7.3.1.2 --- Effect of Li+ ion on AC activation --- p.135 / Chapter 7.3.1.3 --- Separation of [3H]-adenine and [3H]-inositol using column chromatography method --- p.136 / Chapter 7.3.2 --- Measurement of AC and PLC activation using different assay systems --- p.137 / Chapter 7.3.2.1 --- cAMP 125I-Scintillation Proximity Assay System --- p.137 / Chapter 7.3.2.2 --- "D-myo-Inositol 1,4,5-trisphosphate (IP3) [3H]-assay system" --- p.138 / Chapter 7.4 --- Conclusion --- p.139 / Chapter Chapter 8 --- Regulation of prostacyclin receptor coupling --- p.147 / Chapter 8.1 --- Introduction --- p.147 / Chapter 8.2 --- Methods --- p.149 / Chapter 8.3 --- Results and Discussion --- p.150 / Chapter 8.3.1 --- Role of protein kinases on IP-receptor activity --- p.150 / Chapter 8.3.2 --- Effect of SQ22536 on IP-receptor activity --- p.155 / Chapter 8.3.3 --- Role of PKA phosphorylation site in mIP-receptor activity --- p.156 / Chapter 8.4 --- Conclusion --- p.157 / Chapter Chapter 9 --- Prostacyclin receptor isoprenylation --- p.171 / Chapter 9.1 --- Introduction --- p.171 / Chapter 9.2 --- Method --- p.172 / Chapter 9.3 --- Results and Discussion --- p.173 / Chapter 9.4 --- Conclusion --- p.175 / Chapter Chapter 10 --- IP/DP chimeric receptors --- p.178 / Chapter 10.1 --- Introduction --- p.178 / Chapter 10.2 --- Method --- p.179 / Chapter 10.3 --- Results and Discussion --- p.180 / Chapter 10.3.1 --- Property of IP/DP chimeric receptors --- p.180 / Chapter 10.3.2 --- "ID1 chimeric receptor mutant receptors (M4, M5, M6)" --- p.182 / Chapter 10.3.3 --- "Mutant mIP-receptors (Ml, M2, M3)" --- p.183 / Chapter 10.3.4 --- Comparison between M5 and ID1 receptors --- p.184 / Chapter 10.4 --- Conclusion --- p.184 / Chapter Chapter 11 --- General Discussion and Conclusions --- p.193 / Chapter 11.1 --- Introduction --- p.193 / Chapter 11.2 --- Multiple coupling capacity of prostacyclin receptors --- p.193 / Chapter 11.3 --- Factors influencing prostacyclin receptor coupling --- p.196 / Chapter 11.4 --- Prostacyclin receptor cross talk and regulation --- p.198 / References --- p.200

Thesis (Ph.D.)--Hong Kong University of Science and Technology, 2006. / Includes bibliographical references (leaves 215-232). Also available in electronic version.

Two experiments were conducted to determine whether in ewes uterine concentrations of estrogen and progesterone receptors are affected by the presence of a conceptus or by the hormonal milieu associated with extremes in photoperiod to which ewes are exposed. In Exp.1, nine mature ewes were unilaterally ovariectomized by removing an ovary bearing the corpus luteum (CL). The ipsilateral uterine horn was ligated at the external bifurcation and a portion of the anterior ipsilateral uterine horn was removed and assayed for endometrial nuclear and cytosolic concentrations of estrogen receptor (ER) and progesterone receptor (PR) by exchange assays. After a recovery estrous cycle, ewes were bred to a fertile ram. On day 18 of gestation a 10 ml jugular blood sample was collected for measurement of serum concentrations of estradiol -17β (E₂) and progesterone by radioimmunoassay. Ewes were relaparotomized on day 18 and the remaining uterine tissue was removed. Endometrium from both the pregnant and nonpregnant uterine horn was assayed for nuclear and cytosolic ER and PR concentrations. Nuclear and cytosolic ER concentrations on day 10 of the cycle were greater than in endometrium of gravid and nongravid uterine horns on day 18 of gestation (p<.01). Endometrial nuclear PR levels were also greater on day 10 of the cycle than in the pregnant (p<.05) and nonpregnant horn (p<.01) on day 18 of gestation. There were no differences in nuclear and cytosolic ER and PR concentrations between the pregnant and nonpregnant uterine horn on day 18. Serum levels of E₂ and progesterone on day 18 of gestation were 16.56 ± 2.43 pg/ml and 1.74 ± 0.57 ng/ml, respectively. These data suggest that duration of exposure of the uterus to progesterone and(or) the presence of the conceptus causes a reduction in uterine concentrations of ER and PR. Further, an effect of the conceptus, if any, is exerted via a systemic route. In Exp. 2, ten mature ewes were bilaterally ovariectomized in early October. During the onset of the winter solstice (late December), a 10 ml blood sample was collected from five ewes for analysis of serum levels of E₂ and progesterone. Ewes were then laparotomized and approximately one-third to one-half of a uterine horn was removed and assayed for endometrial nuclear and cytosolic ER. The contralateral horn was ligated at the external bifurcation and 10 μg of E₂ in 3 ml of physiological saline was injected into the uterine lumen of the ligated horn. After 48 h, a jugular blood sample was collected for steroid analysis and a section of the E₂ treated horn was removed and assayed for endometrial cytosolic and nuclear ER. This procedure was repeated on the remaining five ewes during the height of the summer solstice (late June). Endometrial nuclear and cytosolic concentrations of ER prior to and after exogenous E₂ stimulation were similar during the winter and summer solstice (p>.05). However, treatment with E₂ increased endometrial nuclear and cytosolic concentrations of ER compared with those of the nonstimulated uterine horn during the winter and summer solstice (p<.05 for each). Serum levels of E₂ prior to luminal treatment of ewes with E₂ during the winter and summer solstice did not differ (16.55 ± 4.05 vs 16.00 ± 3.0 pg/ml, respectively, p>.05). Serum levels of E₂ 48 h after administration of E₂ did not differ among ewes at the winter and summer solstice (18.75 ± 2.4 vs 18.65 ± 1.65 pg/ml, respectively, p>.05). Serum levels of progesterone were basal (<0.10 ng/ml) and did not differ in ewes prior to and after E₂ treatment at the winter and summer solstice (p>.05). These data indicate that physiological factors and(or) hormones such as prolactin and melatonin secreted in response to extremes in photoperiod do not appear to influence uterine concentrations of ER in ovariectomized ewes. / Graduation date: 1991

Thesis (Ph.D.)--Virginia Commonwealth University, 2009. / Prepared for: Dept. of Biochemistry. Title from resource description page. Includes bibliographical references.

Diss. (sammanfattning) Stockholm : Karol. inst., 2002. / Härtill 6 uppsatser.

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).

The present investigation was designed to further our understanding of the interaction of pituitary (luteinizing hormone, LH) and placental (chorionic gonadotropin, CG) gonadotropins with the primate corpus luteum. Studies were performed (1) to characterize the LH/CG receptor population in the

Thesis (M.S.)--University of Michigan, 1982.

Thesis (M.S.)--University of Michigan, 1982.

No description available.