Overweight and obesity are prevalent public health problems that impact physical, mental, and social health. Many studies have evaluated weight loss treatments, but most individuals are unsuccessful at maintaining weight loss long-term. Behavioral and cognitive interventions may be effective in promoting weight loss and weight loss maintenance. One cognitive intervention that has shown potential success in reducing weight and caloric intake is inhibitory control training (ICT). ICT involves trainings where individuals are asked to repeatedly withhold dominant responses to unhealthy or high-calorie food images in an effort to increase food-related inhibitory control abilities. Reductions in caloric intake or weight may occur after as little as one week of ICT; however, it is unclear how more frequent ICT sessions promote weight loss and reduce caloric intake. Further, studies on food-specific ICT are generally poorly powered and it is unclear how ICT affects underlying cognitive and neural mechanisms. One way to measure inhibitory control processes is through the N2 component of the scalp-recorded event-related potential (ERP). The amplitude of the N2 ERP component tends to be larger (i.e., more negative) when an individual inhibits a dominant response during go/no-go tasks compared to non-inhibition go trials. I conducted a quasi-randomized controlled trial where 100 individuals with overweight or obesity were assigned to either a generic (active control; n = 48) or food-specific ICT (experimental group; n = 52). ICTs were completed four times per week for four weeks. Weight and caloric intake were obtained at baseline, immediately after four-weeks of ICT, and at a 12-week follow-up. Participants also completed a high-calorie and a neutral go/no-go task while electroencephalogram (EEG) was recorded at each visit. Results from mixed model analyses suggest that neither weight, caloric intake, nor N2 ERP component amplitude towards high-calorie foods changed at post-testing or at the 12-week follow up for either group. Regression analyses suggest that individuals with lower baseline levels of inhibition may show greater weight loss and reductions in caloric intake after a generic ICT, while individuals with higher baseline levels of inhibition may show greater weight loss and reductions in caloric intake after a food-specific ICT. Self-report ratings indicated the appetitive drive towards food decreased over the course of the study, particularly for individuals with higher levels of baseline inhibition. Overall, generic- or food-specific ICT did not affect weight, caloric intake, or food-specific N2 ERP amplitude. Food-specific ICT may be more effective in reducing caloric intake and weight for individuals with larger inhibition responses to food stimuli, while generic ICT may be more effective in reducing caloric intake and weight for individuals with smaller inhibition responses to food stimuli. ICT may also be targeting other mediating processes, such as the appetitive value of food, as opposed to improving food-specific inhibitory control.

Eating unhealthy foods and eating past satiety are inappropriate behaviors that promote obesity. The ability to effectively inhibit an inappropriate behavior is a key component of cognitive restraint and its impairment has been previously linked to obesity. In this study, a Go/No-Go fMRI task was completed by a cohort of adult women that had experienced initial weight loss followed by various levels of weight regain or continued weight loss. Region of interest fMRI analysis revealed that greater total weight loss was significantly related to decreasing activation in the right inferior frontal gyrus and the right superior frontal gyrus. These results suggest that as weight loss increases fewer cognitive resources are needed in order to maintain levels of inhibitory control. This cognitive efficiency, though only partially supported by better task performance, is supported by greater exercise. An analysis of resting state patterns of correlation between task-activated regions revealed a significant correlation between the right inferior frontal gyrus and the left middle temporal gyrus. The strength of this relationship was significantly correlated with increasing total weight loss and continued weight loss over time. Cognitive restraint was also associated with this fronto-temporal correlation and provides support for cognitive efficiency. Right inferior frontal gyrus was also correlated with left inferior frontal gyrus and this relationship was positively correlated with initial weight loss suggesting that fewer neurocognitive resources were required by those who were able to achieve greater initial weight loss.

Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Bc. Nela Drnková Supervisor: Mgr. Marcela Vejsová, Ph.D. Title of diploma thesis: Evaluation of activity of potencial antifungal substances trought the use of microdilution broth method Background: The aim of this thesis was evaluation of antifungal activity of 70 substances trought the use of microdilution broth method. Substances for testing were synthesized at the Faculty of Pharmacy and they were divided into 5 groups based on chemical structure. Methods: The microdilution broth method was used for testing. The antifungal activity of substances was tested on 8 strains of yeasts and filamentous fungi: Candida albicans, C. tropicalis, C. glabrata, C. krusei, Trichosporon asahii, Aspergillus fumigatus, Absidia corymbifera a Trichophyton mentagrophytes. Results: The most active substances were in group of salicylanilide N- monosubstituted and N,N-disubstituted carbamates and thiocarbamates. The most effective substances were SAL-1L, SAL-1M, SAL-1N. They were effective on Trichophyton mentagrophytes already at concentration of 1,95 µmol.l-1 . The widest spectrum had substance GP-137, it inhibited the growth of all tested strains, but only at high concentrations. Rhodamine...

