Mouse models for neurological disease

Reviews Mouse models for neurological disease Majid Hafezparast, Azlina Ahmad-Annuar, Nicholas W Wood, Sarah J Tabrizi, and Elizabeth MC Fisher The mouse has many advantages over human beings for the study of genetics, including the unique property that genetic manipulation can be routinely carried out in the mouse genome Most importantly, mice and human beings share the same mammalian genes, have many similar biochemical pathways, and have the same diseases In the minority of cases where these features not apply, we can still often gain new insights into mouse and human biology In addition to existing mouse models, several major programmes have been set up to generate new mouse models of disease Alongside these efforts are new initiatives for the clinical, behavioural, and physiological testing of mice Molecular genetics has had a major influence on our understanding of the causes of neurological disorders in human beings, and much of this has come from work in mice ␣-synuclein was found in a German family.2 Both these mutations were associated with more than 90% penetrance of the disease which implicated them in the aetiology of PD in these families ␣-synuclein was then found to be the main component of Lewy bodies;3 these intracytoplasmic inclusions are the pathological hallmark of PD, and this finding provided a direct link between ␣-synuclein and the aetiology of sporadic PD Since then, genetic variability in the ␣-synuclein gene has been found to be a risk factor for sporadic PD,4 and studies on ␣-synuclein in cell and mouse models has furthered our understanding of the pathogenesis.5 Thus, although inherited mutations in ␣-synuclein as a cause of PD are very rare,6 an understanding of the mechanisms by which these mutations cause PD in these families has provided important insights into the common sporadic form of the disease Lancet Neurology 2002; 1: 215–24 Diagnosis Every human illness, whether heritable, infectious, or caused by environmental agents, is modulated to a greater or lesser extent by the genetic make-up of the patient Genetic status contributes to disease susceptibility, including variables such as the age at onset and disease severity Therefore, to understand human diseases it is essential to understand their genetics This link is obvious for so-called “single-gene” disorders such as Huntington’s disease (HD), in which all cases are familial and due to mutation in one gene However, our need to unravel the genetics of disease also extends to the more common disorders, such as dementia or motorneuron degeneration, that may be thought of as largely “sporadic” Fairly uncommon familial forms of sporadic disorders are found in which several family members are affected, indicating that a common gene mutation is inherited within the family Human deficits can have quite variable clinical manifestations, and the discovery of disease-causing genes has widened the clinical diagnostic range of many neurological disorders For example, before the discovery of the Friedreich’s ataxia gene (FRDA),7 clinical diagnostic criteria were developed to help with diagnosis of the condition.8 Now, FRDA gene analysis is possible and the clinical phenotypic range of Friedreich’s ataxia has widened substantially to include patients who present with chorea and predominant spastic paraparesis; in addition, one of us (NWW unpublished) has seen a patient who had age at onset of 62 years In familial disorders, and even some sporadic disorders, a molecular diagnosis can now be made by sequencing of a patient’s DNA in known disease-causing genes to ascertain the mutation status of these genes If a mutation is found, the clinician can make a correct diagnosis and is in a much stronger position to suggest treatments and likely outcome The human condition Dissection of disease pathology Presymptomatic diagnosis These familial disorders illustrate why a genetic analysis of disease is a powerful approach for the elucidation of pathology and, ultimately, for design of treatments for both familial and sporadic forms of the same disorder For example, Parkinson’s disease (PD) is the second most common neurodegenerative disease with a prevalence of one in 350, and a lifetime risk of one in 40 Most cases are sporadic with unknown aetiology, but about 1% of cases are familial; study of these families has greatly furthered our understanding of the disease In 1997, a mis-sense mutation in the ␣-synuclein gene was found in a large kindred with autosomal dominant PD;1 a year later another mutation in Although knowledge of the genetic basis of disease can provide new certainty in diagnosis, it is also taking us into new uncertainty and dilemmas arising from presymptomatic THE LANCET Neurology Vol August 2002 MH, AA-A, SJT, and EMCF are in the Department of Neurodegenerative Disease, and NWW is in the Department of Molecular Pathogenesis, all at the National Hospital for Neurology and Neurosurgery, London, UK Correspondence: Prof Elizabeth Fisher, Department of Neurodegenerative Disease, National Hospital for Neurology and Neurosurgery, Queen Square, London WC1N 3BG,UK Tel +44 (0)20 7676 2037; fax +44 (0)20 7676 2180; email e.fisher@prion.ucl.ac.uk http://neurology.thelancet.com 215 For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease diagnosis These difficulties have been successfully tackled by the HD community.9–11 There are ethical issues with prenatal diagnosis for a disease such as HD, which is undoubtedly severe, but which has a likely age at onset of 50 years There are also obvious difficulties with presymptomatic diagnosis, which range from the protection of privacy of information through to challenges to a patient’s right not to know his or her disease status by interested insurance companies, for example The genetics of disease For the purposes of this review, we have categorised all inherited disorders into six groups: “single-gene” disorders; repeat-expansion disorders; imprinting disorders; chromosomal disorders; polygenic disorders (also called complex traits); and mitochondrial disorders (table 1).12–42 These categories are clinically relevant because they suggest who is at risk of disease and the likely severity of disease The categories may also determine when prenatal or presymptomatic diagnosis is possible (for example, in many single-gene disorders) or currently not possible (as in mitochondrial disorders43) These six different disease types also require technically different approaches to mouse modelling Single-gene disorders These disorders occur when an individual inherits one or two defective copies of a gene from his or her parents, or carries one or more new mutations that gives rise to disease In “dominant” disorders, such as HD or forms of Alzheimer’s disease (AD), the disease will manifest if the patient carries only one copy of the defective gene, along with one normal copy In “recessive” disorders, such as type I proximal spinal muscular atrophy (WerdnigHoffmann disease), both copies need to be defective for the patient to have the disorder.44 If the gene in question is carried on the X chromosome, the disorder will always manifest in males (for example, Duchenne muscular dystrophy and certain forms of colour blindness) because they have only one X chromosome and therefore one copy of all “X-linked” genes Generally, females will present with X-linked diseases only in the rare case of individuals with two copies of the defective gene (there are exceptions to this situation) Females with one copy of a defective X-linked gene generally will not have any symptoms and will be “carriers” of the disorder In fact, as in all biology, real life is never quite as simple as text-book explanations and the presentation of singlegene disorders is actually modified to some extent by the effects of other genes This point is important for understanding of how disease severity can vary in different individuals.45 For example, the