Eye muscle sparing by the muscular dystrophies: Lessons to be learned?

Eye Muscle Sparing by the Muscular Dystrophies: Lessons toBe Learned?FRANCISCO H. ANDRADE,1* JOHN D. PORTER,1–3 AND HENRY J. KAMINSKI1,2

1Departments of Neurology, Case Western Reserve University, University Hospitals of Cleveland, and Department of Veterans AffairsMedical Center, Cleveland, Ohio 441062Department of Neurosciences, Case Western Reserve University, University Hospitals of Cleveland, and Department of Veterans AffairsMedical Center, Cleveland, Ohio 441063Department of Ophthalmology, Case Western Reserve University, University Hospitals of Cleveland, and Department of VeteransAffairs Medical Center, Cleveland, Ohio 44106

KEY WORDS extraocular muscle; animal models; dystrophin; sarcoglycans; dystroglycans;laminins

ABSTRACT The devastating consequences of the various muscular dystrophies are even moreobvious when a muscle or muscle group is spared. The study of the exceptional cell or tissueresponses may prove to be of considerable value in the analysis of disease mechanisms. The smallmuscles responsible for eye movements, the extraocular muscles, have functional and morphologicalcharacteristics that set them aside from other skeletal muscles. Notably, these muscles are clinicallyunaffected in Duchenne/Becker, limb-girdle, and congenital muscular dystrophies, pathologies dueto a broken mechanical or signaling linkage between the cytoskeleton and the extracellular matrix.Uncovering the strategies used by the extraocular muscles to ‘‘naturally’’ protect themselves in thesediseases should contribute to knowledge of both pathogenesis and treatment. We propose thatcareful investigation of the cellular determinants of extraocular muscle-specific properties mayprovide insights into how these muscles avoid or adapt to the cascade of events leading to myofiberdegeneration in the muscular dystrophies. Microsc. Res. Tech. 48:192–203, 2000. r 2000 Wiley-Liss, Inc.

INTRODUCTIONThe muscular dystrophies are a group of genetic

diseases presenting primarily with progressive muscleweakness and wasting. A subset of muscular dystro-phies includes mutations of a transmembrane proteincomplex, the dystrophin-glycoprotein complex (DGC),that constitutes the linkage between the cytoskeletalelements and the extracellular matrix: dystrophin,dystroglycans, sarcoglycans, syntrophins, dystrobre-vin, and laminin-2 (merosin) (Fig. 1). These pathologiesgive rise to the following phenotypes: Duchenne/Beckermuscular dystrophy (DMD, dystrophin deficiency), limb-girdle muscular dystrophy types 2C, 2D, 2E, and 2F(LGMD, g, a, b and d sarcoglycan deficiency, respec-tively) and ‘‘classical’’ congenital muscular dystrophy(CMD, laminin-2 mutation). Although each particulargenetic condition presents characteristic clinical de-tails, the histopathological similarities among patientssuggest common pathogenic pathways. Despite consid-erable efforts, the pathological cascades triggered bythe protein defects remain unclear and there is noeffective treatment for these disorders. Hopefully, spon-taneous and genetically engineered animal models forthese muscular dystrophies will continue to be ofconsiderable value in elucidating disease mechanismsand potentially important treatment strategies.

The reasons for the exceptional sparing or targetingof particular muscle groups in neuromuscular diseaseis an important unresolved question whose answer canprobably be used to the advantage of both basic andclinical scientists (Emery, 1994; Porter and Baker,1996; Porter et al., 1997). The ocular motor system is

differentially affected by various neuromuscular disor-ders such as myasthenia gravis, Grave’s disease, amyo-trophic lateral sclerosis, and muscular dystrophy (Bahnand Heufelder, 1993; Kaminski and Ruff, 1997; Kamin-ski et al., 1992; Mitsumoto et al., 1998). The preferen-tial targeting or sparing of the ocular motor system inthese pathologies may be a function of the uniqueproperties of the extraocular muscles, the ocular motorneurons, the requirements of the visual system, or theparticular disease mechanism. In the context of themuscular dystrophies, the extraocular muscles, respon-sible for movement of the eyes, do not exhibit thepattern of necrosis, fibrosis, and regeneration seen inmost skeletal muscles in either human or animalmodels of these disorders. We propose that understand-ing the cellular strategies that spare the extraocularmuscles in the muscular dystrophies may be exploitedin devising systemic treatments for this group of dis-eases.

BIOLOGY OF EXTRAOCULAR MUSCLESMuch of our knowledge of the responsiveness of

skeletal muscle to neurogenic or myogenic disease is

Contract grant sponsor: National Institutes of Health; Contract grant num-bers: EY11998, EY09834, P30EY11373 (Core Facility for the Visual Sciences);Contract grant sponsor: Research to Prevent Blindness; Contract grant sponsor:Muscular Dystrophy Association; Contract grant sponsor: Department of Veter-ans Affairs Merit Review Award; Contract grant sponsor: Evenor ArmingtonFund; Contract grant sponsor: Ohio Lions; Contract grant sponsor: KnightsTemplar Eye Foundation, Inc.

