Bored at home?—A systematic review on the effect of environmental enrichment on the welfare of laboratory rats and mice

Boredom is an emotional state that occurs when an individual has nothing to do, is not interested in the surrounding, and feels dreary and in a monotony. While this condition is usually defined for humans, it may very well describe the lives of many laboratory animals housed in small, barren cages. To make the cages less monotonous, environmental enrichment is often proposed. Although housing in a stimulating environment is still used predominantly as a luxury good and for treatment in preclinical research, enrichment is increasingly recognized to improve animal welfare. To gain insight into how stimulating environments influence the welfare of laboratory rodents, we conducted a systematic review of studies that analyzed the effect of enriched environment on behavioral parameters of animal well–being. Remarkably, a considerable number of these parameters can be associated with symptoms of boredom. Our findings show that a stimulating living environment is essential for the development of natural behavior and animal welfare of laboratory rats and mice alike, regardless of age and sex. Conversely, confinement and under-stimulation has potentially detrimental effects on the mental and physical health of laboratory rodents. We show that boredom in experimental animals is measurable and does not have to be accepted as inevitable

A biochemical and molecularbiological characterisation and clarification of the toxic mechanism

Titelblatt Inhaltsverzeichnis 0.0 Zusammenfassung I 1.0 Einleitung 1 2.0 Material und Methoden 13 3.0 Ergebnisse 37 4.0 Diskussion 86 5.0 Literaturverzeichnis 103 6.0 Abkürzungsverzeichnis 110 7.0 Danksagung 112 8.0 Lebenslauf 113"Seehasen" sind marine Gastropoden, die den Schutz einer harten äußeren Schale zugunsten einer Reihe von chemischen Abwehrstrategien und defensiven Verhaltensweisen aufgegeben haben. Ihre wohl dramatischste - und sehr erfolgreiche - Waffe ist die gerichtete Abgabe einer purpurfarbenen Tinte bei Kontakt mit einem Prädator. In den späten 80iger Jahren des letzten Jahrhunderts wurde aus der Tinte der indopazifischen Art Aplysia kurodai ein hochmolekulares Glykoprotein aufgereinigt, das sich durch antibakterielle und spezifische zytolytische Eigenschaften auszeichnete - Aplysianin P tötete selektiv Tumorzellen. Der Mechanismus dieses Effekts blieb ungeklärt. In der vorliegenden Doktorarbeit wurde ein zytolytisches Glykoprotein aus Tinte der europäischen Art Aplysia punctata isoliert. APIT (A. punctata ink toxin) ist ein 60 kD-Protein mit spärlicher O-Glykosilierung. Absorptionspeaks bei 390/470 nm enthüllten den Kofaktor Flavin, der sich in anschließenden Experimenten als FAD weiter spezifizieren ließ. Die in eine Aminosäurenfolge übersetzte cDNA von APIT enthält das klassische dinukleotid-bindende Motiv FAD-abhängiger Oxidoreduktasen und ein GG-Motiv, das typisch für L-Aminosäure Oxidasen ist. APIT, wie auch Aplysianin P, löst unter in vitro-Bedingungen den Tod von Tumorzellen aus. Dieser ist durch einen schnellen Verlust von Stoffwechselaktivität und die Permeabilisierung der Zellmembran gekennzeichnet. Es ließen sich keinerlei Hinweise auf apoptotische Prozesse sammeln. Eine Analyse des Proteoms der behandelten Zellen ergab Hinweise auf eine Beteiligung von reaktiven Oxygenspezien am zytolytischen Geschehen. Demgemäß ließ sich zeigen, dass es sich bei APIT um eine L-Aminosäuren Oxidase handelt, die H2O2 über die oxidative Desaminierung von L-Lysin und L-Arginin aus dem Zellkulturmedium generiert. Das produzierte H2O2 ist alleiniger Auslöser der schnellen Zytolyse; diese lässt sich durch Zugabe von Catalase vollständig unterdrücken. Die Sekretion von Oxidasen ist ein bei Tieren nicht unübliches Mittel zur Verteidigung und auch Seehasen scheinen solch eine Strategie der ROS-Generierung zur Abwehr von Feinden einzusetzen."Sea hares" are marine gastropods which have replaced the physical protection of the hard outer shell with an elaborate set of chemical and behavioral defense techniques. The most dramatic - and very successful - one is the dicharge of a purple ink upon predation. In the late 1980's a high-molecular glycoprotein with antibacterial and specific cytolytic properties was purified from ink of the indopacific sea hare Aplysia kurodai - Aplysianin P selectively killed tumor cells. The mechanism of its effect so far has not been elucidated. In the present thesis a cytolytic glycoprotein from ink of the European species Aplysia punctata was isolated. APIT (A. punctata ink toxin) is a 60 kD-protein with sparse O-glycosilation. Absorption peaks at 390/470 nm revealed a flavin cofactor which subsequently turned out to be FAD. The translated sequence of the cloned cDNA contains a classical dinucleotide binding motive of FAD-dependent oxidoreductases and a GG-motive of L-amino acid oxidases. APIT, like Aplysianin P, in vitro triggers tumor cell death which is characterized by a rapid loss of metabolic activity and membrane integrity. No hints on apoptotic events are detected. Proteome analysis of treated cells revealed the involvement of reactive oxygen species (ROS) in the cytolytic process. Accordingly it is demonstrated that APIT functions as L-amino acid oxidase producing H2O2 via oxidative deamination of L-lysine and L-arginine from culture medium. The generated H2O2 is solely responsible for the fast cytolytic response, because the latter is abolished by addition of catalase. Secreted oxidases are a popular means of defence in animals and sea hares seem to share this strategy of self-defence via generation of ROS from a yet not verified natural substrat