same mutation can manifest with a different age at disease onset in the same family, because of the effects of modifier genes elsewhere in the genetic make-up of the patients, as has recently been shown in amyotrophic lateral sclerosis (ALS).46 Although the presence of these modifier genes may be obvious, very few of them have been identified so far Modifier genes will be important targets for therapy in the future Repeat-expansion disorders So far, these disorders all manifest as neurological diseases They arise from simple repeat sequences, in many cases just of three base-pairs, within the DNA of an individual gene, that expand (or contract) as the gene is inherited.47,48 When the number of the repeats reaches a certain threshold, the gene, and the associated protein, become defective Interestingly, the larger the number of repeats, the more severe the resulting disease In addition, since the repeats expand in each generation, the manifestation of disease becomes more severe in each generation Myotonic dystrophy was one of the first such diseases to be identified It arises from an increased number of CTG repeats in the DMPK gene.49,50 Healthy individuals have fewer than 30 copies of this repeat, patients with classic myotonic dystrophy generally have 50 to 300 copies, whereas severely affected individuals with “congenital” myotonic dystrophy have 2000 to 6000 repeats.51 Imprinting disorders Although half the chromosomes are inherited from the mother and half from the father, these sets of chromosomes are not identical They are “imprinted” in some reversible process (probably mainly methylation of the DNA), so the cell can recognise the maternal and paternal contribution of certain genes.52 For a few genes, both the maternally and paternally derived gene copies need to be present if the cell is Table Common or notable disorders of the nervous system that have a known genetic input and available mouse models Disorder Genetic disorder Alzheimer’s disease Single-gene disorder (some forms) Causative genes Recent reviews of mouse models Amyloid plaque protein (APP); presenilin (PS1); 12–21 presenilin (PS2) Amyotrophic lateral sclerosis Single-gene disorder (some forms) Superoxide dismutase (SOD1); alsin Deafness Single-gene, polygenic Several genes known 22–26 27,28 Down’s syndrome chromosome 21 Chromosomal disorder Multiple genes on human 29,30 Huntington’s disease Single-gene disorder, triplet repeat expansion Huntingtin gene 31–34 Parkinson’s disease Single-gene disorder (some forms) ␣-synuclein; parkin 35 Peripheral neuropathy Single-gene disorder (some forms) PMP22, myelin protein zero, connexin 32 and others 36–38 Prader-Willi syndrome Imprinting disorder Probably multiple genes 39 Prion diseases Single-gene disorder Prion (PRNP) 40–42 216 THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease to function properly.53 To have two copies of the maternally derived gene, for example, will not do.54 Many of the known imprinted genes are important for the development of the nervous system Disruption of these genes leads to several neurobehavioural disorders.55 One of these, Prader-Willi syndrome, is due to a deletion on the paternally derived chromosome 15.56,57 This deletion removes several genes that appear to be expressed on the paternal chromosome only and leaves the developing embryo with no messages from these loci 58 Chromosomal disorders These disorders arise when the number of chromosomes an individual has differs from the normal 23 pairs This state of “aneuploidy” is highly lethal The only “monosomy” that survives to term is of the X chromosome, which manifests as Turner’s syndrome Of the trisomies, individuals with extra X and Y chromosomes have been described, as has trisomy 13 (Patau’s syndrome), 18 (Edwards’ syndrome) and 21 (Down’s syndrome) Trisomy of all other chromosomes is lethal prenatally These syndromes probably arise because of altered gene dosage—three copies instead of the normal two—of an unknown number of genes.59 Many other types of more subtle chromosome rearrangements exist and cause disease because of gene-dosage effects.60 Polygenic traits Polygenic traits, or complex traits, arise from the cumulative effect of several different genes These disorders are the cause of the majority of human health-care issues in the more developed countries, and most involve interactions with environmental factors Schizophrenia, stroke, and Tourette’s syndrome, for example, have a genetic component from the interaction of several genes in the patient’s genome that can confer increased risk.61–63 Mitochondrial disorders Finally, when considering genetics and disease we must remember not all our genes are on the nuclear chromosomes; the mitochondria have their own chromosomes.64 These chromosomes contain genes that encode proteins required for mitochondrial function.65 If mutations arise in the mitochondrial genome, they can lead to complex diseases that manifest depending on the number of defective mitochondria Because nerves and muscles are high-energy turnover tissues, they are typically affected by mitochondrial diseases.66–70 Mutations in mitochondrial genes cause a range of neurological disorders such as the syndromes of mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes, and myoclonic epilepsy and ragged red fibres.64 Why model diseases in mice? There are many experiments that can be done in mice that society would deem to be unacceptable in human beings For example, new therapies, including gene therapies, are tested on animal models before the human population.71 In addition, all stages of disease pathology can be investigated in mouse models, whereas in human beings we generally THE LANCET Neurology Vol August 2002 only have access to tissue from endstage disease; the intermediate processes, for example of neurodegeneration, may be obscure.12,31 Animal models also give us access to the developing embryo, so we can investigate the routes giving rise to congenital or perinatal disease.72 Environmental factors, such as diet, can also be rigorously investigated and controlled.73 For genetic investigations of disease, the mouse is the mammal of choice Although human beings and mice diverged from a common ancestor about 80 million years ago, we share almost exactly the same set of genes, with very similar DNA sequences, and produce proteins that carry out the same functions via the same biochemical pathways Thus, a genetic disease of the mouse nervous system is almost certainly going to be caused by a defect in a gene that acts in the human nervous system So we can cross-reference between species, especially because both the mouse and human genome sequences will be completed shortly This similarity has practical applications when we look for disease genes Gene hunting usually entails the collection of family pedigrees In late-onset disease, this task is commonly impossible because the parents of the affected individual are dead and the children are of unknown disease status Mice become sexually mature at about weeks old, so three-generation pedigrees can be generated in a matter of months In addition, some human “matings” are more informative than others, but with mice we can define the matings for a specific investigation Genetics is a statistically based science and depends on the collection of large numbers of individuals from one generation Male mice can easily produce hundreds of offspring, which keeps the geneticists (and presumably the mice) happy Finally, much human disease is genetically heterogeneous, which confounds our attempts at gene hunting This means that what appears to be the same disease in fact derives from mutation in two different genes (that commonly act within the same biochemical pathway) This is the case with AD, which can be caused by single mutations in one of three known genes, presenilin (PS1), presenilin (PS2), and the amyloid plaque precursor protein (APP).74 Since a human family is smaller than geneticists