*Correspondence to: F.H. Andrade, Department of Neurology, Case WesternReserve University and University Hospitals of Cleveland, 11100 Euclid Ave.,Cleveland, OH 44106. E-mail: [email protected]

Received 13 August 1999; accepted in revised form 10 September 1999

MICROSCOPY RESEARCH AND TECHNIQUE 48:192–203 (2000)

r 2000 WILEY-LISS, INC.

based upon information from studies of limb and respi-ratory muscles. In these muscles, ‘‘prototypical’’ musclefiber types respond to diseases with characteristicpathological changes that can be reliably used in diag-nosis. By contrast, some specialized muscle groupsexhibit fundamental departures from ‘‘typical’’ skeletalmuscle structure and function. Fiber type characteris-tics that have been so clearly defined by a concept assimple as ‘‘color’’ (e.g., red, intermediate, and white)suddenly are not so easily resolved. In these specializedmuscles, the classically defined neurogenic and myo-genic pathologic al changes may either be accentuatedor not present at all, and thus may not be of the samediagnostic value. The extraocular muscles representone of these muscle groups in which a unique pheno-type may explain unique responses in disease (Porterand Baker, 1996).

Ocular Motor SystemThe extraocular muscles (four rectus and two oblique

muscles per eye) form the effector arm of the ocular

motor system. These muscles are organized as ‘‘yokedpairs,’’ a term that describes their spatial arrangementand the precise functional coordination between ago-nist and antagonist muscles of both eyes. The overallbehavior of the ocular motor system is geared tomaintain binocular fixation upon visual targets. Thismust be accomplished within very fine tolerances orvisual function deteriorates. There are five ocular mo-tor subsystems: vestibulo-ocular and optokinetic re-flexes, saccades, pursuit, and vergence (Robinson, 1981).Although each subsystem has a distinct central sub-strate, all share the same ‘‘motor plant’’ (extraocularmuscles, motor neurons, eyeball, and the orbital suspen-sory tissues). Some characteristics of the ocular motorplant are of particular interest. As in other motorsystems, the firing rate of the motor neurons innervat-ing a particular extraocular muscle increases when theeye is moving in the ‘‘on’’ direction for this muscle.However, around 18% of motor neurons never ceasefiring for any eye position (Robinson, 1981). Further-more, when the eye is in the ‘‘primary position’’ (looking

Fig. 1. A current model of the organization of the DGC, linking thecytoskeleton to the extracellular matrix. Dystrophin links the cytoskel-eton to b-dystroglycan, the syntrophins and dystrobrevin. b-Dystrogly-can binds a-dystroglycan, and loosely associates with the sarcoglycan

complex (a, b, g, d, possibly e), which in turn links to sarcospan. Theheterotrimeric protein (a2b1g1) laminin-2 is the extracellular ligandof a-dystroglycan.

193DYSTROPHIES SPARE EXTRAOCULAR MUSCLES

straight ahead), more than 70% of all the ocular motorneurons are active, with an average firing rate ofapproximately 100 Hz (King et al., 1981; Robinson,1970). These activity levels are not common in othermotor systems, where high motor neuron activity spurtsoccur only at the initiation of movement (Hennig andLømo, 1985).

Cell Biology of Extraocular MusclesThe extraocular muscles exhibit the greatest diver-

sity amongst mammalian skeletal muscles. Thesemuscles have a unique embryonic origin (Burden, 1998;Couly et al., 1992; Wahl et al., 1994) and develop underthe influence of a variety of factors that are not necessar-ily operative for other skeletal muscles (Brueckner andPorter, 1998; Brueckner et al., 1996, 1999). Mesodermalprimordia condense around the developing eye andinitially proceed through the same general scheme ofmyogenesis seen in other skeletal muscles (Brueckneret al., 1996; Porter and Baker, 1992). Some of thesubsequent events in fiber differentiation diverge fromthe typical skeletal muscle pattern. For example, extra-ocular muscle fiber types retain ‘‘developmental’’ pro-tein isoforms, including embryonic myosin, fetal acetyl-choline receptor (g subunit instead of the adult esubunit) and sarcolemmal-wide expression of polysi-alated neural cell adhesion molecules. Whether theseexpression patterns reflect a ‘‘developmental arrest’’ orsimply an adaptation of so-called embryonic proteinsfor adult functions is not yet clear. The maturation ofthe DGC and its regional specializations clearly isunder the influence of both activity-dependent andactivity-independent mechanisms in most skeletalmuscles (McArdle et al., 1998). The extent to which thevarious elements of the DGC in extraocular musclefollow the expression patterns seen in other skeletalmuscles is a largely unexplored issue (see below).

