京都大学 / 新制・課程博士 / 博士(工学) / 甲第23494号 / 工博第4906号 / 新制||工||1766(附属図書館) / 京都大学大学院工学研究科都市環境工学専攻 / (主査)教授 高野 裕久, 教授 米田 稔, 准教授 上田 佳代 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

Worldwide, pharmaceutical compounds continue to increase in our aquatic environment. The predominant route into nature is through wastewater treatment plants since the elimination of residual pharmaceuticals is still not mainstream in WWTPs. Fluoxetine is an antidepressant which is commonly prescribed to treat human depression. Wastewater residual fluoxetine is typically found in waters around the world, and can thus affect exposed organisms, such as fish and invertebrates. However, how fluoxetine may affect mating behaviour in exposed organisms remains poorly understood, and particularly so in invertebrates. This is hampering our understanding of the consequences of our medicine leaking into nature because mating behaviour often affects fitness, and invertebrates are key organisms in food chains. Therefore, I here experimentally investigated long-term effects of environmental relevant concentration of fluoxetine (20 ng L-1) on mating behaviours of male and female freshwater isopod Asellus aquaticus. I demonstrate that fluoxetine reduced male mating attempts with receptive females. Further, there was a tendency for fluoxetine exposure to increase latency to form pre-copula. There was no effect of fluoxetine exposure on male latency to encounter females or female responses toward males. These results indicate that fluoxetine also can affect isopods by reducing mating behaviour. In the long-term, if reproduction is delayed or reduced, it may cause a reduction in populations and thus, alter the whole ecosystem.

This study aims to explore if it is possible to form separate categories of aspirated voiceless stops in a second language, distinct from the equivalent categories in the native language, for native speakers of a language with an intermediate degree of aspiration, and if such category formation is eased by long-term exposure to another language in which aspirated voiceless stops exist. Two groups of adult native Japanese speakers who had lived in Sweden for a long and short time respectively were recorded when reading a list of sentences containing word-initial, utterance-medial /p t k/ in Japanese and English. Both groups produced higher VOT values for the English stops than for the Japanese stops. The results were significant for /t/ and /k/ and for the long-term residents' /p/, but not for the short-term residents' /p/, presumably because of a low number of tokens. The results are nevertheless interpreted as evident of the possibility of phonetic category formation even though there is only a small difference from the corresponding category in the native language. Since both groups had established new phonetic categories, degree of exposure to Swedish is interpreted as likely not to be a relevant factor.

L'association entre l'exposition prolongée aux particules fines (PM2.5) et la mortalité pourrait varier selon les méthodes de contrôle de la confusion. L'objectif était de comparer les estimations de cette association issues d’approches avec différentes méthodes pour le contrôle de la confusion. En utilisant la cohorte du Système de surveillance des maladies chroniques du Québec (SISMACQ), les estimations d'association provenant d’une régression de Cox avec contraste interindividuel et d’une régression conditionnelle logistique stratifiée sur les cas avec contraste intra-individuel ont été comparées. De plus, les estimations d’association de modèles de Cox ajustés pour des variables individuelles et locales, et ajustés seulement pour des variables locales, en utilisant la cohorte Santé et environnement du recensement canadien (CSERcan) pour la province de Québec ont été comparées afin d’évaluer comment les estimations d’association varient avec les variables incluses dans les modèles. Quatre-vingt-dix-huit millions de personées-années du SISMACQ ont été enregistrées. Des augmentations de la mortalité de 3,0% (Intervalles de confiance [IC à 95%] 1,8-4,3) et de 9,3% (IC à 95%] 4,1-14,8) par augmentation de 3,4 µg/m3 ont été observées pour la régression de Cox et la régression logistique, respectivement. Vingt et un millions de personnes-années de la cohorte CSERcan ont été enregistrées. Des augmentations de la mortalité de 2,2% (IC à 95% : 1,9-2,5) et de 0,8% (IC à 95% :0,6-1,0) par 1 µg/m3 ont été observées pour le modèle ajusté pour des variables individuelles et locales, et pour celui ajusté pour des variables locales seulement. Ces résultats confirment l'association entre l'exposition prolongée aux PM2.5 et la mortalité, et suggèrent que les méthodes de contrôle de la confusion peuvent influencer l’importance et la variabilité de ces estimations. / The association between long-term exposure to ambient fine particles (PM2.5) and mortality may vary depending on the methods of confounding control. The objective was to compare estimates of association using approaches varying by their confounding control methods. Using the cohort from the Quebec Integrated Chronic Disease Surveillance System (SISMACQ), association estimates from a Cox proportional hazard model with inter-individual contrast and from a conditional logistic regression with intra-individual contrast were compared. Additionally, results from Cox models adjusted for individual and local confounders, and adjusted only for local confounders using the Canadian Census Health and Environment Cohorts (CSERcan) for the province of Quebec were compared to evaluate how estimates vary when adjusting for individual and contextual confounders compared to adjusting for contextual factors only. For the SISMACQ cohort, 98 million person-years were recorded. Increase in mortality of 3.0% (95% CI: 1.8, 4.3) and of 9.3% (95% CI: 4.1, 14.8) were observed per 3.4 µg/m3 increment in exposure for the Cox model and conditional logistic regression. For the CSERcan cohort, 21 million person-years were recorded. Increase in mortality of 2.2% (95% CI: 1.9, 2.5) and of 0.8% (95% CI: 0.6, 1.0) were observed per 1 µg/m3 increment in exposure for the model adjusted for both local and individual confounders, and for the model adjusted only for local confounders. These results confirm the association between long-term exposure tom PM2.5 and mortality, and suggest that confounding control methods can influence the magnitude and variability of estimates of association.