find useful, large numbers of families affected by one disease tend to be collected for single genetic studies When the results of these studies are analysed together for gene hunting, they can be difficult to interpret because more than one defective gene is segregating in the different families In mice, huge families can be bred from one single founder animal As a result, researchers know they are looking for only one gene and can pool results and even DNA samples.75 Why use mice rather than other mammals? For geneticists who study mammalian disease, mice have two unique advantages over all other mammals, including human beings The first is that “inbred strains” of mice have been bred since the 1900s and are all genetically identical This identity is achieved by brother–sister mating over 20 generations; there are currently more than 450 such inbred strains.76 So all genetically determined traits, such as coat colour or response to drugs, will be identical within an http://neurology.thelancet.com 217 For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease inbred line, but may differ between inbred lines Differences between mouse strains have been extensively studied in certain specialist areas; for example, in cerebral ischaemia there are significant differences in blood-vessel structure, thermoregulation, blood flow, vascular responsiveness, and vulnerability in different strains of mice.77 The Mouse Genome Database, which is curated by the Jackson Laboratory (Maine, USA), holds an online detailed list of the physiology, disease susceptibility, and other characteristics of each inbred strain (http://www.informatics.jax.org) We can tease out the effects of environment and genes by comparing the phenotypes and genotypes of the different inbred strains We can also use the genetic differences between the strains to find DNA sequences that affect phenotype The second unique advantage of mice is that we can manipulate the mouse genome to create specific genetic changes Extra genes can be added to create “transgenic” mice, and in fact these transgenic studies illustrate how closely related mice are to human beings; in almost all cases a human gene is fully functional in the mouse and can replace the normal endogenous gene The basis of much modelling of human disease comes from transgenic mice that have been made to carry mutated human genes Other techniques allow us to remove or alter gene sequences, to create gene-targeted mice (known as “knock-out” or “knock-in” mice) The Jackson Laboratory also maintains an online database of transgenic (including gene-targeted) mice, and gives a helpful list of the human diseases modelled by such mice (http://tbase.jax.org) Clinicians often say that the disease they are interested in is not exactly modelled in a mouse Disease manifestation in mice depends on several factors, including the genetic background of the animal, so breeding the mutation onto a different inbred line may alter the phenotype of the mouse However, mice are not minipeople There are very obvious differences such as lifespan, fertility, and size There are also some differences in physiology For certain types of research, such as reproductive biology, these differences are critical For working out the links between genes and disease, what is more important is that the biochemical pathways are the same in human beings and mice, which, largely, they are Mice that not model human disease There are cases in which mouse models give rise to novel phenotypes because the biochemical pathways are unexpectedly different from those in human beings, but even knowledge of these can be useful.78 A good example is the gene-targeting experiments that were done to model the human lysosomal storage disorders Tay-Sachs disease and Sandhoff’s disease in the mouse.79–81 These diseases result from mutations in the genes HEXA (Tay-Sachs) and HEXB (Sandhoff’s disease) that code for ␣ and ␤ subunits of the enzyme hexosaminidase, which catabolyses GM2 ganglioside Both disorders result in severe early-onset neurodegeneration and early death When the Hexa and Hexb mouse genes were knocked out to model the human disorders, both types of mouse showed the expected accumulation of GM2 gangliosides in the brain, but the Hexa-gene-targeted mice had no obvious neurological or behavioural deficits However, the Hexb-gene-targeted mice had rapid, progressive, and fatal neurodegeneration Biochemical studies showed that the mouse sialidase enzyme was much more important in this catabolic pathway than previously thought 79–81 These findings illustrate that even when a mouse does not reproduce a human condition we may be able to find potential therapeutic targets, such as sialidase for the treatment of patients with Tay-Sachs disease Clinical and pathological testing of mice With the recent increase in the use of mouse models, the need for standard protocols for the qualitative and Table The SHIRPA protocol Primary Muscle/lower motor neuron Body position, gait, positional passivity, wire manoeuvre, righting reflex, motor performance, spontaneous activity, tail elevation, visual placing, limb tone, passivity, balance, locomotor activity, limb position, body tone, abdominal tone, urination, defecation Spinocerebellar Body position, gait, righting reflex, motor performance, tail elevation, visual placing, limb tone, balance, locomotor activity, limb position, body tone, abdominal tone, grip strength Sensory Transfer arousal, touch escape, corneal reflex, analgesia, gait, visual placing, toe pinch, limb position, pinna reflex, righting reflex Neuropsychiatric Body position, transfer arousal, startle response, body tone, fear, anxiety, learning/memory, spontaneous activity, locomotor activity, touch escape, righting reflex, irritability, vocalisation, bizarre behaviour, food/water intake, positional passivity, catalepsy, aggression, PPI Autonomic Palprebral closure, tail elevation, temperature, heart rate, piloerection, skin colour, food/water intake, urination, startle response, salivation, respiration rate, defecation Clinical chemistry Haematology: white-cell count, red-cell count, haemoglobin, platelets, three part differential Hepatic: alanine and aspartate aminotransferases, ␥-glutamyltranspeptidase total protein, albumin, total bilirubin Renal: sodium, potassium, chloride/bicarbonate, creatine, urea Diabetes: glucose Bone: alkaline phosphatase, total calcium, inorganic phosphate Lipid: total cholesterol, HDL cholesterol, triglyceride Secondary Extended locomotor activity, balance and coordination, histology, food/water intake, analgesia Tertiary Sophisticated behavioural tests, anxiety and learning and memory, nerve conduction studies, fMRI, and other neuroimaging Details of this three stage hierarchical phenotype assessment procedure can be found at http://www.mgu.har.mrc.ac.uk 218 THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease quantitative description of phenotype has become apparent Because accurate assessment is essential, several tests have become more standardised and portable between laboratories, and helpful new articles have come from mouse-behaviour researchers.82–87 One comprehensive screening method, the SHIRPA (Smithkline Beecham, MRC Harwell, Imperial College, the Royal London hospital phenotype assessment) protocol, has been devised for high-throughput phenotype assessment of mice, and is in use in several laboratories worldwide.88–90 The SHIRPA screening method is hierarchical and has three stages, each comprising a series of individual tests that quantify a mouse’s performance in a wide range of tasks (table 2) These simple tests are designed to detect defects in, for example, the functions of lower and upper motor neurons, muscle, sensory neurons, and behaviour (http://www.mgu.har.mrc.ac.uk).91 The standardisation of SHIRPA makes possible the direct comparison of animals over a time period and between groups The primary and secondary stages of the SHIRPA