The extraocular muscles do not conform to tradi-tional classifications of muscle fiber types, based mostlyon myosin isoform expression. A current classificationscheme specific for extraocular muscles includes sixdistinct fiber types based upon: (1) distribution intoorbital and global layers (close to the bony orbit and theeyeball, respectively); (2) innervation status, single vs.multiple nerve contacts per fiber; and (3) mitochondrial/oxidative enzyme content. This scheme identifies or-bital singly, orbital multiply, global red singly, globalintermediate singly, global pale singly, and global multi-ply innervated fiber types (Fig. 2) (Porter and Baker,1996; Porter et al., 1995, 1997; Spencer and Porter,1988). This diversity of fiber types could represent anadaptation to the functional requirements of the ocularmotor system, or simply a developmental consequenceof the anatomical location of the extraocular muscles(Porter and Baker, 1996; Sohal et al., 1998). However,the normal development of at least one of the fibertypes, the orbital singly innervated fibers, seems todepend on environmental cues during early life (Brueck-ner and Porter, 1998; Brueckner et al., 1999).

Physiology of Extraocular MusclesFunctionally, the extraocular muscles have been char-

acterized as being ‘‘weak,’’ ‘‘fast,’’ and ‘‘fatigue resis-tant.’’ Under isometric conditions, extraocular muscleshave very short contraction (time required to reach

peak twitch force) and half-relaxation (time from peakto half-peak twitch force) times compared to prototypi-cal fast muscles like the extensor digitorum longus.Force measured during maximal tetanic contractions ofextraocular muscles are just fractions of those obtainedfrom limb muscles, even when normalized to musclecross-sectional area (Fig. 3) (Close and Luff, 1974;Frueh et al., 1994; Luff, 1981; Lynch et al., 1994).Moreover, their twitches are unusually shallow and thetwitch-to-tetanus ratio is lower than in most muscles(Close and Luff, 1974; Frueh et al., 1994; Luff, 1981).This characteristic is particularly evident when theforce responses to increasing stimulation frequency areplotted, and a shift down and to the right in theforce-frequency relationship becomes obvious. All theseproperties of extraocular muscles may reflect a combina-tion of factors: (1) faster than normal calcium tran-sients during contraction, accomplished by the abun-dant sarcoplasmic reticulum (Asmussen and Gaunitz,1981); (2) displacement of contractile material by otherintra- and extracellular structures; (3) the presence ofless readily excitable non-twitch fibers; (4) differencesin the kinetics of actomyosin, a possibility at least inthose fibers that express extraocular muscle-specificmyosin (Sartore et al., 1987; Wieczorek et al., 1985).

The passive mechanical load that the extraocularmuscles work against is relatively small (eyeball itself,suspensory connective tissue). This has been one rea-son for explaining the resistance to fatigue demon-strated by the extraocular muscles (Fuchs and Binder,1983). This argument neglects the extra load producedby the co-activation of antagonistic muscles during eyemovements (Collins et al., 1975; Robinson, 1981). More-over, since the absolute force that the extraocularmuscles can generate is also very small (Fig. 3), itwould be more relevant to present the passive mechani-cal load as a fraction of the maximal force produced bythe muscles. Most skeletal muscles work around 25–40% of their maximal force (or velocity), maximizingpower and efficiency (Rome, 1998). Presumably, theoverall design of the eye and its motor system is suchthat the extraocular muscles work within this optimalrange.

SARCOLEMMAL ORGANIZATION OF THEEXTRAOCULAR MUSCLES:AN INCOMPLETE PICTURE

Although the precise functional role of the DGC is yetto be resolved, it is clear that its integrity is vital tomyofiber development and survival. In seeking anexplanation for the overt sparing of extraocular musclein a variety of types of muscular dystrophy, the extremeview might be that extraocular muscles are so differentfrom other skeletal muscles that the DGC may beabsent from this muscle group. However, limited immu-nocytochemical analyses of the DGC of extraocularmuscle have shown that dystrophin is normally presentin these muscles and that the full-length protein isabsent in DMD and in its animal models (Khurana etal., 1995; Matsumura et al., 1992).

We have examined the normal distribution of compo-nents of the DGC in mouse extraocular muscles. First,dystrophin and its homologue, utrophin, exhibit theirnormal distribution to the entire sarcolemma and neu-romuscular junction, respectively (Fig. 5). The direct

194 F.H. ANDRADE ET AL.

dystrophin binding partner, b-dystroglycan, and itsextracellular partner, a-dystroglycan, also are presentin adult extraocular muscle. Finally, the four sarcogly-cans (a, b, g, and d) co-localize with dystrophin and thedystroglycans to the sarcolemma. This distribution ofdystrophin-glycoprotein complex components, identicalto that of traditional skeletal muscles, is observed forall extraocular muscle fiber types, even the phylogeneti-cally primitive multiply innervated fiber types. Re-cently, Crosbie and coworkers (Crosbie et al., 1999)showed that a new DGC component, sarcospan, local-izes to the sarcolemma in both limb and extraocularmuscle.

