Congenital myasthenic syndromesDaniel Hantaı a, Pascale Richardb, Jeanine Koeniga and Bruno Eymarda

Purpose of review

Congenital myasthenic syndromes are a heterogeneous group

of diseases caused by genetic defects affecting neuromuscular

transmission. In this article, a strategy that leads to the

diagnosis of congenital myasthenic syndromes is presented,

and recent advances in the clinical, genetic and molecular

aspects of congenital myasthenic syndrome are outlined.

Recent findings

Besides the identification of new mutations in genes already

known to be implicated in congenital myasthenic syndromes

(genes for the acetylcholine receptor subunits and the collagen

tail of acetylcholinesterase), mutations in other genes have more

recently been discovered and characterized (genes for choline

acetyltransferase, rapsyn, and the muscle sodium channel

SCN4A). Fluoxetine has recently been proposed as an

alternative treatment for ‘slow channel’ congenital myasthenic

syndrome.

Summary

The characterization of congenital myasthenic syndromes

comprises two complementary steps: establishing the diagnosis

and identifying the pathophysiological type of congenital

myasthenic syndrome. Characterization of the type of congenital

myasthenic syndrome has allowed it to be classified as caused

by presynaptic, synaptic and postsynaptic defects. A clinically

and muscle histopathologically oriented genetic study has

identified several genes in which mutations cause the disease.

Despite comprehensive characterization, the phenotypic

expression of one given gene involved is variable, and the

aetiology of many congenital myasthenic syndromes remains to

be discovered.

Keywords

electromyography, genetic diagnosis, microelectrophysiology,

neuromuscular junction molecules, neuromuscular transmission,

treatment

Curr Opin Neurol 17:539–551. # 2004 Lippincott Williams & Wilkins.

aInserm U582 and Unite Clinique de Pathologie Neuromusculaire, Institut deMyologie, and bUnite Fonctionnelle de Cardiogenetique et Myogenetique, Hopital dela Salpetriere, Paris, France

Correspondence to Daniel Hantaı, Inserm U582, Institut de Myologie, Hopital de laSalpetriere, 47 Boulevard de l’Hopital, 75651 Paris Cedex 13, FranceTel: +33 1 42165706; fax: +33 1 42165700;e-mail: d.hantai@myologie.chups.jussieu.fr

Current Opinion in Neurology 2004, 17:539–551

Abbreviations

ChAT choline acetyltransferaseCMAP compound muscle action potentialCMS congenital myasthenic syndrome

# 2004 Lippincott Williams & Wilkins1350-7540

IntroductionCongenital myasthenic syndromes (CMSs) form a

heterogeneous group of genetic diseases characterized

by a dysfunction of neuromuscular transmission. This

dysfunction causes muscle weakness, which is increased

by exertion and usually starts during childhood. The

prevalence of CMS is estimated at one in 500 000 in

Europe, and CMSs are much more uncommon than

autoimmune myasthenia [1].

Knowledge of the mechanisms underlying CMS has

increased considerably in the past 25 years, because of

the pioneering work undertaken by the group of Engel etal. [2]. Acetylcholinesterase deficiency was the first CMS

identified, based on the lack of the enzyme at

neuromuscular junctions [2]. Progressively, the patho-

physiological heterogeneity of CMS was demonstrated:

besides synaptic CMS caused by acetylcholinesterase

deficiency, pre- and postsynaptic CMS were described,

the latter including quantitative deficiency or kinetic

anomalies of the acetylcholine receptor. In the past 15

years, many gene mutations responsible for CMS were

identified, affecting the different acetylcholine receptor

subunits and the collagenic tail of acetylcholinesterase

[3,4 ..]. Mutations in the genes for choline acetyltrans-

ferase (ChAT) [5], rapsyn [6], and more recently the

sodium channel SCN4A have been reported to cause

CMS [7.].

Several reviews have been devoted to CMS, one of the

more recent being that of Engel et al. [8 ..]. The

objectives of the present review are to highlight the

principal phenotypical and pathophysiological character-

istics of CMS, to pinpoint the more recent advances in

the field, and to propose a strategy for the accurate

characterization of these disorders.

Classification of congenital myasthenicsyndromes and recent findingsThe current classification of CMS is based on patho-

physiology, i.e. on the precise identification of the

neuromuscular transmission anomaly. The location of

the dysfunction of neuromuscular transmission (Fig. 1)

[9], which is specific to the different CMSs, is either

presynaptic (generally caused by an anomaly of ChAT),

synaptic (corresponding to an anomaly of the acetylcho-

linesterase collagen tail), or postsynaptic (secondary to an

anomaly of acetylcholine receptor or rapsyn). In the

experience of Engel’s group, postsynaptic CMSs are

three times more frequent than acetylcholinesterase

deficiency and 10 times more frequent than presynaptic

539

CMS [4..]. The different classes and subclasses of CMS

will be described below with reminders of their first

descriptions and main characteristics and with an

emphasis on the latest findings.

Presynaptic congenital myasthenic syndromes

Among the presynaptic CMSs, only those caused by

mutations in the ChAT gene have been fully character-

ized, the others remain to be defined at the genetic

level.