protocol assess the behaviour and pathology of the mice, whereas the tertiary screening is tailored to test specifically the system of interest, such as CNS function A full description of the protocol can be found on the website run by the Medical Research Council Mammalian Genetics Unit at Harwell, UK http://www.mgu.har.mrc.ac.uk/ mutabase/shirpa_summary.html) Approaches to mouse modelling There are two approaches to produce mouse models of human disease The first, the genotype-driven approach, depends on knowing the gene of interest, and then manipulating this gene in the mouse to create the appropriate model The second, the phenotype-driven approach, is not gene dependent, but uses standard genemapping and cloning techniques to identify the causal genetic change in an interesting phenotype Genotype-driven mouse modelling Transgenic mouse lines, created by the addition of extra genes (which has been possible since the early 1980s), are generally used to model dominant disorders, because the mutation gives rise to a phenotype irrespective of the two normal endogenous mouse genes present (figure 1) The extra gene can be of mouse or human origin; for example, transgenic mice carrying mutant human SOD1 genes succumb to progressive motor-neuron degeneration that mimics much of ALS.22 In many settings, more than one transgenic mouse line can be created for an individual disease—in the case of the SOD1 transgenics, there are five different lines each with a different human mutation from a patient with ALS.23,92–94 The histopathology of each mouse line is subtly different, and ultimately these differences will help to elucidate the disease process.22,24 When transgenes insert, they can disrupt the function of genes at or near the site of insertion This disruption can have fortuitous effects For example, when a transgene was inserted into an imprinted stretch of the mouse genome that THE LANCET Neurology Vol August 2002 Transgene (human or mouse) is injected into a fertilised mouse egg The gene incorporates into a mouse chromosome and the transgenic embryos are placed into a foster mother, to develop to term Figure An overview of creating transgenic mice is homologous to the human Prader-Willi syndrome and Angelman syndrome regions, the transgene caused a small deletion that models these diseases, when it is either paternally or maternally inherited.95 Gene-targeted, or “knock-out”, mouse lines, in which the genome is manipulated to remove or alter DNA sequences so a gene cannot function, are mostly used to model recessive disorders This technique, which started in the late 1980s, came about through a fusion of molecular biological research to target specific changes to individual DNA sequences, in combination with working with embryonic stem cells in culture (figure 2) The embryonic stem cells are genetically modified to put the specific changes into the genome This can be a very inefficient process in which literally tens of millions of cells are treated to induce a particular change to one small region of DNA The few cells in which this process has occurred successfully are then selected by use of molecular-biological techniques The targeted embryonic stem cells are then injected into recipient blastocysts, and the chimeric blastocysts are implanted into a foster mother The embryonic stem cells have the ability to form any cell type in the body, including the gonads, sperm, and oocytes The chimeric mice that result from this procedure are bred; if the embryonic stem cells have contributed to the germline, a new strain of mice can be created that carries the targeted mutation There are many examples of gene-targeted knock-out mice that are good models for human neurological disorders These include mouse models of autosomal recessive storage disorders in which enzyme function is diminished or absent (for example, Gaucher’s disease96) and models of X-linked single-gene disorders, such as fragile X syndrome.97 More recently, the same gene-targeting method has been used to create “knock-in” mice, in which gene regions are switched for other sequences These animals help us to understand gene function In human beings, Friedreich’s ataxia is caused by expanded GAA repeats in the frataxin gene, which leads to loss of the protein A knock-in mouse that models this disorder has been created, in which 230 copies of the GAA repeat have been placed in the mouse frataxin gene.98 Creation of transgenic or gene-targeted mice is always an experiment, and may not be straightforward, because http://neurology.thelancet.com 219 For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease A Mouse embryonic stem (ES) cells are grown in culture The ES cells are genetically modified by use of molecular biology techniques, to produce specific changes in DNA Cells with the correct DNA targeting are selected These cells are grown in culture and then injected into recipient blastocytes (very early embryos) Chimeric embryos are placed into a foster mother to develop to term Chimeric pups are born Gene targeted mouse If ES cells have contributed to germ line, gene targeted mice will be produced when the chimeric mice are mated B some mutations are lethal at specific embryonic stages or soon after birth Therefore, the role of the gene in later stages of development and during adulthood cannot be studied, as is the case for NF1 (neurofibromatosis 1) knockout mouse embryos.99 As a result, strategies have been developed to create “conditional mutants”, in which mutant genes can be expressed or deleted at different times or in different tissues Various systems exist for achieving this temporal or spatial control of gene expression For example, genes can be activated by placing the transgene under the control of a regulatory sequence that responds to tetracycline If tetracycline is included in the animal’s diet, the gene will be activated at that time This approach has been used very successfully to help understand HD mouse model pathology.100 For deletion of a gene to create a null mutation, the Cre–loxP system has become widely used In this case, the gene of interest is genetically manipulated so that it is flanked by a pair of DNA sequences called loxP sites These are recognition sites for an enzyme called Cre, which can cut out the DNA between the two sites If a mouse is transgenic for the Cre gene, the gene flanked by loxP sites will be deleted in that tissue In addition, reseachers can ensure that the Cre enzyme is expressed in a tissue-specific manner (for example, only in the brain), a technique that has allowed NF1 to be deleted in adult brain only99 or the prion gene to be similarly disrupted in brains of mice more than weeks old.101 Another type of disease modelling uses gene-targeting methods to create mouse models of the chromosomal disorders.102–104 By use of a series of quite demanding techniques, whole chromosomes or large regions of chromosomes can be transferred into mouse embryonic stem cells These are then injected into host blastocysts, to create chimeric embryos and pups Again, if the transchromosomal embryonic stem cell line has contributed to the gonads, the pups will breed to produce a new line of mice that carry an extra chromosome This approach is being used to produce, for example, mouse models of Down’s syndrome by placement of human chromosome 21 or large regions of the chromosome into mice.102,103 Mouse models with naturally occurring chromosomal aberrations have also been described These are useful models for some human syndromes, including imprinting disorders, because many imprinted regions are the same in human beings and mice (as in, for example, a mouse model of Angelman’s syndrome105) Mitochondrial-disease models Figure (A) An overview of creating gene-targeted mice (B) A chimeric mouse that was generated by injecting mouse ES cells (black pigment) into a host blastocyst (white pigment; photograph courtesy of Dr Aideen O’Doherty) 220 So far, modelling of