Similarly to Sanger sequencing, genomic regions that have homologous loci are also diffi cu lt to assay. Even though this is usually screened for before designing the array, if the va riant to be assessed is within one of these high-homology regions, a probe with suffi cient specifi city will be diffi cult to design. Th e Roche (454) GS FLX sequencer was fi rst introduced com- mercially in 2004. Th is seque ncer works on the principle of \200\230pyro- sequencing \200\231, which uses the pyrophosp hate molecule released on nucleotide incorporation by DNA polymerase to fuel a downstream s et of reactions that ultimately produce light from the cleavage of oxyluciferin by lucifera se (21). Th e current 454 instrument pro- duces an average read length of approximately 400 bp per sample, with a throughput of about a billion bases per day. It is also worth noting that since this technology assays individ- ual SNPs, chromosomal tr anslocations are virtually impossible to detect, given that the only output obtained is sig nal intensity per individual marker, regardless of its position on the genome. Although high-throughput genome-wide genotyping studies have yielded very exciting results, and the technology is now stable, robust, and very reproducible, the use of this approach has always been perceived as transitory and not as the ultimate analytical meth- odology. A s we advance rapidly towards ever more cost-eff ective sequencing technology, it is expecte d that high-throughput geno- typing will be replaced by whole-genome sequencing. Applied Biosystem \200\231s SOLiD sequencer was initially released in 2007 and relies on an innovative ligation-based chemistry (21). One of the main advantages of this approach is t hat it has one of the lowest error rates of all three technologies. Th e current SOLiD inst rument, the 5500xl, has a throughput of 30 billion bases per day with read lengths of up to 75 bp. Massively parallel sequencing In the past fi ve years new chemistries and instruments have been developed to perform sequencing (so-called \200\230next-generation \200\231 or \200 \230massively parallel \200\231 instruments), and such technology has already transformed t he fi eld. Th eir impact on genomics has been so dra- matic that the very nature of genetic experimentation has shift ed towards more comprehensive and complete analysis. Th e requir e- ments for handling these data are also fundamentally diff erent from those associated wi th Sanger sequencing, particularly the compu- tational algorithms used, and their continuou s improvement has In 2006 Illumina introduced their fi rst instrument, the Genome Analyzer, which has since s een several improvements, increasing throughput and reducing the times per run (22). In 201 0 a new instrument was released commercially which, although utilizing the same chemistry, improved throughput dramatically and quickly became the sequencer of choice for the vast ma jority of genome centers. Illumina \200\231s approach is based on the concept of \200\230s equencing by synthesis \200\231 (SBS) to produce sequence reads of 35 \200\223150 bp from t ens of millions of surface-amplifi ed DNA fragments simultane- ously. Th e major features o f this technology are the low turnaround CHAPTER 7 Genetic Techniques, Impact and Diagnostic Issues in Movement Disorders 57 time and the amount of data produced. At the time of writing, Illumina \200\231s HiSeq is c apable of producing over 600 billion bases per eight-day run, with read lengths of up to 15 0 bp. In parallel with whole-exome and whole-genome sequencing, it is now also possible to sequen ce the entire transcriptome (the collection of RNA molecules, including mRNA, rRNA, tRNA, a nd non-coding RNA, usually called RNA-seq) of an individual (30). Th is is potentially a ve ry powerful approach that is still in its early days, partly because the data analysis is m ore complex than that required for DNA sequencing. For example, a single gene will have sev eral transcripts, created by alternative splicing, which may or may not have been previousl y described. Th is has implications for primary data analysis, particularly the mapping of reads to the genome, which means that if a read is uniquely aligned to the wrong gene, such as a paralog, this may result in false-positive vari- ants being called. One of the added benefi ts of RNA-seq is that it detects both sequence variability and expression levels sim ultane- ously. Th is means that tissue-specifi c genes will only be detected (for both expr ession and sequence variability) when using RNA extracted from that same tissue. While the improvements in sequencing have been astonishing, clearly defying Moore \200\231s Law (23), the cost is still high enough to preclude whole-genome sequencing en masse at th e