Collectively, these data suggest that the DGC partici-pates in the same functional roles, sarcolemmal integ-rity, transmembrane signaling, or other as yet undiscov-ered functions, in extraocular muscles as in otherskeletal muscles. In either dystrophin or sarcoglycandeficiency, pathology may be dependent upon the second-ary displacement of other components of the proteincomplex from the sarcolemma. Currently, there is littleunderstanding of such secondary consequences or poten-

tial adaptive changes in the dystrophin-sarcoglycancomplex of extraocular muscles and of how such changesmight participate in the rescue of this muscle group.

LACK OF HISTOPATHOLOGICAL CHANGESIN DYSTROPHIC EXTRAOCULAR MUSCLESIntensive analyses of diaphragm and limb muscles

for the consequences of mutations in the DGC havedemonstrated characteristic patterns of myofiber degen-eration and regeneration, as discussed elsewhere inthis issue. There is now evidence to support cell deathby both necrotic and apoptotic mechanisms, often withspatial and temporal differences in the involvement ofdifferent muscle groups. By contrast, there have beenfew studies of histopathological changes in extraocularmuscles of natural and transgenic models of musculardystrophy. The specialized nature, unique biology, smallsize, and relative inaccessibility of the extraocularmuscles likely contribute toward the lack of interest inthis muscle group in animal models of muscular dystro-phy. However, the recent appreciation that study ofmuscle groups that are spared in muscular dystrophy

Fig. 2. Mouse extraocular muscles have a typical relationship withthe eye and optic nerve (ON). Left: Arrangement of superior rectus(SR), lateral rectus (LR), medial rectus (MR), levator palpebraesuperioris (LPS), and retractor bulbi (RB) is shown in cross-sectionwith the optic nerve in the posterior orbit. Hematoxylin and eosinstain, 5 µm paraffin section; scale bar 5 300 µm. Right: Fiber typecomposition of orbital (orb) and global (glob) muscle layers is shown for

an adult mouse rectus muscle. The orbital layer contains two distinctfiber types: orbital singly (1) and orbital multiply (2) innervatedmuscle fibers. The global layer contains four additional fiber types:global red (3), global intermediate (4), and global white (5) singlyinnervated fibers, and global multiply innervated fibers (6). Toluidineblue stain, 1 µm plastic section; scale bar 5 25 µm.


may provide insights into its pathogenesis and treat-ment has focused attention to these muscles. It is nowevident that the extraocular muscles are spared in atleast three distinct forms of muscular dystrophy, involv-ing cytosolic (dystrophin), intrinsic membrane (sarcogly-can), and extracellular (laminin-2) components of theDGC.

Any analysis of extraocular muscle histopathologymust take into account the unique structural featuresof this muscle group (see above and Spencer and Porter,

1988). Traits that are characteristic of the extraocularmuscles often would be viewed as pathological if pre-sent in other skeletal muscles. Among the musculardystrophies, the extraocular muscles have been beststudied in the models of dystrophin deficiency thatmimic DMD. Several investigators have shown that thestructure of the extraocular muscles is conspicuouslyspared in animal models of dystrophin deficiency, includ-ing the mdx mouse and cmdx dog (Kaminski et al.,1992; Karpati and Carpenter, 1986; Karpati et al.,

Fig. 3. Original force tracings from an extraocular muscle (superior rectus) and a diaphragm musclebundle in response to increasing stimulation frequencies. From the bottom, stimulation frequencies: 10,60, 100, 200, and 300 Hz for superior rectus; 10, 40, 60, 100, and 200 Hz for the diaphragm. Force scale forthe extraocular muscle is 1 Newton/cm2, only a tenth of that for the diaphragm.

Fig. 4. Extraocular muscles lack signs of pathology in musculardystrophy. Light photomicrographs illustrating alterations in gastroc-nemius (gast), main extraocular (EOM), and retractor bulbi (RB)muscles in the gsg -/- mouse, a knockout of the g-sarcoglycan gene. Asin Hack et al. (1998), hindlimb muscles exhibit abundant centralnuclei, fibrosis, and infiltration of inflammatory cells. In contrast, the

extraocular muscles exhibit no signs of pathology. The accessoryextraocular muscles, including the retractor bulbi and levator palpe-brae superioris (not shown), do have centrally-nucleated fibers, but inlower numbers than in gastrocnemius. The presence of central nucleiis a sign of regenerating muscle. Hematoxylin and eosin stain, paraffinsection; scale bar 5 50 µm.