mitochondrial mutations has been more difficult than working with nuclear DNA Currently, two approaches are mainly used Both involve cell fusion of cytoplasm-carrying mutant mitochondria from one cell line with embryonic stem cells or single-cell embryos This stage is followed by reimplantation to create chimeric mice.106–108 Mouse models of mitochondrial disease are beginning to become available, which presents exciting opportunities for furthering our knowledge of these intractable disorders.109 THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease For the vast majority of diseases, particularly complex disorders such as multiple sclerosis or stroke, we not know the causative genes So, from a clinician’s viewpoint, the phenotype approach to mouse modelling may be more relevant because, by definition, the mice have a phenotype of interest Furthermore, once a causative gene is identified in the mouse, we can use the human-mouse DNA sequence data to check the human homologue for mutations in patients with similar phenotypes For example, the shaker mouse was found to have a mutation in an unconventional myosin; screening of human samples found the mutation in samples from patients with Usher syndrome (type 1b) or with non-syndromic deafness.110,111 the affected genes In addition, up to 10% of the F1 progeny are aged and will be SHIRPA-tested at the ages of 1, 3, and months (equivalent to late middle-age for a mouse) to identify late-onset phenotypes Details of this mutagenesis programme can be found at http://www.mgu.har.mrc.ac.uk All mutants produced in this programme are available to other researchers free of charge Almost 200 new neurological or behavioural mutants have already been described,75,91,121,122 including two that have defects in the Pmp22 gene PMP22 is defective in human peripheral neuropathies, and one of the new mutants has exactly the same amino acid change that causes the severe human peripheral neuropathy Déjérine-Sottas syndrome; this mouse appears to be a good model of the disorder.75 Use of mutagens Approaches for elucidation of complex traits Some mouse models have been identified serendipitously when spontaneous mutations occurred; for example the neuromuscular degeneration (Nmd) mouse, which has an autosomal recessive mutation in the Smbp2 gene that results in progressive hind-limb impairment due to spinal muscular atrophy.112,113 But these models are limited, and we cannot rely on spontaneous mutations because they arise infrequently Another option is to introduce mutations into the genome randomly by means of mutagens such as chemicals, radiation, or viruses, and then to assay and collect the resulting phenotypes for research on those of interest ENU mutagenesis in the mouse has been used to identify single-gene traits However, new schemes are being set up (borrowing ideas from fruit-fly genetics) to find mutations that modify such traits or affect known pathways that not produce a readily identifiable phenotype These are “sensitised” screens in which mice with known mutations in biochemical pathways of interest become part of the breeding scheme These mutations can enhance or suppress new phenotypes induced by ENU treatment.45,123 Other, more traditional, approaches for identification of modifier genes or genes that underlie complex traits are based on crossing mouse strains with different phenotypes, or on assaying the physiological or disease status of the progeny and then looking for correlation with the inheritance of individual regions of the genome.124–126 The problem of finding modifiers, or quantitative trait loci, is one of the current big challenges of molecular genetics, because some of these genes may have an extremely important role in biochemical pathways, and may be drug targets However, the methods are not quite routine This type of research illustrates yet again the similarity between mice and human beings, and how data from one species can be used to understand the other A clear example comes from studies of cystic fibrosis, in which wide phenotypic variation exists in families with identical CFTR mutations One of these variations is the presence of an intestinal phenotype, meconium ileus, in 15–20% of patients with cystic fibrosis This phenotype was also seen in a knockout mouse model of cystic fibrosis and subsequently a modifier gene (Cfm1) was mapped to the mouse chromosome 7, which is homologous to human chromosome 19.127 On the basis of this information, Zielenski and co-workers127,128 went on to map a cysticfibrosis modifier locus for meconium ileus to the predicted region of human chromosome 19q13 Another approach for identifying genes that act in the same biochemical pathways involves the crossing of animals with different mutations to see if the individual phenotypes are altered If they are, there is an interaction of some sort between the genes In the past this approach, which was first used in fruit-fly experiments, was largely not feasible because of the dearth of mouse mutants Now that the mammalian genome is being comprehensively mutagenised, increasing Phenotype-driven mouse modelling ENU mutagenesis One of the most widely used agents for induction of mutation is N-ethyl-N-nitrosourea (ENU), which is highly potent and induces mainly single base-pair changes in DNA When an appropriate dose of the chemical is used, one mutation per locus per 1000 gametes is achieved on average.114 Because of the efficiency and simplicity in the use of ENU, several major projects have been launched to extend the availability of mouse models for human disease A list of these centres can be found in the review by Brown and Balling.115 Some of these projects aim to generate mutants from specific regions of the genome,116 and some target the entire genome for dominant91,117 or recessive118 mutations Some projects have produced mutants with phenotypes that affect only specific biological systems such as the immune system,119 whereas other researchers are interested in collecting series of new mutants with deficits affecting the visual system120,121 or the ear.122 One such project, with which we have been directly involved, is the ENU Mutagenesis Program at the MRC Mammalian Genetics Unit in Harwell, UK This project started in early 1998 and consists of two phases The aim of phase I was to produce, within years, large numbers of new mouse mutants—as defined by a SHIRPA analysis—that carry dominantly inherited traits that could model human disease.91 The project was set up in such a way that breeding of mutant mice for initial gene-mapping studies was routine Sperm from mutant mice were frozen to create an archive for long-term storage Phase II of the project, which is underway, involves selection of the phenotypes for further characterisation, and detailed mapping and identification of THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com 221 For personal use Only reproduce with permission from The Lancet Publishing Group Review Mouse models for neurological disease numbers of these experiments are being published and they can be very powerful For example, spinocerebellar ataxia type (SCA1) is caused by increased numbers of the repeat CAG within the SCA1 gene Work in Drosophila melanogaster highlighted modifier genes that act on protein folding and clearance.129 This information prompted a study in which mice that overexpress the heat-shock protein, HSP70, were crossed with mice that express a mutant human SCA1 gene; the double-transgenic animals showed reduced neuronal degeneration of the Purkinje cells, which suggested that the HSP70 chaperone has an effect on neuronal survival.130 Cummings and co-workers130 propose that upregulation of chaperone activity may therefore offer a possible route for suppression of neuronal cell death in this disorder The future The isolation of a gene that is involved in the aetiology of a disorder is just the start for understanding the underlying mechanisms of the disease Many questions