present time. To this end, strategies need to be developed to leverage this tech- nology and simultaneously provide cost-eff ectiveness. Th e fi rst approach was derived from commo n sense: given that the por- tion of the genome that we can most easily understand is the c oding region, sequencing should start by targeting this small portion \200\224the so-called exome (24). Much like genotyping arrays, exome sequencing is a transitory approach that re fl ects the most cost-eff ective approach to \200\230next-generation \200\231 sequencing t hat is cur- rently available. Exome sequencing follows the same principle as described for the technologies above, but in cludes an additional step where the genomic DNA library is hybridized with a pool of short biotin-labeled oligonucleotides that were designed to be comple- mentary to the entire coll ection of exons in the genome. Th ese are then magnetically captured and the non-coding DNA is washed away. Th is creates a library that contains only DNA fragments rep- resentative of the coding region. Th ese high-throughput approaches may play a very signifi cant role in the diagnostics labo ratory in the next three to fi ve years. One can certainly envisage a screening approach th at relies on the geno- typing of disease-specifi c markers using microarrays, while simul- taneously whole-genome and transcriptome are being sequenced to detect novel variability. T h is approach is likely to be very suc- cessful in determining the genetic causes of a high proportion of diseases at a fraction of the total cost that novel/complex diseases face in their diagnostic work-up. Th is approach has already permitted the identifi cation of the genetic causes underlying a signifi cant number of Mendelian dis- eases (24 \200\22329). While this is a powerful appr oach, there are some caveats. Th e fi rst of these is that by sequencing the exome, a sig- nifi cant proportion of variation in regulatory regions that may play a role in the disease is missed. Additionally, no method has yet allowed for the sequencing of the entire collec tion of known exons. Typically, 15 \200\22320% of known coding exons are not captured as th ey are located in complex genomic regions, for which capture probes are diffi cult to desig n. Added to this percentage are all the exons that are not properly described or annotated in the current genome reference build. Moreover, since these technologies all use short-rea ds, determining phase is still a poorly addressed issue, unless relevant family members are available for testing, which increases costs linearly. Th e combination of short-reads wit h the fact that only small dispersed fragments of the genome are being sequenced also means that mid-size insertions or deletions are dif- fi cult to assess, particularly if they are heterozygous. Th e identifi - cation of 1 \200\2235 bp deletions is quite accurate, but a heterozygous deletion of 60 \200\223100 bp is less so. Single-molecule sequencing Th e next major advance in genomics technology will be single- m olecule sequencing. Th e premise behind this technique is that it will be capable of sequen cing one DNA molecule at a time, whether that molecule is a constituent of an amplifi ed sa m- ple or unamplifi ed DNA. Th ere are two primary advantages of single-molecule sequencing . First, individual fragments of DNA do not need to be amplifi ed or, when they are, far fe wer cycles of amplifi cation are required. Secondly, sequencing can occur in real time, mea ning that a complete human genome could be sequenced within minutes. Several approaches to single-molecule sequencing are currently being tested; these range from scan- ning probe te chniques, to fl uorescence techniques, to nanopore approaches (31 \200\22334). Th e last of these possibly has the greatest promise. Th e most advanced nanopore approach is that bein g developed by Oxford Nanopore, where a molecule of DNA is passed through an alpha-haemolys ine pore with an exonuclease attached that cleaves the DNA molecule one base at a time. Th e bases are dropped through the nanopore, which is embedded in a membrane. As the DNA passe s through the nanopore, the conductance across the membrane changes: diff erent base pairs change the conductance in diff erent ways, allowing each base to be accurately identifi ed. In some cases, however, whole-genome sequencing has been per- formed instead. Th is has usu ally occurred when a small number of individuals, with a very defi ned and oft en rare phen otype, is being studied. Th e major diff erences in this approach relate to the cost of the analysis, handling much larger amounts of data, and ulti- mately interpreting these data. It is worth noting that, so far, all genome sequencing projects that have revealed new muta tions could