Congenital myasthenic syndromes caused by ChAT mutations

These CMSs usually manifest at birth or in the neonatal

period with bulbar disorders and respiratory insufficiency

with apnoea [10,11] or even sudden death [11,12]. These

episodes are triggered by fever, fatigue and overexertion.

Apart from these bouts, the myasthenic signs are often

modest or not present. Cholinesterase inhibitors are

effective. Microelectrophysiology shows, after prolonged

10 Hz repetitive stimulation, a reduction in amplitude of

the miniature endplate potentials. These anomalies are

characteristic of a defect in the resynthesis of acetylcho-

line or in the filling of synaptic vesicles [10]. Ultra-

structural examination shows that, when muscle is at

rest, the synaptic vesicles are of reduced size.

Ohno and collaborators [5] described the first mutations

in CHAT, the gene encoding ChAT and located in

10q11.2. ChAT is a presynaptic protein localized in the

nerve terminals, where it catalyses acetylcholine produc-

tion. As shown in knockout mice, ChAT affects

synaptogenesis and coordinates synaptic maturation

[13]. Mutations lead to a reduction or even abolition of

the catalytic capacity of the enzyme [5]. Fourteen

mutations have been reported to date, mostly of the

missense type [5,14 .,15.]. Recent structural studies

indicate that whereas half of the missense mutations

are positioned in the molecule such that they affect

enzyme activity directly, the remaining mutations are

distant from the active site and must exert indirect

Figure 1. Pathophysiological classification of congenital myasthenic syndromes

axon terminal

basal lamina

AChR

rapsyn

muscle fibre

Na+ channel

ACh

ChAT

AChE Q, T

Presynaptic defectsDefects in ACh resynthesis (AR) CHATPaucity of synaptic vesiclesLambert-Eaton like CMS

Synaptic defectsEndplate AChE deficiency (AR) COLQCOLQ

CHAT

Postsynaptic defectsAChR kinetic anomalies slow channel syndrome (AD) α>ε, β.δ fast channel syndrome (AR)AChR deficiency (AR) AChR α>ε, β.δ rapsyn (AR) RAPSNAnomaly of muscle Na+ channel α-subunit SCN4A

RAPSN

SCN4A

α > ε, β,δ

ε>α ,β,δ

Incompletely characterized CMSCMS with plectin deficiencyFamilial limb girdle myastheniaCMS with tubular aggregates

No identified defects

CHRNB1CHRND

CHRNA1CHRNE

α

The eight genes involved in congenital myasthenic syndromes (CMSs) are named CHRNA1, CHRNB1, CHRND, CHRNE, COLQ, CHAT, RAPSNand SCN4A. The coded protein and the gene location are, respectively, as follows: (1) a (2q24–q32), b (17p11–p12), d (2q33–q34), e (17p13)subunits of acetylcholine receptor (AChR); (2) collagenic tail (ColQ) (3p24–2) of acetylcholinesterase (AChE); (3) ChAT (10q11.2); (4) rapsyn(11p11). The endplate species of acetylcholinesterase is composed of one, two, or three homotetramers of T globular catalytic subunits attached to acollagenic tail (ColQ), anchoring them in the synaptic basal lamina. Rapsyn stabilizes the acetylcholine receptor aggregates and links them to thepostsynaptic cytoskeleton. Heredity is either autosomal recessive (AR) or autosomal dominant (AD). Modified from the classification proposed by theEuropean Neuromuscular Center (ENMC) [9].

Neuromuscular disease: muscle540

effects [16 ..]. The possibility that the I336T ChAT

mutation found in three consanguineous Turkish

families was a founder was postulated [14 .].

Other presynaptic myasthenic syndromes still incompletely

characterized

The paucity of synaptic vesicles was described in one

patient with early-onset CMS. The density of acetylcho-

line synaptic vesicles was reduced by 80% and the

number of quanta released was drastically reduced [17].

The exact cause of this CMS is still unknown.

The first case of Lambert–Eaton-like CMS was first

reported in a child [18]. His myasthenic syndrome was

characterized by a good response to guanidine and by

electrophysiological anomalies identical to those of a

Lambert–Eaton syndrome, i.e. diminished action poten-

tials markedly potentiated by tetanic stimulation. A

second case presented with severe hypotonia and

respiratory distress at birth [4..]. No mutation was found

in the gene coding for the presynaptic calcium channel.

Three patients were reported with a sporadic myasthenic

syndrome with associated signs of attack of the central

nervous system (cerebellar ataxia or nystagmus) [19].

None presented, as in the Lambert–Eaton syndrome,

with a reduction of the action potentials or with

potentiation after high frequency stimulation. Microelec-

trophysiology revealed a marked reduction in the

spontaneous or nerve stimulation-induced release of

acetylcholine quanta.

Synaptic congenital myasthenic syndrome:

acetylcholinesterase deficiency

CMSs caused by acetylcholinesterase deficiency were

first described in 1977 [2]. Since then, many cases of

partial or complete deficiency of the enzyme located in

the synaptic basal lamina have been reported [20]. The

first symptoms usually arise in the neonatal period, and

the symptoms are severe with a significant lethal risk.

However, the disease may start later, during infancy, and

is not so severe. Several observations point to the

diagnosis of acetylcholinesterase deficiency: autosomal

recessive heredity, repetitive compound muscle action

potential (CMAP) after single stimulation (Fig. 2), the

absence of response to cholinesterase inhibitors, and

slowed [20] but inconstant [21] pupil responses to light.

