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VU Research Portal Behavioral Phenotyping of Complex Traits in Inbred and Mutant Mice Maroteaux, G.P. 2014 document version Publisher's PDF, also known as Version of record Link to publication in VU Research Portal citation for published version (APA) Maroteaux, G. P. (2014). Behavioral Phenotyping of Complex Traits in Inbred and Mutant Mice. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal ? Take down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. E-mail address: [email protected] Download date: 23. Sep. 2021 BEHAVIORAL PHENOTYPING OF COMPLEX TRAITS IN INBRED AND MUTANT MICE Gregoire Pierre Maroteaux Cover and chapters Artwork: Anton Rammelt Printed by www.offpages.nl ISBN 978-94-6182-464-6 Copyright © G. Maroteaux, 2014 All rights reserved. No part of this thesis may be reproduced, stored in a retrieval system, or transmitted in any form or by any means without prior permission of the author. The printing of this thesis was sponsored by: VRIJE UNIVERSITEIT BEHAVIORAL PHENOTYPING OF COMPLEX TRAITS IN INBRED AND MUTANT MICE ACADEMISCH PROEFSCHRIFT ter verkrijging van de graad Doctor aan de Vrije Universiteit Amsterdam, op gezag van de rector magnificus prof.dr. F.A. van der Duyn Schouten, in het openbaar te verdedigen ten overstaan van de promotiecommissie van de Faculteit der Aard- en Levenswetenschappen op donderdag 25 september 2014 om 9.45 uur in de aula van de universiteit, De Boelelaan 1105 door Gregoire Pierre Maroteaux geboren te Versailles, Frankrijk promotor: prof.dr. M. Verhage copromotoren: dr. R.O. Stiedl dr. S. van der Sluis leescommissie: prof.dr. J.D. Armstrong prof.dr. B.M. Spruijt prof.dr. A.B. Smit dr. M. Kas dr. T. Pattij TABLE OF CONTENTS Chapter 1 General introduction 11 Behavioral phenotyping 13 Genetically engineered mice: 15 key to human genetic disease mechanisms Importance of the synapse in psychiatric disorders 18 Aim of this thesis 22 Chapter 2 High throughput phenotyping of avoidance 25 learning in mice discriminates different genotypes and identifies a novel gene Introduction 28 Material & methods 29 Results 32 Discussion 40 Supplemental 43 Chapter 3 Heterozygous Munc18-1 mice exhibit an increased 57 anxiety-like phenotype but no cognitive impairment Introduction 60 Material & methods 60 Results 62 Discussion 68 Conclusions 71 Supplemental 72 Chapter 4 Functional characterization of the PCLO S4814A 89 variant associated with major depressive disorder reveals cellular but not behavioral differences Introduction 92 Results 93 Discussion 106 Material & methods 107 Supplemental 114 Chapter 5 Random mutagenesis by transposon-based gene 125 trap insertion in mice identifies a role of Ubn1 in avoidance learning and fear conditioning Introduction 128 Material and methods 129 Results 133 Discussion 139 Supplemental 143 Chapter 6 General discussion 145 General aim 147 Limitations of standalone tests 148 Home cage relevance 149 The validation of new paradigms 152 Limitations of a new paradigm 154 Accuracy of animal models 155 Future directions 156 Final remarks 157 Chapter 7 Material & Methods 159 Laboratory animals 161 The test battery 161 Automated home cage observation and data analyses 169 Statistical analysis 170 References 173 Abbrevations 187 Samenvatting 189 Summary 193 Acknowledgment 195 Chapter General introduction 1 BEHAVIORAL PHENOTYPING In 1839, Darwin described the behavioral phenotype of 12 species of finches of the Galapagos 1 Islands. He demonstrated the correlation between feeding behavior, morphology of the beak, G and the environment of finches (Darwin, 1839) exemplifying the impact of genetic radiation on eneral morphological differences underlying behavioral adaptation even before the understanding of the genetic basis (Mendel, 1865). Behavioral phenotyping is the description and analysis of introduction internally coordinated responses (actions or inactions) of whole living organisms (individuals or groups) to internal and/or external stimuli (Levitis et al., 2009). The behavioral response of an organism is a complex process dependent on gene expression, physiology and interaction with the environment. Distinct behavioral deviations from normally observed responses can be predictive signs for neurological pathologies and psychiatric disorders in human. Thus, the understanding of the biological mechanisms involved in those behaviors is essential. The complexity of behavior requires the use of