theoretically have arrived at the same result by perform- ing exome sequencing, since the mutations were all coding and in known genes. Th is is not advocating exome sequ encing in place of whole-genome sequencing; it simply shows that our understanding of the e ff ects of genetic variability is still very much confi ned to the coding portion of the ge nome. As we continue to gather evidence for, and increase our knowledge of, functional eff ects of non-coding variability, a whole new range of novel, pathogenic, non-coding mutation s is likely to be discovered. Th e implications of this technique are enormous. Th e ability to sequence one DNA molecule in its entirety (presumably paralleliz- ing reactions will allow for increased coverage), with costs that are negligible compared with current methods, which allows an entire genome to be sequenced in a matter of hours, or even minutes, instead of weeks, and which allows one to clearly determine phase (since only one molecule is being sequenced at a time) is li kely to revolutionize the fi eld of genetics. Oxford Textbook of Movement Disorders 58 Future challenges: de \201 ning variability in human genomes Our knowledge of the human ge nome has seen great improvements over the last few years with the arrival of these high-thr oughput tech- nologies, but also as a result of the Human Genome and HapMap projects. Th is has allowed us to perform many genome-wide asso- ciation and exome-sequencing studies that have elucidated, to some extent, the genetic basis of several complex neurological disease s. Although we understand genetic diversity among humans fairly well at the level of common variants, we still know very little about the amount and type of variation below that leve l of frequency. Since common variants have not fully explained all the heritability in thes e diseases, it is expected that less common and rarer alle- les will also contribute. Th is is the case with some of the known genes for Parkinson \200\231s disease: common variabili ty in SNCA confers risk for PD, and rare mutations are the cause of the disease in a few fa milies. It will soon be possible to re-sequence large numbers of healthy controls to determ ine a baseline for \200\230normal \200\231 variation, which will be fundamental for future studies of complex diseases. Th is is one of the goals of the 1000 Genomes Project, althou gh a much larger number of individuals, from a variety of genetic back- grounds, need to be sequenced to better capture the spectrum of variability in the human genome (35). expected that it will soon reach the $1000 mark, thus becoming a viable approach to clinica l assays and a point of entry for health insurance or medical care. Th e question is whethe r we are able to keep pace with technology and be in a position to make sense of the raw da ta these technologies will generate. Conclusions and future prospects Despite the remarkable advances in genome analysis methods , the current methodologies have some limitations that should be high- lighted. Even in mon ogenic diseases, where it would be expected that identifying a pathogenic mutation would be easiest, there are two major problems. Th e fi rst relates to technology \200\224for examp le, inhomogeneous coverage of the target region. Th is problem leads to a widely varying nu mber of reads covering diff erent regions of the exome which cause, in conjunction with une qual sequencing quality, the detection of false-positive and false-negative variants. Sange r re-sequencing can eliminate false-positive variants, but false-negative variants are pote ntially a major problem because if the fi ltering strategy relies on the detection of mutat ions in the same gene in diff erent aff ected individuals, these might be missed. Th ese pr oblems are likely to be overcome with further technological progress. Th e main genetic pro blem is locus heterogeneity, which is the presence of multiple distinct disease genes/disea se loci for one clinical phenotype. Some neurological disorders, such as Charcot \200\223 M arie \200\223Tooth disease (with over 40 loci), hereditary ataxia, and hereditary spastic p araplegia, show a high degree of locus hetero- geneity. Th e main biological problem is pro ving the causality of a mutation. Ultimately, causality has to be shown using cell and ani- mal model systems, although currently segregation is usually the minimum requirement. With the increase in knowledge, and especially in understand- ing, of human genome variabil ity, we will also see great improve- ments in our ability to correctly annotate variability leading to functional changes. Th ese annotations will add a whole new layer of data to ge nome variation, making these results much more read- ily interpretable in terms of their po tential impact(s) on