1988; Khurana et al., 1995; Porter et al., 1998; Ragusaet al., 1996). There are no signs of myofiber degenera-tion, connective tissue accumulation, or central nucleicharacteristic of regenerated muscles in the principal(rectus and oblique) extraocular muscles in any of theanimal models. Although studies in the mdx mousesuggest the occurrence of renewed pathology in hind-limb muscles of aged mice (Pastoret and Sebille, 1995),

there are no alterations in the principal extraocularmuscles of old mdx mice (Porter et al., 1998). Interest-ingly, the accessory extraocular muscles, the levatorpalpebrae superioris and retractor bulbi, exhibit mildpathological changes, the most prominent being theaccumulation of central nuclei.

Some forms of CMD result from primary mutationsin the a2 subunit of laminin-2. An early study of

Fig. 5. Extraocular muscles exhibit the same distribution patternof dystrophin and associated glycoproteins as in other skeletal muscles.Immunocytochemical localization shows extraocular sarcolemmal dis-tribution of dystrophin (dys), a-dystroglycan (a-dg), b-dystroglycan(b-dg), a-sarcoglycan (a-sg), b-sarcoglycan (b-sg), g-sarcoglycan, andd-sarcoglycan (not shown) in the normal adult muscle. Utrophin

expression in extraocular muscles is restricted to neuromuscularjunctions as in limb muscles; utrophin (utr) co-localizes with acetylcho-line receptors identified by a-bungarotoxin (a-bt). Adult mouse extra-ocular muscle, except for b-sg micrograph, which shows retractor bulbiand extraocular muscle (small fibers, top left). Scale bars 5 50 and10 µm.


extraocular muscles in the dy/dy mouse model oflaminin-2 deficiency (Pachter et al., 1973) reportedultrastructural changes. This report contrasts withclinical observations that patients with laminin 2-defi-cient CMD have full ocular motility (Mendell et al.,1995). A re-examination of dy/dy extraocular muscles(Porter and Andrade, unpublished data) showed alter-ations identical to that described for the dystrophin-deficient mdx mouse. The extraocular muscles exhibitno signs of sarcolemmal disruption (by either EvansBlue or analysis of calcium homeostasis (Porter andKarathanasis, 1998) and there is no evidence of myofi-ber degeneration or regeneration in the principal extra-ocular muscles. By contrast, central nuclei are readilyapparent in levator palpebrae superioris and retractorbulbi muscles.

Recently, the extraocular muscles have been exam-ined in an animal model of sarcoglycan deficiency, theg-sarcoglycan knockout mouse (Porter et al., unpub-lished data). In the absence of g-sarcoglycan, the gsg -/-mice present with generalized skeletal muscle pathol-ogy and cardiomyopathy by 20 weeks of age (Hack etal., 1998). g-Sarcoglycan deficiency leads to secondaryloss of a-, b-, and d-sarcoglycan but does not disturbsarcolemmal localization of dystrophin, b-dystroglycan,and laminin-2. The pattern of morphological alter-ations in extraocular muscle was identical to thatdetected in both mdx and dy/dy mice, that is principalmuscle sparing and mild changes in accessory extraocu-lar muscles (Fig. 4).

In summary, the extraocular muscles exhibit thesame lack of histopathological changes in response tovarious muscular dystrophies that affect the DGC,regardless of whether the primary mutation involvesdystrophin, laminin-2, or a sarcoglycan. The occurrenceof both fully protected principal extraocular musclesand partially susceptible accessory extraocular musclesprovides an excellent model for understanding themechanisms that spare some muscle groups in muscu-lar dystrophy.

LACK OF OCULAR MOTOR ALTERATIONS INDYSTROPHIC PATIENTS

Many of the molecular defects underlying the variousmuscular dystrophies have been characterized in thelast decade. Previously, dystrophies were defined byclinical presentations based on the characteristic differ-ential involvement of muscle groups. In general, theextraocular muscles are spared by the muscular dystro-phies that result from defects of the DGC.

A distinct differential involvement of muscle groupsis typical of DMD: legs are usually affected more thanarms, proximal muscle groups more so than distalmuscles. Indeed, an even more specific pattern can bedetected. Quadriceps, triceps, wrist extensors, ankledorsiflexors, and neck flexors are more affected thantheir antagonists (Emery, 1987; Engel et al., 1994). Incontrast, striated muscle of the sphincters and bulbarmuscles are largely spared.