Acetylated adenosine diphosphoribose (OAADPR) is a product of protein deacetylation catalysed by class III of histone deacetylases called sirtuins. Sirtuins deacetylate histones and other proteins by unique mechanism coupled with consumption of stoichiometric amount of NAD+ . Sirtuins and OAADPR are implicated in the regulation of gene transcription, signalling and metabolic pathways and lifespan extension, thus preventing the development of age-related diseases. Even though, sirtuins are well studied, the exact biological role of OAADPR remains mainly unknown. Its further exploration is restricted by OAADPR's proneness to enzymatic hydrolysis. Therefore, non-hydrolysable analogues of OAADPR are needed to establish its biological function. These analogues are also expected to be competitive inhibitors of sirtuins, which may reveal their potential as therapeutic agents. A series of OAADPR analogues in which the acetate moiety was replaced with alkylcarbonate functionality has been synthesized. The studies of alkylcarbonate migration on furanoside scaffold have established the stability of alkylcarbonate vs. acetate under various conditions. Generally, alkylcarbonates are more stable than acetate under acidic or neutral conditions whereas under basic conditions they seem to be less stable....

Este estudo teve por objetivo avaliar a ação inibitória de duas soluções de quitosana através da determinação da concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM) de duas soluções de quitosana de diferentes procedências. A primeira solução de quitosana derivada de camarão, do tipo comercial Fluka, de PM 600.000 g/mol, G.A. 76% e a segunda solução de quitosana derivada de lula, de PM: '10 POT.7' g/mol e G.A.: 83%. As duas soluções foram ajustadas ao pH 5,0 e na concentração de 0,5% em solução acética a 1%. A melhor atividade antimicrobiana da quitosana ocorre em pH igual ou menor que cinco e ela sofre precipitação em pH superiores a 6,5. Estas duas características foram determinantes na escolha do pH utilizado no teste da CIM. Para confirmar que a inibição do crescimento dos enteropatógenos ocorreu pela ação da quitosana e não pelo pH ácido dos meios, vários testes foram realizados. A avaliação do crescimento dos enteropatógenos em ágar MacConkey, pH 5,0 (ótimo para quitosana) e 7,4(ótimo para o cultivo das bactérias utilizadas), o inóculo de cada bactéria foi preparado segundo o tubo 0,5 de Mc Farland (controle positivo), e a avaliação foi repetida utilizando o inóculo diluído 1:1000 para contagem do número de colônias, e não apresentaram diferenças significativas. Para confirmar estes dados foram realizados os controles negativos das duas soluções de quitosana e do meio de cultura MacConkey em pH 5,0 e 7,4, incubados a 37°C e lidos por 72 horas, sem qualquer alteração. A avaliação da reação de precipitação foi feita em caldo Müeller Hinton em pH 4,0; 5,0; 6,0; 7,0 e 8,0 para as duas soluções de quitosana (v/v), incubados a 37°C e lidos por 72 horas. A avaliação da CIM das duas soluções de quitosana para os enteropatógenos foi realizada por diluição seriada e os inóculos comparados ao tubo 0,5 de Mc Farland, sendo 10 'mü'L da suspensão bacteriana acrescida a cada tubo, incubados a 37°C por 24 horas. A leitura da ausência de turvação visível caracterizou a CIM. Todos os tubos que não apresentaram turvação visível foram plaqueados em ágar MacConkey, em pH 7,4 e incubados a 37°C por 24 horas para determinação da CBM, a qual foi determinada pela menor concentração capaz de causar morte total da população dos enteropatógenos. Todas as técnicas foram realizadas em triplicata para confirmação dos resultados / The aim of this study was to evaluate the inhibitory action of chitosan solutions derived from shrimp (Fluka commercial type, MW 600.000 g/mol, acetylation degree of 76%) and squid (MW '10 POT.7' g/mol, acetylation degree of 83%) through determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against enteropathogens: Salmonella enterica, Shigella sonnei and Escherichia coli EPEC. Those solutions were set to pH 5,0 and a 0,5% concentration in a 1% acetic acid solution. The best antimicrobial activity of chitosan occurs in pH less than or equal to 5,0 and it shows precipitation in pH greater than 6,5. Those features were decisive to choose the pH used in the MIC test. In order to confirm that the growth inhibition of enteropathogens occurred by the action of chitosan and not for the acid pH of the environments, growth evaluation tests of enteropathogens were accomplished in MacConkey agar, pH 5,0 (excellent for chitosan) and pH 7,4 (excellent for culture of used bacteria). The inoculum of each bacterium was prepared comparing with the 0,5 tube of McFarland (positive control) and the evaluation was repeated using the inoculum diluted in a salt solution 1:1000 to count the number of colonies, which did not show significant differences. The reaction evaluation of precipitation of chitosan was done in Müeller Hinton broth with pH ranging 4,0 - 8,0 for both solutions of chitosan (v/v), which were incubated at 37°C and read for 72 hours. The MIC evaluation for both solutions of chitosan for the enteropathogens was done by serial dilution and the inocula were compared to the 0,5 tube of McFarland, adding 10 'mü'L of bacterial suspension to each tube, which were incubated at 37°C for 24 hours. The MIC was distinguished by the absence of visible turbidity. Each tube that did not show visible turbidity was spread on MacConkey agar plates in pH 7,4, and incubated at 37°C for 24 hours to find the MBC, which was determined by the smallest concentration able to cause total death to the enteropathogen population. In both cases, the solutions of chitosan presented a high antimicrobial activity against the enteropathogens Salmonella enterica and Escherichia coli EPEC. However, the higher antimicrobial activity was observed in the enteropathogen Shigella sonnei

糖尿病是一种呈现流行趋势的代谢紊乱综合症，现如今，全球大约有3.46亿糖尿病患者， 这庞大的数字给各国的公共健康安全支出带来了严重的财政负担。 其中，二型糖尿病（T2DM）占90%。其特点是周围组织的胰岛素抵抗以及后期损伤的胰岛β细胞的功能。在饮食后，小肠会分泌两种肠促胰岛素，葡萄糖依赖性促胰岛素多肽（GIP）和胰高血糖素样肽-1（GLP-1）。两种多肽的主要功能是促进餐后胰岛细胞中胰岛素的分泌，另外他们还可以通过其自身的G蛋白偶联受体，GIPR和GLP-1R发挥其他作用，如葡萄糖依赖性的刺激胰岛素的生成，刺激胰岛β细胞的增殖，抑制细胞的凋亡等。这些功能也使肠促胰岛素成为糖尿病治疗的一种手段，比如Exendin-4和DPP4抑制剂。 然而，除了在胰岛中的作用，肠促胰岛还可能和脂质代谢相关，其中GIP和脂质代谢的报导研究的更加深入。在肥胖的状态下，血液中GIP含量高于正常水平；GIPR基因敲除老鼠和GIPR的抑制剂喂养的小鼠可以抵抗高脂饮食诱导的肥胖和2型糖尿病；GIP还可以直接调节脂肪细胞的脂肪生成和脂解。这些数据表明GIP在肥胖和糖尿病的发生过程中可能存在促进作用,这使得GIP治疗药物的开发需要谨慎的对待。 / 为了进一步研究GIP在脂肪细胞中发挥的生物学效应，在本研究中，我们利用腺病毒介导技术通过在脂肪细胞中过表达GIPR来增加GIP的活性，然后检查GIP在脂肪细胞中所起的作用。实验结果表明,GIP可以通过cAMP-PKA信号通路迅速并且长期的刺激脂肪细胞的炎症反应，增强IKKβ-NFκB信号通路和增加炎症基因的表达。更深入的机制研究表明，JNK 信号通路也参与GIP诱导的炎症反应，抑制JNK通路可以大部分恢复GIP增加的炎症因子的表达和IKKβ的磷酸化水平。由于长期的炎症反应，脂肪细胞的胰岛素信号通路受到GIP的损伤，在GIPR过表达的脂肪细胞中，胰岛素刺激的AKT磷酸化水平和葡萄糖吸收能力都被GIP降低，葡萄糖转运蛋白4（Glut-4）的表达水平也同时减少。因此，本研究结果表明GIP可能在肥胖的发展过程中，通过诱导脂肪细胞的炎症反应来损伤胰岛素敏感性而最终导致2型糖尿病的发生。 / Diabetes mellitus is a type of metabolic syndrome that has prevailed all over the world with the development of economic and over-nutrient lifestyle. It is estimated to 346 million diabetes patients in the worldwide most recently. The huge population put a major burden on the cost of public health care to all the countries. Among the types of diabetes, type 2 diabetes (T2DM) makes up 90% of recorded cases. The characteristics of T2DM are insulin resistance of peripheral tissues and impaired pancreatic cell function and mass. Two major incretins GIP (glucose-dependent insulinotropic peptide) and GLP-1 (glucagon-like peptide 1) are secreted from gut in response to food ingestion. The prominent role of GIP and GLP-1 is to stimulate glucose-dependent insulin release in pancreatic β cell. In addition, they both exert multiple biological effects via their relative G-protein coupled receptors, GIPR and GLP-1R, including glucose-stimulated