function and, ultimately, biology. Th is integrated knowledge will sig nifi - cantly enhance our ability to interpret the genome in the context of observed human phenotypes, eventually leading to the era of personalized genomics. In cases where the causative genes are known and screening of these needs to be performed, using approaches like genome or exome sequencing will discover not only the causative mutat ion for the investigated disease, but also mutations causing other mono- genic disorders an d genetic risk factors for complex disorders. Th is creates an obvious ethical problem. Towards the \200\230$1000 \200\231 genome and personalized medicine Th ese high-throughput technologies, by enabling vast data genera- tion, together with our ever-increasing unders tanding of human genome variability and its impact on biology, will provide the groundwork for true genetic disease risk prediction and therapeu- tic approaches. Targeted treatments, which selectively block the impact of certain variants, might also be suggested. One of the most signifi cant issues that still needs to be addressed is that of phase. None of the current technologies allows us to deter- mine genome-wide maps of phased haplotypes , which means that we are still neglecting one of the most fundamental aspects of the human genome \200\224its diploid nature. Th e coming years will see a great increase in the description of variants found in exomes and genomes; some of these will simply be rare non-functional variants while others will ha ve important functional implications for the biology of the human genome. Th e diagnostic p ower of whole-genome sequencing in the context of genetically heterogeneous Mendelian disea se has already been shown (36) and it is likely that the identifi cation of rare heterogene - ous alleles, by means of whole-genome sequencing, will be the only way to defi nitively d etermine genetic contribution to the associated clinical phenotypes. Direct-to-consumer genetic test companies are the fi rst approach to this paradigm. Here, a ny individual is able to send a sample of their DNA, from which the companies will generate whole-genome genotyping data. Although this is not a diagnostic test per se , the implicat ions for diagnosis are obvious. Furthermore, the fact that such companies are providing thi s service and that customers are interested in it, shows that this approach appeals to the general population. Th is system has one severe limitation: only a very lim- ited and selec ted subset of markers is medically actionable, since only known variants that it is possibl e to design assays for can be tested. Th is clearly suggests that, although genotyping meth ods may become one aspect of personalized medicine in the future, they will not be the corn erstone of personal genomics. References 1. Ross CA, Tabrizi SJ . Huntington \200\231s disease: from molecular pathogenes is to clinical treatment. Lancet Neurol 2011; 10 : 83 \200\22398 . 2. Zimprich A, Biskup S, Leitner P, et al. Mutations in LRRK2 cause autosomal-dominan t par kinsonism with pleomorphic pathology. Neuron 2004; 44 : 601 \200\2237 . A mere three years ago, a whole genome sequence would cost $700 000; today that cost has d ropped to $7 000 and it is CHAPTER 7 Genetic Techniques, Impact and Diagnostic Issues in Movement Disorders 59 21. Margulies M, Egholm M, Altman WE, et al . Genome sequencing in microfabricated high-den sit y picolitre reactors. Nature 2005; 437 : 376 \200\2238 0 . 3. Valente EM, Abou-Sleiman PM, Caputo V, et al . Hereditary early-onse t Parkinson \200 \231s disease caused by mutations in PINK1. Science 2004; 304 : 1158 \200\2236 0 . 22. Bentley DR, Balasubramanian S, Swerdlow HP, et al. Accurate whole human genome sequenci ng using reversible terminator chemistry. Nature 2008; 456 : 53 \200\2239 . 4. Pais n-Ru z C, Jain S, Evans EW, et al. Cloning of the gene containing mutations t hat cause PARK8 -linke d Parkinson \200\231s disease. Neuron 2004; 44 : 595 \200\22360 0 . 23. Moore GE . Cramming more components onto integrated circuits. Proc 5. Bonifati V, Rizzu P, van Baren MJ, et al . Mutations in the DJ- 1 gene asso- ciated with autosomal recessive early-onse t parkinsonism. Science 2003; 299 : 256 \200\2239 . IEEE 1998; 86 : 82 \200\2235 . 24. Ng SB, Buckingham KJ, Lee C, et al. Exome sequencing identifi es the 6. Kitada T, Asakawa S, Hattori N, et al . Mutations in the parkin gene cause autosomal rec essive juvenile parkinsonism. Nature 1998; 392 : 605 \200\2238 . 7. Polymeropoulos MH, Lav edan C, Leroy E, et al . Mutation in the alpha-synuclei n gene identifi ed in families with Parkinson \200\231s disease. Science 1997; 276 : 2045 \200\2237 . cause of a Mendelian disorder. Nat Genet 2010; 42 : 30 \200\2235 . 