Diagnosis using muscle biopsies is indicated by no or

poor visualization of acetylcholinesterase at the neuro-

muscular junction. Acetylcholinesterase deficiency is

related to mutations in the COLQ gene coding for the

collagenic tail of acetylcholinesterase [22–24]. At the

neuromuscular synapse, acetylcholinesterase is present

as asymmetric acetylcholinesterase, which is made up of

three homotetramers each comprising four globular

catalytic subunits linked together by a collagenic tail

(ColQ; Q for ‘queue’ in French, which means ‘tail’) of

trimeric helicoidal structure. The collagenic tail concen-

trates and anchors the enzyme within the synaptic basal

lamina.

Twenty-four recessive mutations have been described to

date (Fig. 2). They are more often homozygous than

heterozygous, and nonsense than missense [4 ..]. The

fact that the same homozygous G240X mutation was

detected in several Arab families and in one Iraqi Jewish

patient suggests that it is not uncommon in Near and

Middle East countries [25]. Depending on their localiza-

tion, COLQ mutations have different consequences: in

the N-terminal proline-rich attachment domain, they

prevent attachment of the acetylcholinesterase catalytic

subunits to the collagenic tail; in the mid-part they

prevent the trimerization of the collagenic tail; in the C-

terminal domain they most often impair anchoring of the

enzyme within the synaptic basal lamina [24,26,27.,28.].

This impaired attachment of the collagenic tail to the

synaptic basal lamina has recently been elegantly and

comprehensively tested using purified C-terminal do-

main mutant ColQ applied to frog neuromuscular

junctions [29 ..].

To date there is no effective treatment for this type of

CMS.

Postsynaptic congenital myasthenic syndromes

Among these postsynaptic CMSs, two categories are

described: CMS in connection with a kinetic anomaly of

the acetylcholine receptor; and CMS with a decreased

number of acetylcholine receptors at the neuromuscular

junction. In the latter category, besides the CMSs with

acetylcholine receptor deficiency as a result of numerous

mutations in the different acetylcholine receptor subunit

genes, those caused by mutations in the rapsyn gene

were identified recently [6] and are far from infrequent.

Congenital myasthenic syndrome caused by acetylcholine

receptor kinetic anomalies

Slow channel syndrome is the most frequent kinetic

anomaly of the acetylcholine receptor. This entity, of

autosomal dominant inheritance, is characterized by a

prolonged opening time of the acetylcholine receptor

[30]. Fifteen autosomal dominant missense point muta-

tions causing a gain of function of the acetylcholine

receptor were identified [4..]. Although most of the

mutations were found in the acetylcholine receptor asubunit [31], other subunits are also concerned [32]. The

mutations are located in two transmembrane domains

taking part in the formation of the acetylcholine receptor

pore through which passes the sodium flux [33], M1 for

mutations of the a and b subunits and M2 for those,

more frequent, affecting the a, b, d and e subunits [34];

Congenital myasthenic syndromes Hantaı et al. 541

an area of the extracellular domain of the a subunit close

to the acetylcholine binding site (mutations aG153S and

aV156M) [4 ..].

The functional consequences of the various mutations

were studied in intercostal muscle biopsy or by

expressing the mutation in cell systems [35]. The

prolonged opening time of the acetylcholine receptor is

dependent either on the slowed closing of the channel or

on the increased affinity of the acetylcholine receptor for

its ligand [36]. In addition, a new mechanism that

involves not only delayed closure but also delayed

opening of the channel in the case of a dS268F mutation

was recently published [37].

Clinical expression may vary from early onset and

severe to late onset and moderate [30,38]. The

arguments in favour of the diagnosis are autosomal

dominant heredity, no response to cholinesterase

inhibitors, and repetitive CMAP after a single stimula-

tion. The last two characteristics are also found in

acetylcholinesterase deficiency (Fig. 2). The selectivity

of muscle involvement with a prevalent atrophic deficit

of the finger extensors and of the cervical muscles is

suggestive of slow channel syndrome. Remodelling of

the ultrastructure of the endplate is observed with

calcium deposits, destruction of the postsynaptic folds,

vacuolizations and tubular aggregates [30]. The diag-

nosis leads to the therapeutic use of quinidine, a blocker

agent able to normalize the acetylcholine receptor

opening time [39]. Fluoxetine has recently been shown

to be an alternative to quinidine when the latter is not

tolerated by the patient [40 .].

A peculiar case has been reported of a slow channel

syndrome with recessive transmission, occurring in a

Figure 2. Main features of slow channel syndrome and of acetylcholinesterase deficiency

Acetylcholinesterase deficiency- autosomal recessive

- slowed pupil response

α-BGT fasciculin

control

patient

AChR AChE

AChR AChE

COLQ

In common

- no response to cholinesterase inhibitors

- repetitive CMAP after single stimulation

1 2 3 4 5 6 7 8 9 102 mV 2 ms

Repetitive CMAP

Slow channel syndrome- autosomal dominant

CHRNA1

AChR

M1 M2 M3 M4

s-s

NH2COOH

In both diseases, cholinesterase inhibitors are inefficient, and a repetitive motor response is evidenced by electrophysiological study. Recessivelytransmitted endplate acetylcholinesterase deficiency is demonstrated by the absence of fluorescent fasciculin staining at the endplate. Collagenic tailgene mutations will be determined. Slow channel syndrome is transmitted as an autosomal dominant trait. The most common mutations involveacetylcholine receptor a subunit in the pore region (transmembrane M1 and M2 domains) or in the vicinity of the acetylcholine binding site. Othermutations not shown here are within the b, d and e subunits. AChE, Acetylcholinesterase; AChR, acetylcholine receptor; a-BGT, a-bungarotoxin;CMAP, compound muscle action potential.