remain What is the function of the gene product? How does a mutation in this gene disrupt its function? Why does disrupted function lead to pathology, and via which pathways? When disease processes start? The next set of questions turn to therapies Can we target conventional therapies to help alleviate or cure the disorder, given our knowledge of the affected cellular processes? Is gene therapy a feasible approach and, if so, how can we test it so we get appropriate temporal and spatial gene expression? Mouse models can help answer every one of these questions And as the number of models increases, and as we refine our ability to manipulate the genome, we will be able to undertake more sophisticated investigations into disease The pace of investigation is changing radically as the range of technologies available to us increases; for example, through microarray profiling (which gives us temporal and spatial patterns of expression of all the genes within a tissue) and proteomics (which allows us the high-throughput analysis of protein expression and variation) With these tools to help us understand mouse models, we will be able to identify more biochemical and cellular pathways to target in our therapeutic approaches Moreover, because there are variations in drug response and side-effects, which are genetically determined, mouse models can play a significant part in elucidating the interaction between genes and drugs (pharmacogenomics) However, there are outstanding questions that still need to be addressed Although large-scale production of mutant mice has become a reality, there is still a need for the References Polymeropoulos MH, Lavedan C, Leroy E, et al Mutation in the alpha-synuclein gene identified in families with Parkinson’s disease Science 1997; 276: 2045–47 Kruger R, Kuhn W, Muller T, et al Ala30Pro mutation in the gene encoding alpha-synuclein in Parkinson’s disease Nat Genet 1998; 18: 106–08 Spillantini MG, Schmidt ML, Lee VM, Trojanowski JQ, Jakes R, Goedert M Alpha-synuclein in Lewy bodies Nature 1997; 388: 839–40 Farrer M, Maraganore DM, Lockhart P, et al Alphasynuclein gene haplotypes are associated with 222 Search strategy Data for this article were identified from the authors’ knowledge of the subject, personal files, and textbooks A Pubmed search was carried out with the terms “mouse model”, and individual neurological disorders of interest, such as “Prader-Willi syndrome”, “mitochondrial disease”, “mitochondrial disorders”, “deafness”, and with the terms “Cre loxP”, “mutagenesis”, “ENU” and “N-ethyl-N-nitrosourea” Only papers and books published in English were reviewed development of more robust and faster tests for accurate and more specialised assessment of phenotypes Many of these, such as neuroimaging for the mouse, are now available, especially for study of the CNS.131 Nevertheless, we still need to know more about the mouse—its physiology, anatomy, histology, and, importantly, behaviour.132 A greater understanding of mouse behaviour and possible correlations with the human condition, would enable us to design better tests for identification of mouse models of complex behavioural disorders such as schizophrenia Several government agencies, particularly in the USA, have started initiatives to support the study of mouse pathology and mouse behavioural testing, and these skills are clearly essential to capitalise on the potential of mouse models We note that of paramount importance in all the investigations undertaken, especially since the use of mouse models is likely to increase in the foreseeable future, is the welfare of the mice themselves.133 A worldwide scientific effort has resulted in the production of valuable mouse mutants These mutants model human disease largely because our biochemistry is so similar Mice have become essential for teasing out the complex processes of disease pathology, and for directing rational therapeutic approaches With the availability of the mouse genome sequence, and with new techniques for the creation of more subtle models, the mouse is definitely here to stay Authors’ contributions MH, EMCF, and SJT wrote the first draft of the review AA-A did the literature search NWW critically evaluated the paper Conflict of interest EMCF is a member of the ENU Mouse Mutagenesis Programme Consortium (based at the MRC Mammalian Genetics Unit, Harwell) Role of the funding source MH and EMCF are funded by the Motor Neurone Disease Association; AA-A is funded by the University of Malaya and the Violet M Richards Charitable Trust; NWW, SJT, and EMCF are funded by the UK Medical Research Council; EMCF is funded by the Wellcome Trust; Sarah Tabrizi is a UK Department of Health Clinician Scientist None of these funding bodies had a role in the preparation of this review or in the decision to submit for publication Parkinson’s disease Hum Mol Genet 2001; 10: 1847–51 Goedert M Alpha-synuclein and neurodegenerative diseases Nat Rev Neurosci 2001; 2: 492–501 Vaughan J, Durr A, Tassin J, et al The alphasynuclein Ala53Thr mutation is not a common cause of familial Parkinson’s disease: a study of 230 European cases Ann Neurol 1998; 44: 270–73 Campuzano V, Montermini L, Molto MD, et al Friedreich’s ataxia: autosomal recessive disease caused by an intronic GAA triplet repeat expansion Science 1996; 271: 1423–27 Harding AE Friedreich’s ataxia: a clinical and genetic study of 90 families with an analysis of early diagnostic criteria and intrafamilial clustering of clinical features Brain 1981; 104: 589–620 Harper P Huntington’s disease, 2nd edn London: WB Saunders, 1996 10 Wexler N Huntington’s disease In: Pence GE, ed Classical cases in medical ethics Accounts of cases that have shaped medical ethics, 3rd edn Boston: McGraw-Hill, 1999: 388–416 11 Almqvist EW, Bloch M, Brinkman R, Craufurd D, Hayden MR A worldwide assessment of the frequency of suicide, suicide attempts, or psychiatric hospitalization after predictive testing for THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com For personal use Only reproduce with permission from The Lancet Publishing Group Mouse models for neurological disease Huntington disease Am J Hum Genet 1999; 64: 1293–304 12 Janus C, Westaway D Transgenic mouse models of Alzheimer’s disease Physiol Behav 2001; 73: 873–86 13 Bornemann KD, Staufenbiel M Transgenic mouse models of Alzheimer’s disease Ann N Y Acad Sci 2000; 908: 260–66 14 Chapman PF, Falinska AM, Knevett SG, Ramsay MF Genes, models and Alzheimer’s disease Trends Genet 2001; 17: 254–61 15 Duff K Transgenic mouse models of Alzheimer’s disease: phenotype and mechanisms of pathogenesis Biochem Soc Symp 2001; 67: 195–202 16 Emilien G, Maloteaux JM, Beyreuther K, Masters CL Alzheimer disease: mouse models pave the way for therapeutic opportunities Arch Neurol 2000; 57: 176–81 17 Hock BJ Jr, Lamb BT Transgenic mouse models of Alzheimer’s disease Trends Genet 2001; 17: S7–12 18 Masliah E, Rockenstein E Genetically altered transgenic models of Alzheimer’s disease J Neural Transm Suppl 2000; 59: 175–83 19 Price DL, Wong PC, Markowska AL, et al The value of transgenic models for the study of neurodegenerative diseases Ann N Y Acad Sci 2000; 920: 179–91 20 Richardson JA, Burns DK Mouse models of Alzheimer’s disease: a quest for plaques and tangles ILAR J 2002; 43: 89–99 21 Van Leuven F Single and multiple transgenic mice as models for Alzheimer’s disease Prog Neurobiol 2000; 61: 305–12 22 Nicholson SJ, Witherden AS, Hafezparast M, Martin JE, Fisher EM Mice, the motor system, and human motor neuron pathology Mamm Genome 2000; 11: 1041–52 23 Gurney ME, Pu H, Chiu AY, et al Motor neuron degeneration in mice that express a human Cu,Zn superoxide dismutase mutation Science 1994; 264: 1772–75 24 Newbery HJ, Abbott CM Of mice, men and motor neurons Trends Mol Med 2002; 8: 88–92 25 Deng HX, Siddique T Transgenic mouse models and human neurodegenerative disorders Arch Neurol 2000; 57: 1695–702 26 Doble A, Kennel P Animal models of amyotrophic lateral