25. Bilg var K, Ozt rk AK, Louvi A, et al . Whole-exom e sequencing identi- fi es rec essive WDR62 mutations in severe brain malformations. Nature 2010; 467: 207 \200\22310. 26. Varela I, Tarpey P, Raine K, et al. Exome sequencing identifi es frequent mutation of t he SWI/SNF complex gene PBRM1 in renal carcinoma. Nature 2011; 469 : 539 \200\2234 2 . 8. West A, Periquet M, Lincoln S, et al. Complex relationship between Parkin mutations and Parkinson disease. Am J Med Genet 2002; 114 : 584 \200\2239 1 . 9. Sanger F, Nicklen S, Cou lson AR . DNA sequencing with chain-terminatin g 27. \207 al \237 kan M, Chong JX, Uricchio L, et al . Exome sequencing reveals a novel mutation for autosomal recessive non-syndromi c mental retarda- tion in the TECR gene on ch romosome 19p13. Hum Mol Genet 2011 ; 20 : 1285 \200\2239 . inhibitors. Proc Natl Acad Sci USA 1977; 74 , 5463 \200\2237 . 10. Sanger F, Air GM, Barrell BG, et al. Nucleotide sequence of bacteri- ophage phi X174 DNA. Nature 1977; 265 : 687 \200\2239 5 . 28. Johnson JO, Mandrioli J, Benatar M, et al . Exome sequencing reveals VCP mutations as a cause of familial ALS. Neuron 2010; 68 : 857 \200\2236 4 . 29. Wang JL, Yang X, Xia K, et al . TGM6 identifi ed as a novel causative gene of spinocerebellar ataxias using exome sequ encing. Brain 2010; 133 : 3510 \200\22318 . 11. Lander ES, Linton LM, Birren B, et al. Initial sequencing and analysis of the human genome. Nature 2001; 409 : 860 \200\22392 1 . 12. Sidransky E, Nalls MA, Aasly JO, et al. Multicenter analysis of glucocer- ebrosidase mu tations in Parkinson \200\231s disease. N Engl J Med 2009; 361 , 1651 \200\2236 1 . 30. Mortazavi A, Williams BA, McCue K, Schaeff er L, Wold B . Mapping and quantifying mamma lian transcriptomes by RNA-Se q. Nat Methods 2008; 5 : 621 \200\2238 . 13. Lambert JC, Heath S, Even G, et al . Genome-wid e association study identifi es variant s at CLU and CR1 associated with Alzheimer \200\231s disease. Nat Genet 2009; 41 : 1094 \200\2239 . 31. Lund J, Parviz BA . Scanning probe and nanopore DNA sequencing: core 14. Mart nez-Arias R, Calafell F, Mateu E, Comas D, Andr s A, Bertranpetit J . Sequen ce variability of a human pseudogene. Genome Res 2001; 11 : 1071 \200\2238 5 . techniques and possibilities. Methods Mol Biol 2009; 578 : 113 \200\2232 2 . 32. Wallace EV, Stoddart D, Heron AJ, et al . Identifi cation of epigenetic DNA modifi cati ons with a protein nanopore. Chem Commun 2010; 46 : 8195 \200\2237 . 15. Martinez-Arias R, Comas D, Mateu E, Bertranpetit J . Glucocerebrosidase pseudogene vari ation and Gaucher disease: recognizing pseudogene tracts in GBA alleles. Hum Mutat 2001; 17 :191 \200\2238. 33. Stoddart D, Heron AJ, Klingelhoefer J, Mikhailova E, Maglia G, Bayley H . Nucleobase re cognition in ssDNA at the central constriction of the alpha-hemolysi n pore. Nano Lett 2010 ; 10 : 3633 \200\2237 . 16. Hruska KS, LaMarca ME, Scott CR, Sidransky E . Gaucher disease: muta- tion and polymorp hism spectrum in the glucocerebrosidase gene (GBA). Hum Mutat 2008; 29 : 567 \200\2238 3 . 34. Maglia G, Heron AJ, Stoddart D, Japrung D, Bayley H . Analysis of single nucleic acid m olecules with protein nanopores. Methods Enzymol 2010; 475 : 591 \200\22362 3 . 17 . Th e International HapMap Project. Nature 2003; 426 : 789 \200\2239 6 . 18. Singleton AB, Farrer M, Johnson J, et al. Alpha-synuclein locus triplica- 35. 1000 Genomes Project Consortium, Abecasis GR, Altshuler D, Auton A, et al. 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This page intentionally left blank CHAPTER 8 Overview of Parkinsonism and Approach to Di \200 erential Diagnosis Sabine Spiel berger and Werner Poewe Summary Parkinsonism is one of the most common movement disorders and may be the clinical m anifestation of a variety of neurodegen- erative, structural, toxic, metabolic, infectious, and vascular dis- orders. Since therapy and prognosis diff er considerably between diff er ent forms of parkinsonism, an early correct diagnosis is of great clinical signifi cance bu t may be challenging on clinical grounds alone. diff erential diagnosis within this heterogenic group of parkinso- nian syndromes is a matt er of clinical acumen and judicious use of the ancillary investigations discussed below. Epidemiology of parkinsonism Parkinsonism is one of the most common movement disorders amon g the elderly. Published reports on the prevalence of parkin- sonism estimate a range of 2 \200\22315% in the population aged over 65 (1 \200\2235), with incidence increasing with ag e. A community-based sur- vey documented a prevalence rate of 52.4% for parkinsonism in the population over the age of 85 (2), and therefore, although it occurs worldwide, parkinsoni sm is more frequent in regions with longer life expectancy. By far the most common cause of parkinsonism is Parkinson \200\231s