1,720,962 research outputs found
Associations of Nicotinic Acetylcholine Receptor Subunits Expressed in the Rat Habenula
No full textPh.D.The goals of this study were to define the nicotinic acetylcholine receptor (nAChR) subtypes expressed in the rat habenula. This was done using three main techniques: single and sequential immunoprecipitation using subunit-specific antibodies, single-point binding with α-conotoxin mII, and saturation binding with [3H]-epibatidine and competition binding with nicotine, cytisine, AT-1001, and sazetidine-A.Single immunoprecipitation experiments revealed that every subunit probed was expressed in the habenula in significant quantities. The α5 subunit was expressed in approximately 26% of nAChRs, the α4 subunit was expressed in approximately 40% of receptors, α3 was expressed in approximately 32% of receptors, β2 was expressed in approximately 48% of receptors, and β4 was expressed in approximately 51% of nAChRs in the habenula. Sequential immunoprecipitations demonstrated that every subunit expressed was significantly associated with every other subunit with one notable exception: the α4 and α5 subunits were not discernably associated in the habenula. Additionally, we found that approximately 20% of nAChRs in the habenula contain both the β2 and β4 subunits. Finally, through binding assays with [3H]-epibatidine and α-conotoxin MII, we determined that there are no detectable α6-containing nAChRs in the habenula.Saturation binding assays determined that nAChRs in the habenula have a Kd for [3H]-epibatidine of 78 pM. Additionally, we found that approximately 20% of nAChRs have a high affinity for Sazetidine-A (a ligand with a high specificity for β2-containing nAChRs) and approximately 70% of nAChRs in the rat habenula had a high affinity for AT-1001 (a ligand with a high specificity for β4-containing nAChRs). From this data, and the data from the immunoprecipitation reactions, we hypothesized that the β2β4* nAChRs in the habenula indicate binding properties much like those of β4-containing nAChRs.Finally, nAChR subunits in the olfactory bulb and olfactory tubercle were probed by single and sequential immunoprecipitation. In the olfactory bulb, the α5 subunit was expressed in approximately 28% of nAChRs, the α4 subunit was expressed in approximately 48% of receptors, α3 was expressed in approximately 41% of receptors, β2 was expressed in approximately 71% of receptors, and β4 was expressed in approximately 33% of nAChRs
Pharmacological Properties of alpha4beta2 and alpha3beta4 Nicotine Acetylcholine Receptors: Ligand Binding, Activation, Desensitization, and Interspecies Differences
No full textPh.D.Smoking cessation medications using nicotinic ligands, including nicotine, cytisine, and varenicline, have traditionally revolved around selective activation of alpha4beta2 nicotinic acetylcholine receptors (nAChRs). However, nAChR desensitization may be the primary action of nicotine and may be a viable approach for smoking cessation medication. The alpha3beta4 nAChR subtype has been implicated in side effects of nicotinic drugs as well as nicotine addiction. The purpose of the work here is to investigate pharmacological properties of these two nAChR subtypes, with a focus on mechanisms of ligand binding, activation, desensitization, and interspecies differences between human and rat receptors.The first aim of this dissertation was to dissect the properties of six ligands (nicotine, cytisine, sazetidine-A, varenicline, epibatidine, and 5-I-A85380) at alpha4beta2 nAChRs. We developed a method for determining the dissociation rates of unlabeled ligands. These ligands are more potent at desensitizing than at activating alpha4beta2 nAChRs. A correlation was found between duration of desensitization and binding affinity.The second aim was to develop methods for using microfluidic laminar stream solution exchange (MLSSE) to study nAChRs expressed in HEK293 cell lines. The variables considered in this technique were drug exposure time and washout time. We demonstrated that MLSSE can be applied to a broad range of nAChRs.The final aim studied interspecies differences in alpha3beta4 drug affinity. All tested drugs besides acetylcholine have higher potency for human alpha3beta4 nAChRs than rat nAChRs. AT-1001, a new high affinity alpha3beta4 ligand, does not select for desensitization over activation in alpha3beta4 or alpha4beta2 nAChRs. Furthermore, the differences in affinity of AT-1001 between alpha3beta4 and alpha4beta2 nAChRs result in longer desensitization in alpha3beta4 nAChRs compared to alpha4beta2 nAChRs. This is consistent with the affinity of a ligand for a receptor subtype being the major determinant for desensitization length.The investigations in my dissertation contribute to a better understanding of the pharmacological properties of alpha4beta2 and alpha3beta4 nAChRs. The methods developed will help in future studies. Moreover, the results provide important insight in developing new nicotinic therapeutics based on nAChR desensitization and indicate that investigators should be aware of species differences in studying nicotinic ligands
REGULATION AND STOICHIOMETRY OF α4β2* NEURONAL NICOTINIC RECEPTORS
No full textPh.D.Nicotinic acetylcholine receptors (nAChRs) are present throughout the nervous system where they mediate and modulate neurotransmission. The α4β2* subtype is the most prevalent high affinity nAChR expressed in the brain. Models of nicotine addiction and dependence implicate this receptor subtype as central to smoking behavior. This work examines the regulation, expression, and pharmacology of this receptor subtype.In Part I, the density of nAChRs in the rat cortex is examined following a two phase treatment paradigm which emulates a chronic smoker who quits and begins a smoking cessation therapy. Rats were administered saline or nicotine then maintained on these treatments or switched to treatment with saline, varenicline, a currently approved FDA smoking cessation therapy, or sazetidine-A, a new ligand efficacious in reducing nicotine self-administration in animals. Assessments of nAChR radioligand binding density and α4 and β2 subunit protein densities reveal that varenicline maintains nicotine-induced up-regulation while sazetidine-A allows receptors to return to basal levels.Part II examines high and low sensitivity α4β2 nAChRs, corresponding to subunit stoichiometries of (α4)2(β2)3 and (α4)3(β2)2, respectively. In this section, transfection ratios of rat α4 and β2 subunit cDNA in HEK293 cells are used to create a model of high and low sensitivity α4β2 nAChRs. Through characterization of this model system, a method to detect nAChR stoichiometry using two concentrations of acetylcholine was validated. Applying this method to the rat motor cortex indicates that expression of the low sensitivity α4β2 nAChR predominates in this area. The effect of nicotine treatment on α4β2 nAChR stoichiometry was also assessed.In Part III, the pharmacology of six concatameric α4β2β4* nAChRs expressed in HEK293 cells is characterized alongside non-linked α4β2 and α4β4 nAChRs. Stable cell lines for each unique nAChR permutation were created and demonstrated to produce nAChRs that assemble as full length products which bind [3H]-Epibatidine with high affinity. Sazetidine-A, a β2 selective ligand, and AT-1001, a β4 selective ligand, were used to determine the presence of α4β2 and α4β4 binding sites on each concatamer permutation. These data suggest that the 3rd subunit in the pentameric sequence occupies the non-ligand binding site of the receptor
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Get PDFWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
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