Antitumor polyketide biosynthesis by an uncultivated bacterial symbiont of the marine sponge Theonella swinhoei

Bacterial symbionts have long been suspected to be the true producers of many drug candidates isolated from marine invertebrates. Sponges, the most important marine source of biologically active natural products, have been frequently hypothesized to contain compounds of bacterial origin. This symbiont hypothesis, however, remained unproven because of a general inability to cultivate the suspected producers. However, we have recently identified an uncultured Pseudomonas sp. symbiont as the most likely producer of the defensive antitumor polyketide pederin in Paederus fuscipes beetles by cloning the putative biosynthesis genes. Here we report closely related genes isolated from the highly complex metagenome of the marine sponge Theonella swinhoei, which is the source of the onnamides and theopederins, a group of polyketides that structurally resemble pederin. Sequence features of the isolated genes clearly indicate that it belongs to a prokaryotic genome and should be responsible for the biosynthesis of almost the entire portion of the polyketide structure that is correlated with antitumor activity. Besides providing further proof for the role of the related beetle symbiont-derived genes, these findings raise intriguing ecological and evolutionary questions and have important general implications for the sustainable production of otherwise inaccessible marine drugs by using biotechnological strategies

Widespread Occurrence and Genomic Context of Unusually Small Polyketide Synthase Genes in Microbial Consortia Associated with Marine Sponges

Numerous marine sponges harbor enormous amounts of as-yet-uncultivated bacteria in their tissues. There is increasing evidence that these symbionts play an important role in the synthesis of protective metabolites, many of which are of great pharmacological interest. In this study, genes for the biosynthesis of polyketides, one of the most important classes of bioactive natural products, were systematically investigated in 20 demosponge species from different oceans. Unexpectedly, the sponge metagenomes were dominated by a ubiquitously present, evolutionarily distinct, and highly sponge-specific group of polyketide synthases (PKSs). Open reading frames resembling animal fatty acid genes were found on three corresponding DNA regions isolated from the metagenomes of Theonella swinhoei and Aplysina aerophoba. Their architecture suggests that methyl-branched fatty acids are the metabolic product. According to a phylogenetic analysis of housekeeping genes, at least one of the PKSs belongs to a bacterium of the Deinococcus-Thermus phylum. The results provide new insights into the chemistry of sponge symbionts and allow inference of a detailed phylogeny of the diverse functional PKS types present in sponge metagenomes. Based on these qualitative and quantitative data, we propose a significantly simplified strategy for the targeted isolation of biomedically relevant PKS genes from complex sponge-symbiont associations