Neuromuscular disease: muscle542

consanguineous family in connection with a homozygous

mutation of the e subunit (eL78P) located in the

extramembrane region. This mutation was pathogenic

only if present on two alleles [41]. In addition, a slow

channel syndrome associated with a chromosomal

translocation 2q31–9p27 was described [42].

Fast channel syndromes are of autosomal recessive

transmission, although a case of autosomal dominant

transmission was reported recently [43.]. The diagnosis

is made by microelectrophysiology showing a shortening

of the acetylcholine receptor opening time [44]. Clinical

severity is variable. Arthrogryposis was reported in one

case [45]. The patients are responsive to the combina-

tion of 3,4-diaminopyridine and cholinesterase inhibitors.

Eight mutations were identified affecting a, d and esubunits and are located either in the extracellular

domain, in the M3 transmembrane domain (mutation

aV285I), or in the cytoplasmic loop between the M3 and

M4 domains (e mutations only) [8 ..]. Of the two

mutations present in the patient, one is a nonsense

mutation whereas the other is responsible for the kinetic

anomalies. This second mutation can modify the kinetics

of the receptor by various mechanisms that can be

determined on intercostal biopsy or after in-vitro

expression in cell models. A recent article thus details

the detrimental effects of a V132L mutation located in

the acetylcholine receptor a subunit within the signature

cystine loop on acetylcholine binding and channel gating

[46 ..]. The different mechanisms underlying fast chan-

nel syndromes are the topic of a recent review [47.].

Congenital myasthenic syndromes with predominant

acetylcholine receptor deficiency (with absent or only slight

kinetic anomalies)

These account for approximately half of CMS patients

[4..]. The majority are related to mutations of the

acetylcholine receptor. No peculiar clinical findings point

to this type of autosomal recessive CMS, whose severity

is variable. Nevertheless a founder effect in the Gypsy

population of the e1267delG mutation has been pro-

posed [48]. An extensive study on five disease loci in the

different Gypsy groups has demonstrated a strong

founder effect and a carrier rate of 3.74% for this

mutation [49].

Cholinesterase inhibitors are most often active and 3,4-

diaminopyridine can provide additional benefit.

The described mutations are numerous (60 or more),

either homozygous or heterozygous [4 ..]. They are of all

types: missense mutations, chromosomal deletions,

insertions, deletions. The mutations are located on the

whole gene encoding the acetylcholine receptor esubunit, most being located in the extracellular domain

and in the cytoplasmic loop between the M3 and M4

transmembrane domains [4 ..]. Recently, a chromosomal

microdeletion was identified for the first time in CHRNE[50], showing that this type of mutation may be missed

by standard screening techniques. Lately a frameshift

mutation in exon 7 of CHRNE (e553del7) was shown to

provoke skipping of the preceding exon both in muscle

tissue and when expressed in COS cells [51 .].

Mutations in the promoter were also described [52,53].

Interestingly, the injection of the corresponding recom-

binant in the rat allowed the authors to demonstrate that

a mutation in an N-box of the CHRNE promoter leads to

less acetylcholine receptor synaptic expression [50].

Another experimental approach, namely the cell expres-

sion of green fluorescent tagged acetylcholine receptor,

allowed others to show that mutations affecting cysteine

470 of the e subunit prevent acetylcholine receptor

surface expression [54].

More rarely, other subunits of acetylcholine receptor a, band d subunits are implicated [4..]. The preponderance

of mutations of the e subunit may be caused by the

possibility of the re-expression of the g fetal acetylcho-

line receptor isoform in the case of null mutations of

CHRNE [55,56].

Curiously, CMS not only affects humans but also South

African Red Brahman calves. These calves suspected of

myasthenia have been shown to bear a homozygous 20

basepair deletion mutation in bovine CHRNE. This

mutation leads to a non-functional allele and a severe

phenotype [57,58.].

Congenital myasthenic syndromes with mutations of the rapsyn

gene (RAPSN)

These were first identified in 2002 [6]. Rapsyn is a

43 000 Mr postsynaptic cytoplasmic protein, which

participates in acetylcholine receptor assembly at the

neuromuscular junction [59] and allows its anchoring to

the cytoskeleton by b-dystroglycan among other mole-

cules [60]. Most mutations of this gene located in 11p11

were identified in the tetratricopeptide repeat domain,

and cell expression studies revealed that the co-

expression of mutant rapsyn and acetylcholine receptor

subunits impair the recruitment of acetylcholine recep-

tors to rapsyn clusters, an essential step for the anchoring

of the acetylcholine receptor to the cytoskeleton [6].