disease (PD), followed by drug-induced parkinsonism an d vascular parkinsonism (1, 4). Atypical parkinsonian disorders, such as multiple system at rophy (MSA), progressive supranuclear palsy (PSP), and corticobasal syndrome (CBS), are rar e causes of parkinsonism, with estimated prevalences ranging from 1 to 6 per 100 000 (6, 7) . Ancillary tests for enhancement of diagnostic accuracy include drug-challenge testing to as sess dopaminergic responsiveness, neuroimaging to evaluate nigrostriatal presynaptic dopami nergic function or evidence for structural abnormalities, and genetic test- ing to identify common genetic subtypes of Parkinson \200\231s disease. Th erapeutic management of parkins onism mainly depends on the underlying pathology. Levodopa is the single most eff ective wa y to treat the motor symptoms of Parkinson \200\231s disease, but it may also be at least p artially eff ective in other types of parkinsonism, so that a trial with levodopa will be p art of the treatment approach in most patients presenting with parkinsonism. Aetiopathogenesis Th e etiology of symptomatic forms of parkinsonism is diverse (see Table 8.1), but is usually defi nable in terms of functional or structural disorders aff ecting n igrostriatal dopamine transmission or downstream signaling pathways, although this is only partially understood in degenerative parkinsonism. Up to 5% of clinical cases of PD may be caused by Mendelian mutations, and recent genome-wide association studies (GWAS) have ident ifi ed eleven risk loci associating with PD, some of which overlap with the known causative mutations such as alpha-synuclein (8). Taken together, however, such loci only about doubl e a person \200\231s risk of developing PD, suggesting that there are multiple additional a etiological fac- tors, both genetic and environmental. Nevertheless, genetic fi nd- ings in PD suggest that dysfunction at the level of the mitochondria, synuclein processing, and en dolysosomal traffi cking are all likely to be important in PD pathogenesis. Aetiopathogenes is of other types of degenerative parkinsonism, such as MSA, PSP, or CBS, is even Essential background Classi \201 cation of parkinsonism A parkinsonian syndrome is a prese nting or important clinical feature in a variety of distinct conditions including neurodege n- erative, metabolic, toxic, infl ammatory, and autoimmune disor- ders, as well as vascula r or neoplastic disorders. In addition, there are rare cases without an identifi able organ ic cause which satisfy criteria for psychogenic parkinsonism. For pragmatic clinical pur- p oses, parkinsonian disorders are best grouped into two principal categories: degenerative p arkinsonism, where a variety of sporadic and genetic degenerative diseases cause nigrostria tal neurodegen- eration, and symptomatic or secondary parkinsonism, caused by non-degenerat ive lesions of the same system or other sites of the striatopallidothalamic-cortical motor circuitry (see Table 8.1). Several of these disorders are discussed in detail in other chap ters in this book. In the absence of reliable and valid biomarkers, the Oxford Textbook of Movement Disorders 62 Table 8.1 Overview of principal causes of parkinsonism Neurodegenerative parkinsonism Sporadic Sporadic Parkinson \200\231s disease (PD) Multiple system atrophy (MSA) Progressive supranu clear palsy (PSP) Corticobasal syndrome (CBS) Dementia with Lewy bodies (DLB) Genetic Genetic Parkinson \200\231s disease (PARK 1 \200\22318) Huntington \200\231s disease (Westp hal variant) SCA mutations (SCA2, SCA3) Frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17) Wilson disease Neuronal brain iron accumulation syndrome (NBIA) Pa rkinsonism \200\223amyotrophic lateral sclerosis \200\223dementia complex of Guam PSP-like parkinsonism in Guadeloupe X-linked dystonia parkinsonism (DYT 3, XDP, Lubag) Dopa-responsi ve dystonia (DYT 5, Segawa syndrome) Rapid-onset dystonia parkinsonism (DYT 12) Dentatorubr al-pallidoluysian atrophy Fragile X-associated tremor \200\223ataxia syndrome Neuroacanthoc ytosis Symptomatic parkinsonism Infectious Post-encephalitic (encephalitis lethargica, Japanese B, post-streptococcal, other viral inf ections (HIV, PML)) Prion disease Toxoplasmosis Toxic Carbon monoxide Cyanide MPTP Manganese Methanol Rotenone Drug-induced Dopamine-receptor blockers Dopamine-depleting drugs Other drugs (e.g. valproic acid, calciu m-channel blockers) Brain tumors Supratentorial tumors (e.g. frontal meningioma) Brainstem tumors (rare) Metabolic Hypoparathyroidism Extrapontine myelinolysis Chronic liver failure Fahr \200\231s disease Miscellaneous Normal pressure hydrocephalus Hemiatrophy-hemiparkinson syndrome Psychogenic HIV, human immunode \201 ciency virus; MPTP, 1-methyl-4-phenyl-1,2,3-tetrahydropyridine ; PML, progressive