sclerosis Amyotroph Lateral Scler Other Motor Neuron Disord 2000; 1: 301–12 27 Ahituv N, Avraham KB Auditory and vestibular mouse mutants: models for human deafness J Basic Clin Physiol Pharmacol 2000; 11: 181–91 28 Steel KP Inherited hearing defects in mice Annu Rev Genet 1995; 29: 675–701 29 Dierssen M, Fillat C, Crnic L, Arbones M, Florez J, Estivill X Murine models for Down syndrome Physiol Behav 2001; 73: 859–71 30 Reeves RH, Baxter LL, Richtsmeier JT Too much of a good thing: mechanisms of gene action in Down syndrome Trends Genet 2001; 17: 83–88 31 Menalled LB, Chesselet MF Mouse models of Huntington’s disease Trends Pharmacol Sci 2002; 23: 32–39 32 Sipione S, Cattaneo E Modeling Huntington’s disease in cells, flies, and mice Mol Neurobiol 2001; 23: 21–51 33 Shelbourne PF Of mice and men: solving the molecular mysteries of Huntington’s disease J Anat 2000; 196: 617–28 34 Heintz N, Zoghbi HY Insights from mouse models into the molecular basis of neurodegeneration Annu Rev Physiol 2000; 62: 779–802 35 Barbieri S, Hofele K, Wiederhold KH, et al Mouse models of alpha-synucleinopathy and Lewy pathology: alpha-synuclein expression in transgenic mice Adv Exp Med Biol 2001; 487: 147–67 36 Bennett CL, Chance PF Molecular pathogenesis of hereditary motor, sensory and autonomic neuropathies Curr Opin Neurol 2001; 14: 621–27 37 Garbay B, Salles J, Knoll A, et al Trembler as a mouse model of CMT1A? Ann N Y Acad Sci 1999; 883: 262–72 38 Martini R Animal models for inherited peripheral neuropathies: chances to find treatment strategies? J Neurosci Res 2000; 61: 244–50 39 Nicholls RD, Ohta T, Gray TA Genetic abnormalities in Prader-Willi syndrome and lessons from mouse models Acta Paediatr Suppl 1999; 88: 99–104 40 Brandner S, Klein MA, Frigg R, et al Neuroinvasion of prions: insights from mouse models Exp Physiol 2000; 85: 705–12 41 Scott MR, Supattapone S, Nguyen HO, DeArmond SJ, Prusiner SB Transgenic models of prion disease Arch Virol Suppl 2000; 113–24 42 Telling GC Prion protein genes and prion diseases: THE LANCET Neurology Vol August 2002 studies in transgenic mice Neuropathol Appl Neurobiol 2000; 26: 209–20 43 Thorburn DR, Dahl HH Mitochondrial disorders: genetics, counseling, prenatal diagnosis and reproductive options Am J Med Genet 2001; 106: 102–14 44 Sendtner M Molecular mechanisms in spinal muscular atrophy: models and perspectives Curr Opin Neurol 2001; 14: 629–34 45 Nadeau JH Modifier genes in mice and humans Nat Rev Genet 2001; 2: 165–74 46 Giess R, Holtmann B, Braga M, et al Early onset of severe familial amyotrophic lateral sclerosis with a SOD-1 mutation: potential impact of CNTF as a candidate modifier gene Am J Hum Genet 2002; 70: 1277–86 47 Cummings CJ, Zoghbi HY Trinucleotide repeats: mechanisms and pathophysiology Annu Rev Genomics Hum Genet 2000; 1: 281–328 48 Fischbeck KH Polyglutamine expansion neurodegenerative disease Brain Res Bull 2001; 56: 161–63 49 Thornton C The myotonic dystrophies Semin Neurol 1999; 19: 25–33 50 Zeesman S, Whelan DT, Carson N, et al Parents of children with spinal muscular atrophy are not obligate carriers: carrier testing is important for reproductive decision-making Am J Med Genet 2002; 107: 247–49 51 Baraitser M The genetics of neurological disorders, 3rd edn Oxford: Oxford University Press, 1997 52 Paulsen M, Ferguson-Smith AC DNA methylation in genomic imprinting, development, and disease J Pathol 2001; 195: 97–110 53 Davies W, Isles AR, Wilkinson LS Imprinted genes and mental dysfunction Ann Med 2001; 33: 428–36 54 Boer H, Holland A, Whittington J, Butler J, Webb T, Clarke D Psychotic illness in people with PraderWilli syndrome due to chromosome 15 maternal uniparental disomy Lancet 2002; 359: 135–36 55 Weinstein LS The role of tissue-specific imprinting as a source of phenotypic heterogeneity in human disease Biol Psychiatry 2001; 50: 927–31 56 Hanel ML, Wevrick R The role of genomic imprinting in human developmental disorders: lessons from Prader-Willi syndrome Clin Genet 2001; 59: 156–64 57 Nicholls RD, Knepper JL Genome organization, function, and imprinting in Prader-Willi and Angelman syndromes Annu Rev Genomics Hum Genet 2001; 2: 153–75 58 Runte M, Huttenhofer A, Gross S, Kiefmann M, Horsthemke B, Buiting K The IC-SNURF-SNRPN transcript serves as a host for multiple small nucleolar RNA species and as an antisense RNA for UBE3A Hum Mol Genet 2001; 10: 2687–2700 59 Hernandez D, Fisher EM Mouse autosomal trisomy: two’s company, three’s a crowd Trends Genet 1999; 15: 241–47 60 Stankiewicz P, Lupski JR Genome architecture, rearrangements and genomic disorders Trends Genet 2002; 18: 74–82 61 Comings DE Clinical and molecular genetics of ADHD and Tourette syndrome: two related polygenic disorders Ann N Y Acad Sci 2001; 931: 50–83 62 Hademenos GJ, Alberts MJ, Awad I, et al Advances in the genetics of cerebrovascular disease and stroke Neurology 2001; 56: 997–1008 63 Prasad S, Semwal P, Deshpande S, Bhatia T, Nimgaonkar VL, Thelma BK Molecular genetics of schizophrenia: past, present and future J Biosci 2002; 27 (suppl 1): 35–52 64 Schapira AHV, DiMauro S Mitochondrial disorders in neurology Boston: Butterworth-Heinemann, 2002 65 Shanske AL, Shanske S, DiMauro S The other human genome Arch Pediatr Adolesc Med 2001; 155: 1210–16 66 DiMauro S, Andreu AL, Musumeci O, Bonilla E Diseases of oxidative phosphorylation due to mtDNA mutations Semin Neurol 2001; 21: 251–60 67 Orth M, Schapira AH Mitochondria and degenerative disorders Am J Med Genet 2001; 106: 27–36 68 Pulkes T, Hanna MG Human mitochondrial DNA diseases Adv Drug Deliv Rev 2001; 49: 27–43 69 Schaefer AM, Taylor RW, Turnbull DM The mitochondrial genome and mitochondrial muscle disorders Curr Opin Pharmacol 2001; 1: 288–93 70 Zeviani M, Klopstock T Mitochondrial disorders Curr Opin Neurol 2001; 14: 553–60 71 Vogler C, Barker J, Sands MS, Levy B, Galvin N, Sly WS Murine mucopolysaccharidosis VIL: impact of therapies on the phenotype, clinical course, and http://neurology.thelancet.com Review pathology in a model of a lysosomal storage disease Pediatr Dev Pathol 2001; 4: 421–33 72 Kwiatkowski DJ, Zhang H, Bandura JL, et al A mouse model of TSC1 reveals sex-dependent lethality from liver hemangiomas, and up-regulation of p70S6 kinase activity in Tsc1 null cells Hum Mol Genet 2002; 11: 525–34 73 Carrie I, Guesnet P, Bourre JM, Frances H Diets containing long-chain n-3 polyunsaturated fatty acids affect behaviour differently during development than ageing in mice Br J Nutr 2000; 83: 439–47 74 Bertram L, Tanzi RE Of replications and refutations: the status of Alzheimer’s disease genetic research Curr Neurol Neurosci Rep 2001; 1: 442–50 75 Isaacs AM, Davies KE, Hunter AJ, et al Identification of two new Pmp22 mouse mutants using large-scale mutagenesis and a novel rapid mapping strategy Hum Mol Genet 2000; 9: 1865–71 76 Beck JA, Lloyd S, Hafezparast M, et al Genealogies of mouse inbred strains Nat Genet 2000; 24: 23–25 77 Small DL, Buchan AM Animal models Br Med Bull 2000; 56: 307–17 78 Elsea SH, Lucas RE The mousetrap: what we can learn when the mouse model does not mimic the human disease ILAR J 2002; 43: 66–79 79 Cohen-Tannoudji M, Marchand P, Akli S, et al Disruption of murine Hexa gene leads to enzymatic deficiency and to neuronal lysosomal storage, similar to that observed in Tay-Sachs disease Mamm Genome 1995; 6: 844–49 80 Phaneuf D, Wakamatsu N, Huang JQ, et al Dramatically different phenotypes in mouse models of human Tay-Sachs and Sandhoff diseases Hum Mol Genet 1996; 5: 1–14 81 Sango K, Yamanaka S, Hoffmann A, et al Mouse models of Tay-Sachs and Sandhoff diseases differ in neurologic phenotype and ganglioside metabolism Nat Genet 1995; 11: 170–76 82 Crawley JN, Paylor R A proposed test battery and constellations of specific behavioral paradigms to investigate the behavioral phenotypes of transgenic and knockout mice Horm Behav 1997; 31: 197–211 83 Crawley JN What’s wrong with my mouse? New York: John Wiley, 2000 84 McIlwain KL, Merriweather