These mutations are responsible for a reduction of

rapsyn and consequently of the acetylcholine receptor

itself at the neuromuscular junction. The reduction in

rapsyn expression is not specific, because it is also

observed in primary acetylcholine receptor deficiencies.

The inheritance of this CMS is autosomal recessive.

Since the first four cases were published, nearly 50 other

cases have been reported [61–63,64.,65,66 .,67.,68]. Half

Congenital myasthenic syndromes Hantaı et al. 543

of them bear the homozygous N88K. The other half

bears N88K on one allele and a second mutation on the

other allele. This second mutation is localized all along

the rapsyn molecule, and nearly 20 different mutations

have been identified to date (Fig. 3a) [6]. Missense

mutations predominate (approximately two-thirds of

cases). When the second mutation is not identified by

direct sequencing, the search for a chromosomal micro-

deletion of RAPSN is recommended [69].

Two E-box mutations were identified in the rapsyn

promoter [64.] (Fig. 3b). Seven of the eight patients

reported originated from the Jewish population of Iraq

and Iran and had already been described for their

peculiar clinical phenotype: benign CMS with facial

malformations (mandibular prognathism, elongated face)

[70].

A founder effect of the frequently identified N88K

mutation is likely at least in the European or Indo-

European population [61,62,71..], although the exis-

tence of other founders has been proposed [72]. The

high frequency of the N88K mutation may lead to cases

of pseudo-dominant inheritance.

Genotype–phenotype correlation is not easy. Analysis

of the corpus of clinical observations confirms the

existence of two phenotypes: (1) a neonatal form,

even antenatal (with arthrogryposis multiplex congeni-

ta), with major respiratory disorders and severe

progression of the disease; and (2) mild forms

beginning during childhood or in adulthood. On the

basis of 16 cases, a distinction between early and late

onset was proposed [66.]. The importance of the

identification of the late-onset cases is to avoid

improper immunotherapy. Patients with the rapsyn

mutation responded well to cholinesterase inhibitors

[66 .] or to the combination of cholinesterase inhibitors

and 3,4-diaminopyridine [67.].

In summary, mutations in RAPSN and the resulting

rapsyn deficiency appear to be an important cause of

Figure 3. Diagram depicting the main domains of rapsyn and the localization of the identified mutations

Tetratricopeptide repeats (TPR) necessaryto rapsyn self-association

AChR

TPR1 TPR2 TPR3 TPR4 TPR5 TPR6 TPR7 coiledcoil RING

Q3K

L14P

46in

sCA

25V

F81

LY

86X

N88

KR

91L

C97

X

Q12

4X

A14

2DR

151P

V16

5M

553i

ns5

IVS

4-2A

→G

A24

6V

Y26

9X

G29

1D

E33

3X

1083

_108

4dup

CT

1177

delA

A

(a)

(b)

–38A→G –27C→G

E box E box

β-d

ystr

ogly

can

(a) Seven tetratricopeptide repeat domains (TPR1–7) are necessary for rapsyn to self-associate. The coiled-coil domain binds to the large cytoplasmicloop of the acetylcholine receptor (RACh) subunits. The RING domain binds rapsyn to b-dystroglycan. A serine phosphorylation site is located atcodon 406. Modified from Ohno et al. [6]. (b) Two E-box (CANNTG-type sequence, on grey background), to which myogenic factors can bind arelocated upstream of the transcription initiation site in the rapsyn gene promoter region. Localization of the two different rapsyn promoter mutations.Adapted from Ohno et al. [64.].

Neuromuscular disease: muscle544

CMS associated with endplate acetylcholine receptor

deficiency.

Congenital myasthenic syndrome with plectin deficiency

Plectin is a highly preserved structural protein of the

cytoskeleton expressed in several cell types, including

skeletal muscle and the postsynaptic membrane. Plectin

deficiency was described in a patient presenting with

progressive myopathy, associated with myasthenic syn-

drome (involving facial, limb and oculomotor muscles),

and epidermolysis bullosa [73]. The pathophysiology of

this CMS is poorly understood.

Congenital myasthenic syndrome caused by a mutation in the

sodium channel SCN4A

The case was recently reported of a 20-year-old patient

presenting since birth with very short bouts (3–30 min)

of respiratory distress and bulbar paralysis [7 .]. The

diagnosis was made by electrophysiology of the inter-

costal muscle, which revealed the impossibility of

evoking an action potential after nerve stimulation.

Two mutations of SCN4A were identified, including

only one (V1442E) located in the S3/S4 extracellular

domain, which was found to be pathogenic when

expressed in HEK cells. The clinical aspect is quite

different from that usually associated with a SCN4Amutation (dyskalemic paralysis, congenital paramyot-

ony).

Incompletely characterized congenital myasthenic

syndromes

These CMSs are described on clinical or histological

grounds, but their molecular origin and more generally

their pathophysiology remain unknown in the absence of

an exhaustive exploration.

Familial limb girdle myasthenia

Several families have been reported [74,75]. This

previously named ‘myasthenic myopathy’ is of recessive

inheritance. Clinically, the absence of oculobulbar signs

was remarkable. The weakness and fatigability involved

the girdles. The peculiarity of this not yet understood

entity was recently stressed with the publication of five

cases, who all presented with tubular aggregates in their

muscle biopsy and who all responded favourably to

cholinesterase inhibitors [76].