multifocal leucoencephalopathy and relaying this to the internal segment of the globus pallidus (GPi), which in turn proje cts via inhibitory GABAergic projec- tions to the motor segments of the ventral anterior th alamus from where glutamatergic facilitatory input reaches the motor planning and iniating structures of the prefrontal cortex (see Fig. 8.1a). Th is signaling circuit is controlled and modulated by the nigrostriatal dopamine projection, such that dopamine defi ciency or p atholo- gies aff ecting striatal dopamine receptors will lead to changes in the two princip al striatopallidal output pathways targeting the GPi in a monosynaptic fashion (direct path way) or via relays in the exter- nal pallidum (indirect pathway). Th e net eff ect of dysfu nctional dopaminergic input onto these two striatal neuronal populations of projection neur ons is increased fi ring of the GPi, resulting in less clear, but involves oligodendroglial alpha-synuclein pathology in MSA and neuronal tau pathology in PSP and CBS. More detailed discussions are given in the corresponding chap- ters of this volume. Pathophysiology Regardless of etiology, the principal pathophysiological abnor- mality unde rlying the cardinal motor features of parkinsonism \200\224in particular bradykinesia \200 \224is related to dysfunctional motor loops connecting the prefrontal cortex, basal ganglia , and motor thala- mus. According to a model of basal ganglia motor processing origi- nally proposed and developed by Alexander and colleagues (9), the striatum plays a central role in processing sensorimotor information CHAPTER 8 Overview of parkinsonism and approach to differential diagnosis 63 (a) (b) Cortex Cortex Glu Glu Striatum Striatum Glu Glu MSN-D2 MSN-D1 MSN-D1 MSN-D2 y a w h t a p y a w h t a p y a w h t a p y a w h t a p l l a d a d SNc SNc l l a d a d i l l i l l i l l i l l a p o t a i r t s a p o t a i r t s a p o t a i r t s a p o t a i r t s GPe Th Th GPe c i g r e A B A G t c e r i d n c i g r e A B A G t c e r i d n c i g r e A B A G t c e r i D c i g r e A B A G t c e r i D GABA GABA GABA GABA Glu Glu I I STN GPi STN GPi Fig. 8.1 (a) Basal ganglia motor loops with direct and indirect pathways. Black arrows indi cate inhibition; white arrows indicate activation. SNc, substantia nigra pars compacta; GPe , globus pallidus externus; STN, subthalamic nucleus; GPi, globus pallidus internus; SNr, s ubstantia nigra pars reticulata; Th , thalamus; Glu, glutamate; Enk, encephalin; SP, substa nce P; MSN, medium spiny neurons. (b) Impaired basal ganglia motor loops in patients with P arkinson \200\231s disease. Owing to neurodegeneration in the SNc, dopaminergic input into the striatum is reduced, resulting in a loss of excitatory dopaminergic drive on D1 recepto rs and inhibitory dopaminergic input on D2 receptors. Th e net result is increased GPi acti vity via direct and indirect pathways, leading to inhibition of Th and cortex. increased inhibitory input into the motor thalamus, thus reducing thalamocortical motor dri ve with underactivity of the motor parts of the prefrontal cortex leading to impaired movem ent initiation, speed, and amplitude (see Fig. 8.1b). Similar eff ects may also be due to d ownstream events at the level of the pallidum, motor thalamus, or thalamocortical projectio ns, as is the case in diff erent types of secondary parkinsonism. the single most disabling motor feature of PD, impacting on virtu- ally every aspect of act ivities of daily living, including handwrit- ing, getting dressed, handling feeding utensil s, hygiene, speech, and communication, as well as general mobility. Hypomimia with decrease d blink rate, micrographia with decrementing script size within a sentence or even a word ( see Fig. 8.2), or hypophonia with reduced volume of speech are clinical terms frequently us ed to describe isolated features of parkinsonian bradykinesia. Common early complaints of p atients related to bradykinesia include hand- writing diffi culties, buttoning up a shirt, turning keys, or brushing their teeth, while relatives, colleagues, or friends may note cha nges in facial expression, a soft er voice, or reduced arm swing or drag- ging of one leg w hile the patient is walking. With more advanced disease, getting up from an armchair or out of a car seat or turn- ing around in bed, as well as problems with walking become more pro minent among the symptoms reported in the patient interview. A peculiar and severely disabl ing feature of parkinsonian motor impairment, which is more aptly covered under the term of aki- nesia rather than bradykinesia, comprises diffi culties of movement initiation, and m aintenance of a fl ow of sequential or repetitive movements. Th is commonly manifests as he sitation when a patient is about to start walking