Evaluations of eye movements have been occasion-ally performed in dystrophin-deficient patients. Usinginfrared oculography, we measured horizontal eye move-ments in three patients with dystrophin deficiency(Kaminski et al., 1993). These patients were all in their20s, wheelchair-bound, and required assistance with

daily living activities. The amplitude of their saccades(fast, redirecting eye movements) were normal, andsaccadic velocities were slightly slower but withinnormal 95% confidence limits. In this study, sevenadditional patients were evaluated clinically and nonedemonstrated obvious ocular motility abnormalities.Scelsa and coworkers described a single Becker muscu-lar dystrophy patient with mild, bilateral medial andlateral rectus weakness (Scelsa et al., 1996). Horizontaland vertical peak saccadic velocities were significantlyslower. This patient was clearly exceptional, and onemust wonder whether other modifying influences mayexplain the extraocular muscle involvement.

Myotonic dystrophy is the most common adult-onsetmuscular dystrophy and is caused by a triplet repeatmutation in the noncoding region of the ubiquitouslyexpressed protein kinase myotonin. The pathophysiol-ogy of myotonic dystrophy is clearly different fromthose disorders affecting the DGC, and it is presentedfor comparison. Muscle involvement in this disordervaries widely, from nearly asymptomatic to severeweakness at birth and death in early childhood. Thecharacteristic clinical feature, myotonia, is produced byspontaneous repetitive sarcolemmal depolarizations,leading to prolonged contractions. Histological evalua-tion demonstrates dystrophic changes in muscle, butthe pattern of muscle group involvement differs. Thereis sparing of proximal musculature and prominentweakness of muscles innervated by the cranial nerves.In contrast to other dystrophies, ptosis is a characteris-tic feature. Eye movement recordings show slowing ofsaccades and impaired pursuit. These abnormalitiesmay be of central nervous system or muscle origin.Saccadic velocities improve with repeated testing andtreatment with muscle membrane stabilizing drugs,suggesting the extraocular muscles are involved tosome degree but are relatively spared compared withother skeletal muscles and the levator palpebrae (DiConstanzo et al., 1997). Limited pathological studiesfrom the 1970s demonstrate centrally located nuclei insome extraocular muscles but otherwise no unequivocaldystrophic changes (Ginsberg et al., 1978; Kuwabaraand Lessel, 1976).

WHY ARE EXTRAOCULAR MUSCLES SPARED?Skeletal muscles are not all created equal in their

response to neuromuscular disease. While the target-ing or sparing of particular muscles or muscle groups isnot an infrequent occurrence, there often are no overtphenotypic or functional differences between the af-fected and unaffected muscles that might provide cluesto the underlying mechanisms for disease selectivity.For example, the distal myopathies spare seeminglyidentical muscles that are in close proximity to theaffected musculature. Unfortunately, for most neuro-muscular diseases, direct relationships have not beenestablished between muscle or motor neuron group-specific properties and the propensity to disease.

We propose that the sparing of the extraocularmuscles in muscular dystrophy is directly related tomuscle group-specific properties that either prevent oradapt for the cascade of events leading to myofiberdegeneration. Most published data deal with the patho-genesis of dystrophin deficiency in humans and itscorresponding animal model, the mdx mouse. Some


studies on animal models of CMD and LGMD have beenrecently published. The identification of the definitivemechanism(s) for extraocular muscle protection is lim-ited in part because the precise sequence of events thatis triggered by the dystrophic process is not fullyresolved. Nonetheless, the sparing of the extraocularmuscles could be potentially explained by the expres-sion of alternative proteins that could compensate forlost proteins, or by the functional or metabolic charac-teristics of these muscles, which may protect them fromthe consequences of the primary defects. Data on whichfactor, if any, is more important are still lacking,although a clearer understanding of the response of theextraocular muscles to the dystrophic process is slowlybeginning to emerge.

Preservation of Sarcolemmal StabilityIn DMD patients and mdx mice, lack of dystrophin

leads to secondary loss of the other elements of the DGCin limb skeletal muscle: dystroglycans, syntrophins,and sarcoglycans (Straub and Campbell, 1997). Inhuman and animal models of LGMD, a mutation of asingle sarcoglycan results in the loss of some or all thesarcoglycans, with evidence of sarcolemmal disruption(Duclos et al., 1998; Duggan and Hoffman, 1996; Hacket al., 1998; Sewry et al., 1996). While pure laminin-2deficiency does not usually affect the distribution ofother membranes of the complex, it disrupts the sarco-lemma as evidenced by cytosolic enzyme release andvital dye uptake in particular muscles (Pegoraro et al.,1998; Straub et al., 1997; Voit, 1998).

One explanation for the extraocular muscle sparingby these dystrophic disorders is that the loss of a singlemember of the DGC might not be sufficient to destabi-lize the sarcolemma of extraocular muscles. This mightoccur if the composition of the DGC differed, themechanical stresses seen by this muscle group wereless (see Fig. 3; Karpati and Carpenter, 1986; Karpati etal., 1988), or other proteins replaced the missing mem-bers (Hodges et al., 1997; Law et al., 1994; Matsumuraet al., 1992; Settles et al., 1996). Our preliminary dataargue against significant differences in the organiza-tion of the DGC in extraocular muscle sarcolemma.Dystrophin, laminin-2, dystroglycans, and sarcogly-cans have the same general distribution in extraocularas in limb and respiratory muscles.