insulin production, cell proliferation and anti-apoptosis in pancreatic β cells. The beneficent effects of incretins potentiate them as targets for the treatment of diabetes. GLP-1 analog, exendin-4 and DDP4 (dipeptidyl peptidase-4) inhibitors (to prevent GIP and GLP-1 from degradation) have been already used in clinical research. However, in addition to their effects on pancreatic β cell, both peptides are also related to lipid metabolism. The role of GIP has been studied more extensively. In obese state, the circulating level of GIP is elevated. GIPR knockout (KO) mice are resistant to high fat diet (HFD) induced obesity, a similar phenotype is found in GIPR antagonist administrated HFD-mice. Moreover, GIP also directly promotes lipogenesis and lipolysis in adipocytes. The rising evidence suggests a potential role of GIP in adipocyte biology and lipid metabolism, which diminishes the enthusiasm of GIP as a candidate therapeutic reagent for T2DM. / In order to further understand the biological effects of GIP in adipocytes, here, we over-expressed GIPR in 3T3-L1 CAR adipocytes via adenovirus-mediated gene transfer technology to enhance the activity of GIP. The results demonstrate that GIP impairs the physiological functions of adipocytes as a consequence of increasing the production of inflammatory cytokines, chemokines, and phosphorylation of IkB kinase (IKK) β through activation of the cyclic AMP-protein kinase A (cAMP-PKA) pathway. Activation of Jun N-terminal Kinase (JNK) pathway is also observed in GIP-induced inflammatory responses in adipocytes. An inhibitor of JNK blocks GIP-stimulated secretion of inflammatory cytokines and chemokines, as well as phosphorylation of IKKβ. The chronic inflammatory response eventually impairs insulin signaling in adipocytes, as demonstrated by reduction of protein kinase B (PKB/AKT) phosphorylation. The subsequently physiological analysis also indicates that GIP inhibits insulin-stimulated glucose uptake, and gene expression analysis reveals a decrease of glucose transporter 4 (Glut-4) in the meanwhile. The results suggest that GIP may be one of stimuli attributable to obesity induced insulin resistance via induction of adipocyte inflammation. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Nie, Yaohui. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 95-111). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgements --- p.v / INTRODUCTION --- p.1 / Chapter Part 1 --- Obesity and Type 2 diabetes --- p.1 / Chapter 1.1 --- Introduction to diabetes --- p.1 / Chapter 1.1.2 --- Physiology of adipocyte --- p.4 / Chapter 1.1.3 --- Mechanism of obesity induced diabetes --- p.10 / Chapter Part 2 --- Incretins and T2DM --- p.12 / Chapter 2.1 --- History of incretins --- p.12 / Chapter 2.2 --- Physiological actions of incretins --- p.14 / Chapter 2.3 --- Molecular mechanism of incretin actions in pancreas --- p.16 / Chapter 2.4 --- Incretins and T2DM --- p.19 / Chapter Part 3 --- Incretins and lipid metabolism --- p.23 / Objective --- p.26 / Methods and materials --- p.28 / Chapter 1 --- Cell culture --- p.28 / Chapter 1.1 --- 3T3-L1 culture and differentiation --- p.28 / Chapter 1.2 --- 3T3-L1 CAR culture and differentiation --- p.29 / Chapter 2 --- Cloning and recombinant adenovirus construction --- p.30 / Chapter 2.1 --- Plasmid construct --- p.30 / Chapter 2.2 --- Construct of recombinant adenoviruses --- p.30 / Chapter 2.3 --- Generation and infection of the adenoviruses --- p.31 / Chapter 3 --- Physiological and morphological assays --- p.32 / Chapter 3.1 --- Lipolysis assay --- p.32 / Chapter 3.2 --- TUNEL assay --- p.32 / Chapter 3.3 --- Glucose uptake --- p.33 / Chapter 3.4 --- Glut-4 localization --- p.33 / Chapter 4 --- Gene expression analysis --- p.35 / Chapter 4.1 --- Quantitative real-time PCR --- p.35 / Chapter 4.2 --- Immunoblot analysis --- p.35 / Chapter 4.3 --- ELISA