Congenital myasthenic syndrome with tubular aggregates

This CMS is associated with tubular aggregates at the

histological muscle examination. The case of three

sisters presenting with a slowly progressive myopathy

beginning in early childhood associated with cardio-

myopathy was reported. A favourable response to

cholinesterase inhibitors was noted [77]. Similar char-

acteristics were described in another family [75]. A

sporadic case was reported recently [78]. In the absence

of thorough investigations of neuromuscular transmis-

sion, the classification of these cases remains delicate,

more especially as the presence of tubular aggregates is

not specific and can be associated with isolated

myopathy, painful cramps [79] and with slow channel

syndromes [31].

Approach to the diagnosis of congenitalmyasthenic syndromesOn the basis of these historical advances in the knowl-

edge of CMS, the diagnostic strategy includes roughly

two successive steps: (1) the association of a clinical-

electrophysiological picture of a myasthenic syndrome,

and data in favour of a congenital origin; and (2) the

recognition of the pathophysiological type, which is

based on clinical data, the type of hereditary transmis-

sion, the response to cholinesterase inhibitors, the results

of electromyography, and finally the muscle biopsy and

molecular genetics. The sequential order of these two

last investigations depends upon the initial clinical-

electromyographical data.

Clinical presentation

The various CMSs share a common clinical presenta-

tion. The onset is in general early. Late appearance of

the symptoms during adolescence, or even in the adult,

is more rarely reported. Some clinical signs suggest an

anomaly of neuromuscular transmission: ophthalmople-

gia and ptosis, dysphonia and swallowing disturbance,

facial paresis, and muscle fatigability. In the young child,

the ptosis is not easy to recognize because hypotonia,

poor mimicry, suction disorders, and weakness of the cry

are in the foreground. The occurrence of bouts and

worsening by exertion are characteristics of the disease.

The favourable effect of cholinesterase inhibitors is a

significant argument in favour of a myasthenic syn-

drome. However, two types of CMS are worsened by

cholinesterase inhibitors: slow channel syndrome and

acetylcholinesterase deficiency. With the proper

myasthenic signs, myopathic signs are often associated:

amyotrophy, tendinous retractions, facial malformation

and scoliosis. The severity of the CMS is variable,

depending upon the severity of the walking deficit, the

bulbar disorders and the respiratory difficulties. Acute

respiratory failure may occur, triggered by infectious

episodes, and is frequent in the first months of life. In

the absence of respiratory assistance, the risk of death is

high [12].

A family history of the disease is an essential argument in

favour of the genetic origin of myasthenic syndrome.

Most CMSs are of autosomal recessive inheritance. Slow

channel syndrome is the only autosomal dominant CMS

characterized hitherto. The progression patterns of

CMSs are highly variable, including in a given patient,

at various periods of life. Myasthenic bouts are fre-

Congenital myasthenic syndromes Hantaı et al. 545

quently triggered by infectious episodes, pregnancy and

even periods. Progressive aggravation of the disease may

sometimes occur late in adulthood, with the appearance

of respiratory insufficiency [27 .]. A favourable progres-

sion is possible after a severe neonatal onset [4..].

Titration of anti-acetylcholine receptor antibodies in the

serum

The absence of antibodies against acetylcholine receptor

and muscle-specific receptor tyrosine kinase [80,81.] is a

prerequisite for the diagnosis of CMS, although an

exception was reported [82 .].

Electromyography

The electrophysiology of neuromuscular transmission is

the determinant for the diagnosis of CMS. This includes

searching for neuromuscular block, repetitive motor

responses and increments [83,84]. The observation of a

neuromuscular transmission block affirms the myasthe-

nic syndrome. The decrement can be absent in CMS,

particularly in patients who are not highly symptomatic,

and in cases of CMS caused by mutations in ChAT [5] or

rapsyn [6]. In the case of a ChAT deficiency, the

decrement may appear only after an initial 5-min 10 Hz

stimulation [14 .]. Repetitive CMAPs are pathognomonic

of two varieties of CMS: slow channel syndrome and

acetylcholinesterase deficiency (Fig. 2). The search for

an increment is imperative. An increment greater than

100% in amplitude and in area is suggestive of a

presynaptic origin.

Muscle biopsy

A first role of the muscle biopsy is to eliminate the

diagnosis of myopathy (congenital myopathy or mito-

chondrial cytopathy). Although non-specific, the pre-

dominance of type I fibres and the marked atrophy of

type II fibres is suggestive of CMS. The presence of

tubular aggregates is frequent, but poses the problem of

the group of CMSs with tubular aggregates. The

neuromuscular junctions are visualized for (acetyl)choli-

nesterase by the histochemical technique of Koelle,

fasciculin or specific antibodies, and for acetylcholine

receptor by fluorescent a-bungarotoxin, which binds to

it. The neuromuscular junctions frequently exhibit

variable anomalies: reduced size, the disappearance of

synaptic folds, all modifications are not specific, however,

to a given CMS.