The clinical hallmark of the dystrophic process ismuscular weakness and wasting. Damage and necrosisof skeletal muscle fibers explain the loss of contractilefunction. The fast twitch fibers of limb skeletal musclesare particularly susceptible to sarcolemmal destabiliza-tion in DMD patients and mdx mice (Karpati andCarpenter, 1986; Webster et al., 1988). Because of theirlarger size and faster contractile properties, thesefibers are subjected to considerable stresses duringrapid contractions, straining and disrupting the weak-ened sarcolemma (Karpati and Carpenter, 1986; Karpatiet al., 1988; Petrof et al., 1993; Weller et al., 1990). Thiswould predict that the intrinsic susceptibility to me-chanical injury is what separates spared from affectedmuscle groups. It is peculiar that two of the factorsfrequently mentioned as detrimental to dystrophic limband respiratory skeletal muscles (activity and eccentriccontractions) are found in the extraocular muscles, aspared muscle group. The combination of small fiber

size, fast speed of contraction, and high activity levels isa distinctive characteristic of these muscles (Asmussenet al., 1994; Frueh et al., 1994; Porter et al., 1995).While the extraocular muscles are very fast and active,the loads they work against are comparatively small.This explains their in vitro properties: fast twitchkinetics and velocity of shortening, but low absoluteand specific forces (Fig. 3) (Asmussen et al., 1994; Closeand Luff, 1974; Frueh et al., 1994). It could also be whythe extraocular muscles are spared by the dystrophicprocess even if sarcolemmal structure is compromised.In other words, even if no alternative ‘‘scaffolding’’proteins are expressed, the functional characteristics ofthe extraocular muscles may be such that the musclesare not susceptible to mechanical injury.

In molecular models of the protein complex linkingcytoskeleton to extracellular matrix, the key domains ofdystrophin are the F-actin and b-dystroglycan bindingsites (Blake et al., 1996; Campbell, 1995). The autoso-mal product utrophin has identical binding sites, butthis protein is normally restricted to the neuromuscu-lar and myotendinous junctions (Guo et al., 1996; Loveet al., 1989; Matsumura et al., 1992; Tinsley et al.,1992; Winder et al., 1995). Nevertheless, utrophin is apotential substitute for dystrophin (Khurana et al.,1990; Love et al., 1989; Tinsley and Davies, 1993). Adystrophy-associated increase in utrophin is not alto-gether unusual, at least in small caliber, presumablyregenerative, fibers that recapitulate myogenic events.Previous studies of limb and diaphragm muscles haveshown increased utrophin levels and apparent expan-sion of cellular localization of utrophin to sites formerlyoccupied by dystrophin in both DMD patients andanimal dystrophin deficiency models (Clerk et al., 1993;Helliwell et al., 1992; Karpati et al., 1993; Khurana etal., 1991; Law et al., 1994; Morris, 1998; Pons et al.,1994; Takemitsu et al., 1991; Tanaka et al., 1991;Taylor et al., 1997). However, the level of utrophinupregulation might not be sufficient to fully compen-sate for the absence of dystrophin, even in the mdxmouse, which shows mild signs of limb dystrophy and alimited life span (Lefaucheur et al., 1995; Matsumuraand Campbell, 1994). Interventions that increase utro-phin to a level sufficient to saturate b-dystroglycanbinding sites and preserve the DGC can structurallyand functionally rescue mdx limb and diaphragm mus-culature (Deconinck et al., 1997; Tinsley et al., 1996).This is a vitally important finding, as it opens the doorfor a feasible treatment for DMD.

The extraocular muscles appear to have the capacityto endogenously increase utrophin expression in re-sponse to dystrophin deficiency, and this ability may beresponsible for its complete sparing in DMD. Thelocalization of utrophin to both neuromuscular junc-tions and non-junctional sarcolemma in mdx extraocu-lar muscles (Matsumura et al., 1992) lends support tothe preservation of sarcolemmal integrity hypothesis.We have shown the upregulation of utrophin proteinlevels in extraocular muscles of mdx mice. More impor-tantly, our data show that mice deficient in bothdystrophin and utrophin exhibit substantial extraocu-lar muscle pathology (Porter et al., 1998), therebysupporting a putative sparing role by utrophin. Further-more, as discussed earlier, the extraocular muscles arealso spared in dy/dy and g-sarcoglycan knockout mice


(Andrade et al., 1998) (Fig. 4). Curiously, in all threedystrophy models, the accessory extraocular muscles(levator palpebrae superioris and retractor bulbi) arefound to have increased the number of central nuclei, arather mild pathological change in response to themutations. This opens the door to interesting functionaland structural comparisons between muscle groups inclose proximity to one another.