assay --- p.36 / Chapter 5 --- Isolation of primary adipocytes --- p.37 / Results --- p.38 / Chapter Part 1 --- Role of GIP in 3T3-L1 cells --- p.38 / Chapter 1.1 --- Differentiation of 3T3-L1 adipocytes --- p.38 / Chapter 1.2 --- GIP slightly stimulates phosphorylation of p-CREB and lipolysis in 3T3-L1 cells. --- p.40 / Chapter 1.3 --- Analysis of gene expression in GIP-treated adipocytes --- p.42 / Chapter 1.4 --- Discussion --- p.44 / Chapter Part 2 --- Role of GIP in GIPR over-expressing 3T3-L1 CAR adipocytes --- p.46 / Chapter 2.1 --- Differentiation of 3T3-L1 CAR adipocytes --- p.46 / Chapter 2.2 --- Functional tests in GIPR over-expressing 3T3-L1 CAR adipocytes. --- p.48 / Chapter 2.3 --- Effect of GIP on cell viability --- p.50 / Chapter 2.4 --- Analysis of gene expression in GIP-treated adipocytes --- p.52 / Chapter 2.5 --- GIP activates inflammatory responses in GIPR over-expressing adipocytes --- p.54 / Chapter 2.6 --- Inhibition of IKKb pathway restores GIP-induced inflammatory responses --- p.56 / Chapter 2.7 --- Effects of GIP on adipocytes are partially dependent on the cAMP-PKA pathway --- p.58 / Chapter 2.8 --- Activation of cAMP-PKA pathway induces adipocyte inflammation. --- p.60 / Chapter 2.9 --- cAMP-Epac pathway is not involved in GIP-induced inflammation --- p.62 / Chapter 2.10 --- GIP stimulates cell stress activated kinases --- p.64 / Chapter 2.11 --- JNK partially mediates GIP-induced adipocyte inflammation --- p.65 / Chapter 2.12 --- Inhibition of JNK pathway partially restores GIP-induced inflammatory responses --- p.67 / Chapter 2.13 --- GIP impairs insulin signaling in GIPR over-expressing 3T3-L1 CAR adipocytes via inducing inflammatory response --- p.69 / Chapter 2.14 --- GIP enhances basal glucose uptake but impairs insulin stimulated glucose uptake in 3T3-L1 CAR GIPR over-expressing adipocytes --- p.71 / Chapter 2.15 --- Discussion --- p.73 / Chapter Part 3 --- Role of GIP in primary adipocytes --- p.78 / Chapter 3.1 --- GIPR expression level in primary adipocytes --- p.78 / Chapter 3.2 --- Analysis of gene expression in primary adipocytes after GIP treatment --- p.80 / Chapter 3.3 --- Discussion --- p.81 / SUMMARY --- p.82 / Chapter Future investigation --- p.83 / Chapter Appendix 1: --- Abbreviations --- p.86 / Chapter Appendix 2: --- Protocols --- p.90 / Preparation of competent cells --- p.90 / Outlines of recombinant adenovirus preparation --- p.91 / Virus titering (TCID50) --- p.92 / Primers for real-time PCR --- p.93 / Chapter Publications and Scientfic activities --- p.94 / Thesis related publication: --- p.94 / Other pubiliations: --- p.94 / Scientific activities: --- p.94 / References --- p.95

Negative life stress and maternal inhibitory control are both critical ingredients involved in the shaping and maintaining of the quality of parenting behaviors. This study explored both how the experience of stressful life events and inhibitory control relate to two particular types of parenting behaviors: harsh/controlling and autonomy-supportive. Given that these two types of parenting have broad implications for children’s developmental trajectories, it is important to further enhance our understanding of the etiological factors that both shape and maintain parenting practices. Utilizing a high-risk sample (i.e. low SES, high presence of documented child maltreatment) of mothers with pre-school aged children, this study did not support the relationship between the experience of stressful life events, maternal inhibitory control and quality of parenting. However, post hoc analyses of life stress using a measure of objective SES did yield a significant link between stress and the presence of autonomy-supportive parenting. This study expands the current understanding of how stress and inhibitory control relate to parenting behaviors. Implications of this study for practice and research are discussed.