MY, Yuva-Paylor LA, Paylor R The use of behavioral test batteries: effects of training history Physiol Behav 2001; 73: 705–17 85 Tecott LH, Wehner JM Mouse molecular genetic technologies: promise for psychiatric research Arch Gen Psychiatry 2001; 58: 995–1004 86 Wehner JM, Radcliffe RA, Bowers BJ Quantitative genetics and mouse behavior Annu Rev Neurosci 2001; 24: 845–67 87 Morris RG Episodic-like memory in animals: psychological criteria, neural mechanisms and the value of episodic-like tasks to investigate animal models of neurodegenerative disease Philos Trans R Soc Lond B Biol Sci 2001; 356: 1453–65 88 Rafael JA, Nitta Y, Peters J, Davies KE Testing of SHIRPA, a mouse phenotypic assessment protocol, on Dmd(mdx) and Dmd(mdx3cv) dystrophindeficient mice Mamm Genome 2000; 11: 725–28 89 Rogers DC, Fisher EM, Brown SD, Peters J, Hunter AJ, Martin JE Behavioral and functional analysis of mouse phenotype: SHIRPA, a proposed protocol for comprehensive phenotype assessment Mamm Genome 1997; 8: 711–13 90 Rogers DC, Peters J, Martin JE, et al SHIRPA, a protocol for behavioral assessment: validation for longitudinal study of neurological dysfunction in mice Neurosci Lett 2001; 306: 89–92 91 Nolan PM, Peters J, Strivens M, et al A systematic, genome-wide, phenotype-driven mutagenesis programme for gene function studies in the mouse Nat Genet 2000; 25: 440–43 92 Bruijn LI, Becher MW, Lee MK, et al ALS-linked SOD1 mutant G85R mediates damage to astrocytes and promotes rapidly progressive disease with SOD1-containing inclusions Neuron 1997; 18: 327–38 93 Ripps ME, Huntley GW, Hof PR, Morrison JH, Gordon JW Transgenic mice expressing an altered murine superoxide dismutase gene provide an animal model of amyotrophic lateral sclerosis Proc Natl Acad Sci USA 1995; 92: 689–93 94 Wong PC, Pardo CA, Borchelt DR, et al An adverse property of a familial ALS-linked SOD1 mutation causes motor neuron disease characterized by vacuolar degeneration of mitochondria Neuron 1995; 14: 1105–16 95 Gabriel JM, Merchant M, Ohta T, et al A transgene insertion creating a heritable chromosome deletion mouse model of Prader-Willi and Angelman syndromes Proc Natl Acad Sci USA 1999; 96: 9258–63 223 For personal use Only reproduce with permission from The Lancet Publishing Group Review 96 Tybulewicz VL, Tremblay ML, LaMarca ME, et al Animal model of Gaucher’s disease from targeted disruption of the mouse glucocerebrosidase gene Nature 1992; 357: 407–410 97 Kooy RF, D’Hooge R, Reyniers E, et al Transgenic mouse model for the fragile X syndrome Am J Med Genet 1996; 64: 241–45 98 Miranda CJ, Santos MM, Ohshima K, et al Frataxin knockin mouse FEBS Lett 2002; 512: 291–97 99 Zhu Y, Romero MI, Ghosh P, et al Ablation of NF1 function in neurons induces abnormal development of cerebral cortex and reactive gliosis in the brain Genes Dev 2001; 15: 859–76 100 Yamamoto A, Lucas JJ, Hen R Reversal of neuropathology and motor dysfunction in a conditional model of Huntington’s disease Cell 2000; 101: 57–66 101 Mallucci GR, Ratte S, Asante EA, et al Post-natal knockout of prion protein alters hippocampal CA1 properties, but does not result in neurodegeneration EMBO J 2002; 21: 202–10 102 Hernandez D, Mee PJ, Martin JE, Tybulewicz VL, Fisher EM Transchromosomal mouse embryonic stem cell lines and chimeric mice that contain freely segregating segments of human chromosome 21 Hum Mol Genet 1999; 8: 923–33 103 Shinohara T, Tomizuka K, Miyabara S, et al Mice containing a human chromosome 21 model behavioral impairment and cardiac anomalies of Down’s syndrome Hum Mol Genet 2001; 10: 1163–75 104 Tomizuka K, Yoshida H, Uejima H, et al Functional expression and germline transmission of a human chromosome fragment in chimaeric mice Nat Genet 1997; 16: 133–43 105 Albrecht U, Sutcliffe JS, Cattanach BM, et al Imprinted expression of the murine Angelman syndrome gene, Ube3a, in hippocampal and Purkinje neurons Nat Genet 1997; 17: 75–78 106 Larsson NG, Rustin P Animal models for respiratory chain disease Trends Mol Med 2001; 7: 578–81 107 Nakada K, Inoue K, Hayashi J Mito-mice: animal models for mitochondrial DNA-based diseases Semin Cell Dev Biol 2001; 12: 459–65 224 Mouse models for neurological disease 108 Wallace DC Mouse models for mitochondrial disease Am J Med Genet 2001; 106: 71–93 109 Jacobs HT Making mitochondrial mutants Trends Genet 2001; 17: 653–60 110 Gibson F, Walsh J, Mburu P, et al A type VII myosin encoded by the mouse deafness gene shaker1 Nature 1995; 374: 62–64 111 Liu XZ, Walsh J, Mburu P, et al Mutations in the myosin VIIA gene cause non-syndromic recessive deafness Nat Genet 1997; 16: 188–90 112 Cook SA, Johnson KR, Bronson RT, Davisson MT Neuromuscular degeneration (nmd): a mutation on mouse chromosome 19 that causes motor neuron degeneration Mamm Genome 1995; 6: 187–91 113 Cox GA, Mahaffey CL, Frankel WN Identification of the mouse neuromuscular degeneration gene and mapping of a second site suppressor allele Neuron 1998; 21: 1327–37 114 Chen Y, Yee D, Dains K, et al Genotype-based screen for ENU-induced mutations in mouse embryonic stem cells Nat Genet 2000; 24: 314–17 115 Brown SD, Balling R Systematic approaches to mouse mutagenesis Curr Opin Genet Dev 2001; 11: 268–73 116 Hagge-Greenberg A, Snow P, O’Brien TP Establishing an ENU mutagenesis screen for the piebald region of mouse chromosome 14 Mamm Genome 2001; 12: 938–41 117 Hrabe de Angelis MH, Flaswinkel H, Fuchs H, et al Genome-wide, large-scale production of mutant mice by ENU mutagenesis Nat Genet 2000; 25: 444–47 118 Herron BJ, Lu W, Rao C, et al Efficient generation and mapping of recessive developmental mutations using ENU mutagenesis Nat Genet 2002; 30: 185–89 119 Nelms KA, Goodnow CC Genome-wide ENU mutagenesis to reveal immune regulators Immunity 2001; 15: 409–18 120 Baird PN, Guymer RH, Chiu D, et al Generating mouse models of retinal disease using ENU mutagenesis Vision Res 2002; 42: 479–85 121 Thaung C, West K, Clark BJ, et al Novel ENUinduced eye mutations in the mouse: models for human eye disease Hum Mol Genet 2002; 11: 755–67 122 Hardisty RE, Mburu P, Brown SD ENU mutagenesis and the search for deafness genes Br J Audiol 1999; 33: 279–83 123 Matin A, Nadeau JH Sensitized polygenic trait analysis Trends Genet 2001; 17: 727–31 124 Flint J, Mott R Finding the molecular basis of quantitative traits: successes and pitfalls Nat Rev Genet 2001; 2: 437–45 125 Moore KJ, Nagle DL Complex trait analysis in the mouse: the strengths, the limitations and the promise yet to come Annu Rev Genet 2000; 34: 653–86 126 Belknap JK, Hitzemann R, Crabbe JC, Phillips TJ, Buck KJ, Williams RW QTL analysis and genomewide mutagenesis in mice: complementary genetic approaches to the dissection of complex traits Behav Genet 2001; 31: 5–15 127 Rozmahel R, Wilschanski M, Matin A, et al Modulation of disease severity in cystic fibrosis transmembrane conductance regulator deficient mice by a secondary genetic factor Nat Genet 1996; 12: 280–87 128 Zielenski J, Corey M, Rozmahel R, et al Detection of a cystic fibrosis modifier locus for meconium ileus on human chromosome 19q13 Nat Genet 1999; 22: 128–29 129 Warrick JM, Chan HY, Gray-Board, Chai Y, Paulson HL, Bonini NM Suppression of polyglutamine-mediated neurodegeneration in Drosophila by the molecular chaperone HSP70 Nat Genet 1999; 23: 425–28 130 Cummings CJ, Sun Y, Opal P, et al Over-expression of inducible HSP70 chaperone suppresses neuropathology and improves motor function in SCA1 mice Hum Mol Genet 2001; 10: 1511–18 131 Hunter AJ, Nolan PM, Brown SD Towards new models of disease and physiology in the neurosciences: the role of induced and naturally occurring mutations Hum Mol Genet 2000; 9: 893–900 132 Bucan M, Abel T The mouse: genetics meets behaviour Nat Rev Genet 2002; 3: 114–23 133 Dennis MB Jr Welfare issues of genetically modified animals ILAR J 2002; 43: 100–09 THE LANCET Neurology Vol August 2002 http://neurology.thelancet.com For personal use Only reproduce with permission from The Lancet Publishing Group