The aim of this Ph.D. thesis was to analyze the morphine-induced changes of frontal brain cortex protein composition in rats exposed to increasing doses of morphine (10-50 mg/kg) for prolonged period of time (10 days). The first part of this work was oriented to the analysis of the phenomenon of hypersensitization/superactivation of adenylyl cyclase (AC), which is regarded as one of the crucial molecular mechanisms causing drastic pathological consequences of drug addiction. The increase of AC activity represents a "compensatory" response and is functionally related to the desensitization of G protein response to prolonged morphine exposure of target cells. The clear desensitization of µ-OR- and δ-OR-stimulated G protein response by morphine was demonstrated in our laboratory by analysis of the dose-response curves of DAMGO and DADLE-stimulated, high-affinity [35 S] GTPγS binding in plasma membranes isolated from frontal brain cortex of rats exposed to morphine according to the same protocol as that used in my Ph.D. thesis (10-50 mg/kg, 10 days). The κ-OR-stimulated [35 S] GTPγS binding was unchanged. It has been determined the amount of all AC isoforms (AC I-IX) in plasma membranes (PM) isolated from control and morphine-treated rats which were sacrificed 24 hours since the last dose of morphine....

(Oberflächen-) Gewässer weltweit sind mit geringen Mengen (ng/L bis wenige µg/L) humaner Pharmazeutika belastet. Diclofenac (DCF; nicht-steroidal, entzündungshemmend) und Metoprolol (MTP; ß-Blocker) gehören entsprechend ihres hohen Verbrauchs zu den am häufigsten gefundenen Substanzen. Deren biologische Aktivität ist nicht auf den Menschen beschränkt. Gut konservierte Enzyme innerhalb der Vertebraten legen Auswirkungen auf Nicht-Zielorganismen wie Fische nahe, die bisher in Langzeituntersuchungen mit umweltrelevanten Konzentrationen unzureichend untersucht wurden. In der vorliegenden Arbeit wurden die physiologischen Effekte von DCF und MTP auf die Nil-Tilapie (Oreochromis niloticus), einem der wichtigsten Aquakulturfische weltweit, untersucht. In vitro konnte anhand primärer Hepatozyten gezeigt werden, dass bereits umweltrelevante Konzentrationen von DCF zu einer erhöhten Genexpression verschiedener Schlüsselenzyme der Detoxifizierung führten. Nach MTP-Exposition waren die Veränderungen weniger eindeutig. Beide Substanzen induzierten die Vitellogenin Genexpression, nur DCF jedoch bereits in umweltrelevanter Konzentration. In vivo wurden in zwei Langzeit-Expositionsversuchen die physiologischen Effekte vom befruchteten Ei bis 80 Tage nach Schlupf in O. niloticus untersucht. Beide Substanzen hatte keinen Einfluss auf Schlupferfolg und Überleben, das Wachstum war nach 80 Tagen nach Schlupf leicht reduziert. Die deutlichsten Auswirkungen waren histopathologische Veränderungen der Kiemen, veränderte Genexpressionen der Gonadotropine und eine erhöhte Expression von Vitellogenin. Die Ergebnisse legen eine stärkere östrogene Aktivität von DCF im Vergleich zu MTP nahe. Zusammenfassend sind die Bedenken gegenüber den Einzelsubstanzen eher gering, negative Auswirkungen auf die Reproduktion und sich verstärkende Effekte bei zeitgleicher Exposition gegenüber DCF und MTP lassen sich jedoch nicht ausschließen und sollten im Weiteren untersucht werden. / Surface waters worldwide are contaminated with low levels (ng/L up to few µg/L) of human pharmaceuticals. Diclofenac (DCF; non-steroidal, anti-inflammatory) and metoprolol (MTP; ß-blocker) are highly consumed and therefore commonly detected. Their biological activity is not restricted to humans. Well conserved enzymes within the vertebrates suggest effects on non-target organisms such as fish, poorly studied in long-term exposure experiments using environmentally relevant concentrations. In the presented work, physiological effects of DCF and MTP on the Nile tilapia (Oreochromis niloticus), an important aquaculture fish species, were studied. Using primary hepatocytes, it was shown in vitro that environmentally relevant concentrations of DCF increased the gene expression of different key enzymes of the detoxification, while MTP exposure had a less clear effect. Both substances induced vitellogenin gene expression, but only after DCF exposure this was significantly elevated already at the environmentally relevant concentration. In vivo, two long-term exposure studies on the physiological effects from the fertilized egg until 80 days post-hatch were evaluated. Both substances did not affect hatching success and survival, while growth was slightly reduced after 80 days post-hatch. Histopathological alterations of the gills, changed gene expression patterns of the gonadotropins and induced vitellogenin gene expression were the most dominant findings. The results indicate a stronger estrogenic mode of action of DCF compared to MTP. Overall, the risk due to a single substance exposure seems to be relatively low but adverse effects on reproduction and additive effects during simultaneous exposure to DCF and MTP cannot be excluded and should be investigated further.