The balance between excitatory and inhibitory synaptic transmission within and across neurons in active networks is crucial for cortical function and may allow for rapid transitions between stable network states. GABAergic interneurons mediate the majority of inhibitory transmission in the cortex, and therefore contribute to the global balance of activity in neuronal networks. Disruption in the network balance due to impaired inhibition has been implicated in several neuropsychiatric diseases (Marin 2012). Both schizophrenia and autism are two highly heritable cognitive disorders with complex genetic aetiologies but overlapping behavioural phenotypes that share common imbalances in neuronal network activity (Gao & Penzes 2015). An increasing body of evidence suggests that functional abnormalities in a particular group of cortical GABAergic interneurons expressing the calcium-binding protein parvalbumin (PV) are involved in the pathology of these disorders (Marin 2012). As deficits in this neuronal population have been linked to these disorders it could be useful to target them and increase their activity. A conserved feature in PV cells is their unusually low input resistance compared to other neuronal populations. This feature is regulated by the expression of leak K+ channels, believed to be mediated in part by TASK and TREK subfamily two-pore K+ channels (Goldberg et al. 2011). The selective blockade of specific leak K+ channels could therefore be applied to increase the activity of PV cells. In this thesis, specific TASK-1/3 and TREK-1 channel blockers were applied in cortical mouse slices in an attempt to increase the output of PV cells. The blockade of either channel did not successfully increase the amplitude of PV cell-evoked inhibitory postsynaptic currents (IPSCs) onto principal cells. However, while the blockade of TASK-1/3 channels failed to depolarise the membrane or alter the input resistance, the blockade of TREK-1 channels resulted in a small but significant depolarisation of the membrane potential in PV cells. Interestingly, TREK-1 channel blockade also increased action potential firing of PV cells in response to given current stimuli, suggesting that TREK-1 could be a useful target for PV cell modulation. These results demonstrate for the first time the functional effects of using specific two-pore K+ channel blockers in PV cells. Furthermore, these data provide electrophysiological evidence against the functional expression of TASK-1/3 in PV cells. It could therefore be interesting to further characterise the precise subtypes of leak K+ channels responsible for their low resistivity. This would help to classify the key contributors of the background K+ conductances present in PV cells in addition to finding suitable targets to increase their activity.

The present study was designed to analyze the transfer of stimulus control in temporal fading procedures. Several aspects of temporal fading procedures were manipulated including sources of inhibitory stimulus control, delays of reinforcement, and rates of increase in the temporal parameter of a fading procedure. In Experiment I, previous research producing transfer of stimulus control in a temporal fading procedure was directly replicated and controls were implemented for the operation of inhibition. The results showed that inhibitory stimulus control is not necessary in order to produce a transfer as participants with neutral stimulus backgrounds also transferred from one dimension to another without errors. However, positive stimulus backgrounds in the fading procedure prohibited the participants from achieving an errorless transfer of discrimination learning. In Experiment II, a fixed trial duration was employed with a constant and equal delay of reinforcement for both new and original stimulus dimensions. In this condition, participants did not transfer from one dimension to another with up to 30-second delays. Control participants were yoked to participants exposed to delayed and fading procedures to examine response latencies under delayed reinforcement for a simultaneous discrimination. There were no discernible response patterns under this condition except that participants continued to emit relatively short response latencies with a 40-second delay of reinforcement. In Experiment III, the effects of different steps of temporal fading on transfer were examined. The results showed that as the step of delay increased (10 sec. per trial), subjects transferred earlier in the fading series. Also, subjects with extremely low steps of delay (.1 sec. per trial) tended to remain with the original stimulus dimension. Experiments I through III demonstrated the necessity of either inhibitory or neutral stimulus backgrounds, differential delays of reinforcement correlated with each stimulus dimension, and relatively rapid increments in delay of the original stimulus dimension to obtain transfers of stimulus control in temporal fading procedures. When excitatory stimulus backgrounds were employed, or no differential delay of reinforcement was present, or the delay of the original stimulus dimension increased slowly, errorless transfers were not obtained. Overall, the results indicate that temporal fading procedures are a reliable, although complexly controlled, means of obtaining transfer between two stimulus dimensions.