or as sudden stops in a sequence of fi ng er Clinical presentation Cardinal features of parkinsonian syndromes A parkinsonian syndrome i s defi ned by the presence of bradyki- nesia in combination with at least one of the follow ing signs: rest- ing tremor, rigidity, and impaired postural refl exes, with the last of th ese not being the result of sensory, proprioceptive, or cerebellar dysfunction (10). Accord ing to the UK Parkinson \200\231s Disease Society Brain Bank criteria (QSBB criteria) brady kinesia is the anchor fea- ture for parkinsonism and is used as an umbrella term including reduced movement speed (bradykinesia in the strict sense) and amplitude (hypokinesia), as w ell as slowness of movement initiation, and decrementing speed and amplitude with or withou t interrup- tions (akinesia) of sequential repetitive movements. Bradykinesia is Fig. 8.2 Micrographia with decrementing script size in a patient with idiopathic PD. Oxford Textbook of Movement Disorders 64 tapping as well as during walking. Th e prototypical example of such motor blocks is freezi ng of gait (FoG), where the patient \200\231s feet appear to be glued to the ground while t he trunk is moving forward. FoG is a major cause of falls in people with parkinsonian disor ders and is commonly induced by changes in gait rhythmicity, such as turn- ing, as well as by environmental stimuli such as narrow passages or people approaching in the opposite dire ction. An extreme form of start hesitation with freezing, where patients are unable to init iate walking but once under way may almost walk normally, has been described as \200\230ga it ignition failure \200\231 (11). (also to avoid aspiration pneumonia) within the fi rst six to eight years of disease. Postural deformities are also common to many types of parkinson- ism. A stooped posture wit h fl exion of the trunk is a hallmark of PD, while in MSA parkinsonism antecollis or mixed anterolat erocollis deviations are oft en seen in the absence of signifi cant trunk fl ex- ion (13). However, patients with PSP typically show some degree of hyperextension of the ne ck and trunk, while severe degrees of trunk fl exion, termed camptocormia, occur in PD and occasion- ally in MSA (14). Th e stooped posture of PD as well as hyperexten- sion of the n eck in PSP have been related to muscular rigidity, but there is ongoing controversy as to w hat extent antecollis posturing in MSA or camptocormia in PD might also be the result of dy stonic muscle contractions or myopathy of extensor muscles (15, 16). Most patients presenting with bradykinetic parkinsonism also exhibit muscular rigidity , ch aracterized by a steady resistance to passive movement which is independent of velocity and may include cogwheel-like rhythmic oscillations. Rigidity in parkin- sonism is oft en desc ribed as lead-pipe rigidity since it is present to the same extent over the whole range of movement. It tends to aggravate when patients are asked to perform active movements of the opposite limb during rigidity testing. Parkinsonian rigidity can aff ect both limb and axia l muscles, with somewhat diff erent distri- butions in the diff erent forms of parkinsonism . Dystonia is a rare presenting feature of idiopathic PD (17), but is not uncommon in autosom al recessive froms with young-onset disease like Parkin, Pink-1, and DJ-1 where focal limb dystonia has been reported in up to 40% of cases (18). Focal limb dystonia with strictly as ymmetrical parkinsonism should raise suspicions of CBS; further examples of dystonia \200 \223parkinsonism syndromes include rare genetic disorders such as X-linked rapid-onset dyst onia \200\223par- kinsonism (Lubag), PARK 9 (Kufor \200\223Rakeb disease), PARK 16 or neuro degeneration with brain iron accumulation (NBIA), Wilson disease or secondary parkinsonian syndromes in manganese, car- bon monoxide, or cyanide poisoning, basal ganglia infarcts, or drug-induced parkinsonism (see Table 8.2). Tremor (involuntary rhythmical sinusoidal oscillation of a body part) is the third cardinal motor feature of parkinsonism. Th e prototypical parkinsonian tremor is a 4 \200\2236 Hz r esting tremor, which decreases or diminishes upon voluntary movements to revert aft er a ne w position is obtained (re-emergent tremor). It may be aggravated by fatigue, stress, or em otions. Unilateral onset and persistent asymmetry defi ne the classical rest tremor of PD, with a subgroup of patients also showing additional pos- tural tremor of higher frequency ( 12). In PD, rest tremor usually involves the limbs in isolation; head, chin, and voice trem or are uncommon. Similarly, classical 5 Hz rest tremor is uncommon in atypical degenerative parkinsonian disorders such as MSA or PSP, where irregular and jerky types of postural or kinetic tremor prevail.