animal models to mimic human neurological and psychiatric diseases. The last decades were marked by the expansion of new technologies in genetics, brain imaging and molecular biology, to increase the precision, the wealth and the reproducibility of newly generated results. However, behavioral phenotyping – due to its complexity – has not yet experienced this revolution and still is hampered by poor reproducibility and low throughput (Crabbe et al., 1999). In this chapter, the reasons for the development of a new approach of behavioral phenotyping are introduced, followed by the relevance of investigating behavior in mice and comparing different genetic backgrounds or targeted mutations as well as in random mutation in different genes. Thereby the focus will be directed towards the importance of presynaptic genes in neurologic and psychiatric disorders. In need of a new behavior approach For decades, scientists strove to develop a variety of appropriate behavioral tests for rodent phenotyping. The most commonly used behavioral tests are based on locomotor activity measures in acute tests of short duration, normally 5 min to less frequently 60 min. Classical examples like anxiety assessment in an open field (OF), an elevated plus maze (EPM), or a dark- light box (DLB) (Crawley and Goodwin, 1980; Hall and L, 1932; Pellow et al., 1985) or learned behavior, such as performance in the Barnes maze task (BM) and the 5-choice serial reaction time task (5-CSRTT) (Barnes, 1979; Robbins, 2002). To describe the effect of a genetic background, a mutation or a drug on behavior, a battery of tests is required to tap into different aspects of behavior such as motor, sensory, cognitive and circadian functions. Such test batteries, like the SHIRPA protocol (Rogers et al., 1997) or IMPReSS protocol (Brown and Moore, 2012; Brown et al., 2005), are designed to cover muscle, cerebellar, sensory and neuropsychiatric functions. Those sets of experimental procedures contain different tests across several screens of increasing complexity and specialization (Nolan et al., 2000). These test batteries have standardized protocols to improve comparability and reproducibility across studies. However, the succession of tests in those batteries requires repeated human-animal interaction. Carry- over effects can be caused by the order in which tests are performed on the same cohort of animals. Moreover, environmental effects, the lack of consistency in the protocols between 13 laboratories, discrepancies in the data collection methods (visual scoring vs. computerized) and operational definition (e.g. definitions of grooming, rearing, lingering) are also sources of variation in the results that lead to different interpretations (Crabbe et al., 1999; Turner and Burne, 2013; Wahlsten et al., 2003; Würbel, 2002). Considering the huge number of variables that directly or indirectly affect behavioral testing, it is no surprise that behavioral testing is a bottleneck in screening for genes involved in neurological disorders and the development of novel drug treatments. Consequently, there is growing consensus that behavioral testing needs a boost toward automation to increase its effectiveness and standardization in the study of animal models of human diseases and therapeutic developments (Crabbe and Morris, 2004; Smith and Eppig, 2012; Tecott and Nestler, 2004). New technology: home cage recording With the development of advanced video-tracking software, computer power and data storage capacity, new behavioral technologies have emerged, allowing automated observation of animals in their home cage over long periods of time (de Visser et al., 2006; Goulding et al., 2008; Jhuang et al., 2010). Automation of observation allows repetitive, objective, and consistent measurement over days or even weeks, rather than minutes or hours. This substantially increases the scientific efficiency with respect to the researcher’s investment of time and effort. Continuous tracking has increased the behavioral throughput and decreased confounding factors like handling, and the frequent adjustment to novel environments that is required in batteries of classical behavioral tests. Furthermore, continuous recording allows investigation of multi-dimensional aspects of behavior like habituation, baseline and challenged behavior (de Mooij-van Malsen et al., 2009; de Visser et al., 2006; Kas et al., 2008), offering the possibility to study longitudinally the progression of