Interleukin-6 (IL-6), oncostatin M (OSM), leukaemia inhibitory factor (LIF) and cardiotrophin-1 (CT-1) are members of the IL-6-type cytokine family that is characterised by sharing the common receptor subunit gp130. While the involvement of these polypeptides in cell differentiation, cell survival, proliferation, apoptosis, inflammation, haematopoiesis, immune response and acute phase reaction has already been demonstrated, the description of their role in development and progression of cardiac hypertrophy is still rather limited. A model has been postulated that declares the transient expression of IL-6-type cytokines as protective, while a continuous cardiac secretion of these proteins seems to be rather harmful for the heart. Within the first part of the study (results 4.1, 4.2 and 4.3) it was shown that OSM induces hypertrophy of primary neonatal rat cardiomyocytes (NRCM), just as its related cytokines LIF, CT-1 and hIL-6/hsIL-6R (hsIL-6R, human soluble IL-6 receptor). Regarding the hypertrophic potentials the LIFR/gp130 utilising cytokines (hLIF, hOSM and hCT-1) are stronger inducers than the OSMR/gp130 utilising mOSM. Human IL-6/hsIL-6R which signals via a gp130 homodimer has the weakest hypertrophic effect. The thorough analysis of typical signalling pathways initiated by IL-6-type cytokines revealed that STAT3 phosphorylation at Y705 seems to be the most important hypertrophy promoting pathway. In addition and in contrast to published work, we clearly demonstrate that classical IL-6 signalling (upon pure IL-6 treatment) has no hypertrophic effect on cardiomyocytes, because they lack sufficient amounts of the membrane-bound IL-6R. This is also true for neonatal rat cardiac fibroblasts (NRCFB). Since these cells can also influence cardiac hypertrophy, signalling pathways and target genes were additionally examined in NRCFB in response to OSM, LIF and IL-6/sIL-6R. One of the key findings of this thesis is the selective change in expression of cytokines and receptors of the IL-6 family in both cell types upon IL-6-type cytokine stimulation. A striking difference between NRCM and NRCFB is the fact that the target gene induction in NRCM is of similar duration upon mOSM and hIL-6/hsIL-6R treatment, while hIL-6/hsIL-6R is capable of promoting the induction of OSMR and IL-6 significantly longer in NRCFB. By searching for transcription factors or intermediate cytokines which could be responsible for this difference, a strong correlation between increased Il6 transcription and amount of mRNA levels for C/EBPβ and C/EBPδ was observed in response to IL-6/sIL-6R stimulation. Interestingly, mOSM also mediates the induction of C/EBPβ and δ, but the initiation is significantly less efficient than in response to IL-6/sIL-6R. Therefore, we assume that mOSM stimulation fails to reach threshold values required for a prolonged IL-6 secretion. Since we additionally observe a slight IL-6R mRNA upregulation in NRCFB, we assume that the combination of IL-6, LIF, C/EBPβ, C/EBPδ and IL-6R expression might be responsible for the observed different kinetics with which IL-6 and OSM stimulate NRCFB. In addition to the aforementioned proteins, members of the renin-angiotensin system seem to support the IL-6-type cytokine mediated hypertrophy. Since it has already been shown that angiotensin II vice versa induces IL-6 expression in NRCM and NRCFB, this enhanced expression of AT1α and ACE could be of crucial interest for the hypertrophy supporting phenotype. The second part of the presented work dealt with the characterisation of the receptor complexes of rat OSM. The central question of this analysis was, whether rOSM, just like mOSM, only binds the type II (OSMR/gp130) receptor complex or is able to utilise the type II and type I (LIFR/gp130) receptor complex. Using different experimental approaches (knock-down of the OSMR expression by RNA interference, blocking of the LIFR by LIF-05, an antagonistic LIF variant, and generation of stably transfected Ba/F3 cells expressing the newly cloned rat OSMR/gp130 or LIFR/gp130 receptor complex) we can clearly show that rat OSM surprisingly utilises both, the type I and type II receptor complex. Therefore it closely mimics the human situation. Furthermore, rOSM displays cross-species activities and stimulates cells of human as well as murine origin. Its signaling capacities closely mimic those of human OSM in cell types of different origin in the way that strong activation of the JAK/STAT, the MAP kinase as well as the PI3K/Akt pathways can be observed. Therefore, the results obtained in the last section of this thesis clearly suggest that rat disease models would