Two types of information are decisive: (1) the absence of

acetylcholinesterase at the neuromuscular junction

establishes the diagnosis of acetylcholinesterase defi-

ciency; a study by ultracentrifugation on sucrose gradient

will generally reveal the absence of asymmetrical

(synaptic) forms of the enzyme; and (2) a significant

reduction in the number of acetylcholine receptors,

further quantified by binding with iodinated a-bungar-

otoxin, points to a primary anomaly of acetylcholine

receptor or rapsyn.

The expression of the fetal g subunit of the acetylcho-

line receptor argues in favour of a primary anomaly of

the acetylcholine receptor e subunit [55,56]. Immuno-

cytochemical study of the expression of other markers of

the neuromuscular synapse can also be performed: agrin,

muscle-specific receptor tyrosine kinase, rapsyn, neur-

egulin, a-dystrobrevin or utrophin [85]. It is aetiologi-

cally suggestive if there is a major and selective

reduction of the expression of a protein, but the primary

nature of the deficit is, however, not established (a

deficit in rapsyn is found in CMS with mutations in both

the rapsyn gene and the acetylcholine receptor subunit

genes).

Molecular genetics

The diagnosis of CMS can be confirmed by molecular

analyses in the eight genes whose mutations are so far

known to cause CMS: four genes encoding the various

acetylcholine receptor subunits (CHRNE, CHRNA1,CHRNB1, CHRND), the genes encoding rapsyn

(RAPSN), the collagen tail of acetylcholinesterase

(COLQ), choline acetyltransferase (CHAT), and the

sodium channel (SCN4A). With the exception of the

Gypsy e1267del mutation [48] and the RAPSN N88K

mutation [6,71 ..], a search has been made to identify

‘private’ mutations. Analysis of the coding sequences

and flanking intronic regions by direct sequencing after

polymerase chain reaction amplification of each fragment

on genomic DNA is required. Many mutations have

been described to date, and the two predominant genes

in postsynaptic CMS appear to be CHRNE and RAPSN.In approximately half of the cases, the analysis of these

genes does not identify a mutation causing the disease,

suggesting that other genes could be involved. When a

new mutation is identified, its pathogenic character can

be demonstrated by expression studies of this mutation

in HEK cells, COS cells or oocytes, but other experi-

mental models can also be used.

Microelectrophysiology of neuromuscular transmission

Microelectrophysiology of the intercostal muscle can be

used to specify the pre- or postsynaptic location of the

dysfunction of neuromuscular transmission, and in

postsynaptic CMS, to find kinetic anomalies of the

acetylcholine receptor [4..]. The complexity of these

techniques (patch clamp) and the risks of general

anaesthesia in a myasthenic patient limit the indications

for this exploration, more especially because the expres-

sion of the mutations in experimental cell systems by

itself allows the pathophysiological characterization of

CMS. In addition, the study of other muscles under local

anaesthesia was proposed: quadriceps [86] and ancone

[15].

Neuromuscular disease: muscle546

Difficulties of diagnosisThe diagnosis of CMS is not always easy. Faced with

sporadic CMS beginning after the neonatal period, the

diagnosis of seronegative autoimmune myasthenia may

be difficult to eliminate, more especially as long periods

of remission are possible in both afflictions and bouts can

occur in the adult CMS patient during pregnancy [38].

Muscle-specific receptor tyrosine kinase (MuSK) anti-

bodies have been detected in more than

half of the patients presenting with seronegative

(no acetylcholine receptor antibodies) auto-immune

myasthenia [80,81 .]. In case of uncertainty, the absence

of MuSK antibodies must be verified before establishing

a diagnosis of CMS.

Three recent observations have stressed that it is

sometimes difficult to draw clear boundaries between

autoimmune myasthenia and CMS [82,87,88.]. The first

reported two sisters carrying heteroallelic mutations of

the acetylcholine receptor a subunit, both presenting

with neonatal myasthenic syndrome, but one developed

autoimmune myasthenia as an adult, attested by the

transitory presence of anti-acetylcholine receptor anti-

bodies and a favourable response to plasmaphereses and

corticotherapy [82]. The authors suggested that the

genetic anomaly of the acetylcholine receptor could

constitute the factor triggering autoimmune myasthenia.

The second observation concerned a patient presenting

with acquired slow channel syndrome beginning at 30

years of age. The passive transfer of the serum of this

patient to a mouse reproduced kinetic anomalies of the

acetylcholine receptor, which demonstrated its autoim-

mune origin and excluded a congenital affliction [87].

The third observation reported an acetylcholine recep-

tor-seronegative, MuSK-seropositive myasthenia gravis

patient, in whom no acetylcholine receptor or MuSK

deficiency was found in muscle biopsies despite the

electrophysiological impairment. Mutation analysis of

MUSK did not reveal mutations but polymorphisms. The

authors concluded that circulating anti-MuSK antibodies

may not have caused the myasthenic syndrome in this

patient [88.].

Phenotype–genotype correlations andprognosisThe genotype and clinical phenotype are not correlated

in CMS. Mutations in different synaptic proteins give

similar clinical pictures: the occurrence of apnoeic

episodes in early childhood was reported in CMSs with

a deficit in ChAT, in those caused by primary anomalies

of the acetylcholine receptor, of the acetylcholinesterase

or of rapsyn. Arthrogryposis has been described in CMS

caused by mutations in the gene encoding rapsyn [66.]

and the acetylcholine receptor d subunit [45]. The same

mutation could lead to very different clinical phenotypes:

for example, the homozygous N88K rapsyn mutation

leads either to a very severe neonatal form or to a late-

onset and benign form [6,66 .]. Finally, variability within a

family was noted in certain cases of CMS.

