N-ethyl-N-nitrosourea mouse mutants in the dissection of behavioural and psychiatric disorders

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Abstract

Twin and adoption studies have consistently implicated genetics in the aetiology of psychiatric and behavioural disorders. The identification of the genes and molecular pathways that are associated with these traits using linkage studies has been difficult because psychiatric disorders are almost always non-mendelian, heterogeneous, involve multiple genetic loci and are influenced significantly by environmental factors. Mouse models that are based on intermediate signatures of psychiatric disease and pharmacological responsiveness hold promise as a complementary approach to dissecting the molecular basis of neurobehavioural disorders. This has been made possible by the development and refinement of gene targeting technologies and the use of super-efficient chemical mutagens. N-ethyl-N-nitrosourea (ENU) mutagenesis in the mouse, when coupled to a battery of sensitive behavioural screens, is an effective way of creating and identifying novel mouse behavioural mutants. Here, the concept of screening for ENU mutants is introduced while progress with two behavioural screens, an “anxiety” screen and a circadian screen, are presented. It is hoped that the study of mouse mutants that have arisen from these screens will provide new insights into the genetic basis of abnormal behaviour and that they might lead to the development of novel therapeutic compounds for human psychiatric disease.

Section snippets

The mouse as a model of psychiatric disease

Twin and adoption studies have implicated genetics in the aetiology of attention-deficit hyperactivity disorder, autism, bipolar disorder, depression, anxiety disorders, and schizophrenia Folstein and Rutter, 1977, Heston, 1966, McGuffin et al., 1996, McGuffin et al., 2003, Rietveld et al., 2003. Yet, despite the great advances that have been made in genetics in the last 50 years, the underlying causative factors remain largely unknown. Many groups have undertaken linkage studies to identify

Spontaneous mouse mutants

The study of spontaneous mouse mutants has led to important insights into gene function and disease phenotypes. An example of a spontaneous mutant relevant to psychiatric disorders is the mouse mutant coloboma. Coloboma was first identified because it exhibited head bobbing, hyperactivity and ocular dysmorphology (Hess et al., 1992), and was later shown to exhibit altered hippocampal plasticity (Steffensen et al., 1996) and neurodevelopmental abnormalities (Heyser et al., 1995). Genetic mapping

Gene targeting and reverse genetics

The use of homologous recombination in embryonic stems cells has permitted scientists to generate targeted mutations in specific genes. This approach is referred to as reverse genetics, as one begins with a particular hypothesis about the function of a gene and proceeds to study the phenotypic consequences of a targeted mutation in that gene. Gene targeting has revolutionised mouse behavioural genetics, playing a vital role in the understanding of the molecular basis of memory Reisel et al.,

Chemical mutagenesis and forward genetics

Forward genetics differs from reverse genetics, as it begins not with an interest in a particular gene, but in a phenotype (e.g., an abnormal behaviour). Relying on chemical mutagenesis to produce random mutations, forward genetics has been used extensively in several species to study gene function by establishing screens for various phenotypes Driever et al., 1996, Nusslein-Volhard and Wieschaus, 1980. Pioneering studies in screening specifically for behavioural mutants were introduced by

The case for ENU

The chemical mutagen N-ethyl-N-nitrosourea (ENU) has been established as the most appropriate mutagen for gene function studies in the mouse. This preference is based on several properties specific to ENU. Firstly, ENU induces mutations in rodents at a rate that is far greater than that of other mutagens. Several groups have estimated that phenotypic effects associated with every mammalian gene could be detected by screening only ∼1000 animals carrying ENU-induced mutations Coghill et al., 2002

Implementing an ENU mutagenesis screen

A number of institutes worldwide have established mouse ENU mutagenesis screens with the goal of modelling human disease and prescribing a function to each mammalian gene. Many groups have incorporated screens for behavioural phenotypes (See Table 1). The simplest form of screen that can be carried out is one for dominant ENU-induced mutations. With this approach, males are treated with ENU and subsequently mated with wild-type females (Fig. 1). Progeny from these crosses, which can each

Mouse mutants and anxiety

A number of mouse paradigms have been developed to explore the biological basis of anxiety. The most widely used are the open field, elevated plus maze and dark–light box (Rodgers, 2001). These devices all rely on the innate aversion of mice to open spaces and bright areas, which conflicts with their natural tendency to explore novel environments. An open field is a large square or round arena that is brightly lit and is normally white in colour. When placed in this environment, mice

Mouse mutants and circadian behaviour

Circadian behaviour incorporates the regulation of diverse processes such as the sleep–wake cycle, locomotor activity, temperature regulation, metabolism, water/food intake and levels of circulating hormones. This behaviour is normally synchronised to external environmental cues such as sunrise/sunset, a process known as entrainment. However, in the absence of such cues, these inherent rhythms persist. Disturbances in circadian parameters have been associated with a number of psychiatric and

Histology and pathology

Human studies have implicated abnormal brain pathology in psychiatric illness. For instance, reduced hippocampal volumes have been observed in schizophrenic patients Pegues et al., 2003, Velakoulis et al., 1999, glial cell reduction in the prefrontal cortex has been associated with mood disorders (Ongur et al., 1998) and asymmetry of the caudate nucleus has been linked to ADHD Filipek et al., 1997, Hynd et al., 1993. Given these observations, histological and pathological studies of mouse

Conclusion

The use of forward genetics approaches, such as ENU-mutagenesis screening, to identify mouse behavioural mutants is still in its infancy although results from systematic screens would suggest that this approach is valid in identifying and elaborating upon mammalian gene function. To date, no phenotypic screen has been unsuccessful in identifying novel molecular components and mechanisms. As more sophisticated phenotypic screening tools are developed, it should be possible to identify and study

Acknowledgments

DAK is supported by a Christopher Welch Scholarship for some of this work.

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