allow evaluation of the relevance of OSM for human biology much better than murine models. / Interleukin-6 (IL-6), Oncostatin M (OSM), Leukämie inhibierender Faktor (LIF) und Cardiotrophin-1 (CT-1) sind Mitglieder der IL-6-Typ Zytokin-Familie, welche durch die gemeinsame Nutzung der Rezeptoruntereinheit gp130 charakterisiert ist. Während eine Beteiligung dieser Proteine bei Zelldifferenzierung, Zellüberleben, Proliferation, Apoptose, Entzündung, Hämatopoese, Immunantwort und Akut-Phase-Reaktion bereits gezeigt wurde, ist die Beschreibung ihrer Rolle bei der Entstehung und dem Fortschreiten der kardialen Hypertrophie deutlich limitierter. Es wurde bereits ein Modell postuliert, nach dem die kurzzeitige Expression dieser Zytokine schützend wirkt, während eine andauernde kardiale Sekretion eher schädlich für das Herz zu sein scheint. Im ersten Teil der Arbeit (Ergebnisse 4.1, 4.2 und 4.3) konnte gezeigt werden, dass OSM wie auch seine verwandten Zytokine LIF, CT-1 und hIL-6/hsIL-6R (hsIL-6R, humaner löslicher IL-6 Rezeptor) Hypertrophie-induzierend auf primäre neonatale Ratten-Kardiomyozyten (NRCM) wirkt. Hinsichtlich ihres hypertrophen Potentials sind die Zytokine, welche über LIFR/gp130 signalisieren (hLIF, hOSM und hCT-1), die stärkeren Induktoren im Vergleich zu mOSM, welches den OSMR/gp130 Rezeptorkomplex bindet. Die Stimulation mit humanem IL-6/hsIL-6R hatte hingegen die schwächste hypertrophe Wirkung. Unsere genaue Analyse der typischen IL-6-Typ Zytokin vermittelten Signalwege enthüllte die Phosphorylierung von STAT3 an Y705 als offenkundig wichtigsten hypertrophen Weg. Zusätzlich dazu konnten wir auch zeigen, dass klassisches IL-6 Signalling (ohne sIL-6R) keinen hypertrophen Einfluss auf NRCM hat, da diesen Zellen ausreichende Mengen des membranständigen IL-6R fehlen. Diese Beobachtung steht in klarem Kontrast zu bereits publizierten Arbeiten. In den ebenfalls untersuchten neonatalen Ratten-Kardiofibroblasten (NRCFB) verhält es sich, was den IL-6R angeht, genauso wie in NRCM. Da auch diese Zellen eine kardiale Hypertrophie mit beeinflussen können, wurden in ihnen die gleichen Signalwege und Zielgene nach Stimulation mit OSM, LIF und IL-6/sIL-6R untersucht. Die selektive Expressionsregulation von Zytokinen und Rezeptoren der IL-6-Familie in beiden Zelltypen nach IL-6-Typ Zytokin Stimulation ist hierbei einer unserer wichtigsten Befunde. Ein gravierender Unterschied zwischen NRCM und NRCFB besteht darin, dass die mOSM und hIL-6/hsIL-6R vermittelte Geninduktion in NRCM von vergleichbarer Dauer ist, wohingegen sie sich in NRCFB unterscheidet. Bei der Suche nach Transkriptionsfaktoren oder intermediären Zytokinen, welche für diesen Unterschied verantwortlich sein könnten, beobachteten wir nach IL-6/sIL-6R Stimulation eine deutliche Korrelation zwischen der Il6-Transkription und den mRNA Mengen von C/EBPβ und C/EBPδ. Auch OSM ist in der Lage beide Transkriptionsfaktoren zu induzieren, jedoch viel ineffizienter als IL-6/sIL-6R. Wir vermuten, dass mOSM einen bestimmten Schwellenwert, der für die verlängerte IL-6 Sekretion benötigt wird, nicht erreicht. Da wir zusätzlich noch eine schwache Zunahme der IL-6R mRNA in NRCFB beobachten konnten, gehen wir davon aus, dass die Expression von IL-6, LIF, C/EBPβ, C/EBPδ und IL-6R für die unterschiedlichen Kinetiken, mit denen IL-6 und OSM NRCFB stimulieren, verantwortlich sein dürfte. Es scheinen auch Mitglieder des Renin-Angiotensin-Systems die IL-6-Typ Zytokin vermittelte Hypertrophie zu unterstützen. Da schon gezeigt wurde, dass Angiotensin II reziprok die IL-6 Expression induziert, könnte diese verstärkte Synthese von AT1α und ACE von größter Bedeutung für den Hypertrophie-unterstützenden Phänotyp sein. Der zweite Teil der Arbeit (4.4) beschäftigte sich mit der Charakterisierung der Rezeptorkomplexe des Ratten-OSM. Die zentrale Frage hierbei bestand darin, ob rOSM wie mOSM nur den Typ II (OSMR/gp130) Rezeptorkomplex bindet, oder wie das hOSM sowohl den Typ II als auch den Typ I (LIFR/gp130) Rezeptorkomplex benutzen kann. Mit Hilfe unterschiedlicher experimenteller Strategien (knock-down der OSMR Expression durch RNA-Interferenz, LIFR-Blockade durch antagonistisches LIF-05, und die Generierung von stabil transfizierten Ba/F3-Zellen, welche die hierzu klonierten OSMR/gp130 oder LIFR/gp130 Rezeptorkomplexe der Ratte exprimieren) konnten wir eindeutig zeigen, dass Ratten-OSM überraschenderweise beide Rezeptorkomplexe benutzt. In dieser Hinsicht verhält sich es sich wie das humane Homolog. Des Weiteren besitzt Ratten-OSM Kreuz-Spezies-Aktivität und stimuliert humane und murine Zellen. Das Signal-Potential von rOSM ist dem von humanem OSM auf Zellen unterschiedlichen Ursprungs sehr ähnlich. Das Zytokin ist befähigt JAK/STAT, MAP Kinase und PI3K/Akt Signalwege potent zu aktivieren. Deshalb deuten die Daten des zweiten Teils dieser Arbeit darauf hin, dass Krankheitsmodelle in Ratten die Evaluierung der Relevanz des OSM für die humane Biologie deutlich besser widerspiegeln würden als murine Modelle.