Prognosis is difficult to assess. A favourable outcome is

possible in cases of CMS initially thought to be severe

because of respiratory or bulbar bouts. In contrast, motor

and respiratory degradation occurring late in adulthood

has been reported in patients initially only slightly

affected [27.]. As indicated above, knowledge of the

primary molecular anomaly of CMS does not enable the

prediction of disease progression. The response to

treatments known to ameliorate neuromuscular transmis-

sion is a significant prognostic factor: thus in acetylcho-

linesterase deficiency, the absence of amelioration by

cholinesterase inhibitors or any other drug may be

alarming.

TreatmentNon-specific measures are essential: immediate treat-

ment of respiratory distress, the prevention of infections

and of malnutrition as a result of swallowing disorders,

and orthopaedic surveillance of spinal complications and

retractions. Drug contraindications must be respected as

for any other myasthenic syndrome. In the case of CMS,

there is no reason to apply the immunosuppressive

therapy used for myasthenia gravis. Cholinesterase

inhibitors are efficient in all CMSs, with the exception

of slow channel syndrome and acetylcholinesterase

deficiency, which they can even worsen. They exert a

preventative effect on the respiratory decompensations

of CMS caused by ChAT mutations [4 ..]. 3,4-Diamino-

pyridine, whose mode of action is presynaptic, is

sometimes effective in pre- or postsynaptic CMSs [39].

Patients suffering from slow channel syndrome benefit

from the regulatory action of acetylcholine receptor

blockers: quinidine is effective by correcting the

prolonged opening of the acetylcholine receptor [89],

but is formally contraindicated in all the other forms of

CMS. A favourable effect of fluoxetine was recently

demonstrated in some patients, and is of interest despite

the large amount needed [40 .]. At present, there is no

specific treatment for acetylcholinesterase deficiency.

ConclusionAlthough the epidemiology of the CMSs is poorly

understood, these disorders constitute the major cause

of the myasthenic syndrome in the young child and are a

minor cause of adult myasthenic syndrome. The

diagnosis is often difficult to ascertain because of the

frequent absence of a family history of the disease, and

because of the pre-eminence of the myopathic signs

compared with myasthenic signs. The early onset of the

first symptoms, the presence of fluctuations, the

demonstration of a neuromuscular block, repetitive

CMAP after single stimulation, and the cholinesterase

Congenital myasthenic syndromes Hantaı et al. 547

inhibitor test all enable the rectification of the diagnosis

and the proposal of an effective treatment and genetic

counseling. The numerous studies devoted to CMS over

more than 20 years have demonstrated the patho-

physiological heterogeneity of CMS. Characterization

of the CMS is based on the mode of transmission, the

search for a CMAP, the positive or negative response to

cholinesterase inhibitors, the study of motor endplates,

which is easily done on a deltoid muscle biopsy, and

molecular genetics. It will thus be possible to identify

the majority of CMSs: a primary anomaly of one of the

various acetylcholine receptor subunits, of rapsyn,

acetylcholinesterase, ChAT or even SCN4A. However,

the origin of a significant fraction of CMSs remains

unknown. Numerous molecules of the neuromuscular

junction are potential candidates for CMS and may be

tested (Fig. 4) [90]. Therefore, in non-identified cases,

various investigations will be used: genetic linkage

analysis in the case of large families, the demonstration

in the muscle biopsy of a selective deficit in one given

synaptic molecule, or microelectrophysiology of the

intercostal muscle. Collaboration between clinicians,

morphologists, geneticists, and neurobiologists is essen-

tial for a complete characterization of the CMSs and for

the understanding of the fundamental mechanisms of

neuromuscular transmission based on human pathology.

AcknowledgementsWe thank Claire Legay and Hanns Lochmuller for critical reading of themanuscript. This study was supported by the Direction de la RechercheClinique de l’Assistance Publique, Hopitaux de Paris (PHRC #AOM01036), Association Francaise contre les Myopathies, GIS-MaladiesRares, and Reseau Inserm de Recherche Clinique.

Figure 4. Neuromuscular junction molecules that are or might be involved in congenital myasthenic syndrome

α -CGRP

acetylcholineChAT

α 2, α 4, β 2, γ 1 lamininα 3, α 4, α 5 type IV collagenAChE Q, Tagrinneuregulin

ankyrindystrophin

Na+ channel

AChR-α ,β ,δ ,εα 7A, B,β 1 integrinα ,β dystroglycanMuSKErbB2, ErbB4

rapsynutrophin

plectin

K+ channelCa++ channel

ErbB3

synaptobrevinsynaptotagminsynaptophysin

Molecules with proved implication in congenital myasthenic syndromes are indicated in bold letters. The molecules, the genes of which are potentialcandidates, are indicated in normal letters. AChE, Acetylcholinesterase; AChR, acetylcholine receptor; MuSK, muscle-specific receptor tyrosinekinase. Adapted from Sanes and Lichtman [90].

Neuromuscular disease: muscle548

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