Enhanced Calcium HomeostasisThe loss of calcium homeostasis is generally consid-

ered to be an early step in the pathogenesis of musculardystrophy. The disruption of the cytoskeleton-to-extracellular matrix linkage, via a genetic defect in anyone component of the DGC, is thought to destabilize thesarcolemma, increase calcium influx, and trigger celldeath that is mediated by calcium-activated proteases(Campbell, 1995). As the disease progresses, calciumhomeostasis is further altered by proteolysis of a keycalcium sequestration enzyme, the sarcoplasmic reticu-lum (SR) calcium-ATPase (Leberer et al., 1988). Al-though regenerated mdx limb muscles do not undergorepeated necrotic cycles, intracellular calcium contentremains elevated (Turner et al., 1988, 1991). However,if utrophin is overexpressed beyond a certain thresholdin gene insertion experiments in mdx mice, calciumhomeostasis is restored (Deconinck et al., 1997). Thus,when the sarcolemmal defect is only partially resolved,as in the native mdx strain, improved cellular calciumhandling mechanisms may remain long-term contribu-tors to calcium homeostasis and myofiber viability indystrophin deficiency. For example, it has been shownthat fast skeletal muscle fibers of mdx mice upregulatethe production of parvalbumin, a cytosolic calcium-binding protein (Gailly et al., 1993). The muscle fibertype most affected by the dystrophin deficiency (Web-ster et al., 1988) increases its calcium buffering capac-ity, presumably in response to a higher calcium load.

The differential response of muscle groups in modelsof muscular dystrophy may represent the ability ofsome muscles to manage the pathological increase inintracellular free calcium concentration. That is, extra-ocular muscles might be spared in these pathologies,even if sarcolemmal stability is compromised, by meansof a constitutively high calcium sequestration capacity.Consistent with this hypothesis, the functional proper-ties of the extraocular muscles normally require asubstantial calcium handling capacity (Porter andKarathanasis, 1998, 1999; Porter et al., 1997; Porter,1998). These are exceptionally fast contracting andactive muscles, also capable of sustained tetanus forextended periods (Asmussen et al., 1994; Close andLuff, 1974; Frueh et al., 1994; Katoh et al., 1998). Thesmall myofibers, unique troponin T isoform composi-tion, abundant mitochondria and SR, and high SRcalcium and calcium-ATPase content of extraocularmuscles (Briggs et al., 1988; Porter and Karathanasis,1998; Spencer and Porter, 1988) are consistent withthese functional traits. Extraocular muscles may modu-late cytosolic free calcium levels by both passive andactive mechanisms, as most of their fibers are high inparvalbumin (Celio and Heizmann, 1982), and mainlyexpress the fast isoform of calcium-ATPase (Jacoby andKo, 1993; Tullis and Block, 1996). It is likely that the

pCa/force properties of extraocular muscles mean thatthey increment force in a more graded manner and overa wider range of cytosolic calcium concentrations (Briggset al., 1988; Schachat et al., 1987). Thus, the ability tomodulate free cytosolic calcium both more finely andabove the levels typically seen in skeletal muscleswould appear to be required for day-to-day function ofextraocular muscle. For this reason, it is plausible toconsider enhanced calcium homeostasis as a potentialprotective mechanism of extraocular muscles in muscu-lar dystrophy.

There is evidence that a substantial calcium seques-tration capacity protects the extraocular muscles fromtoxic agents that act via increases in intracellularcalcium concentration. The aminoacyl local anestheticsincrease intracellular calcium via frank sarcolemmalbreaks and are myotoxic to the extent that they havebeen used to induce necrosis prior to the study ofmuscle regeneration. By contrast, we have shown thatthe local anesthetics elicit only mild and limited dam-age when applied to mammalian extraocular muscles(Porter et al., 1988). Likewise, calcium ionophoreA-23187 (calcimycin) fails to kill extraocular musclefibers at concentrations that are lethal to virtuallyevery other cell type (Khurana et al., 1995). These datashow that the extraocular muscles exhibit substantialcapacity to regulate intracellular free calcium concentra-tion. However, they are not direct evidence that calciumscavenging capacity plays a causal role in the protec-tion of this muscle group in the muscular dystrophies.

Free Radical ScavengingMorphological similarities between the muscular dys-

tropies and the myopathies produced by vitamin E orselenium deficiency have suggested a common role forreactive oxygen species (‘‘free radicals’’) in the pathogen-esis of these disorders. This suspicion was strengthenedby evidence of increased oxidative stress in the musclesof patients and animal models of muscular dystrophy(Hunter and Mohamed, 1986; Jackson et al., 1989; Karand Pearson, 1979; Mechler et al., 1984). Curiously,most studies demonstrated increased lipid peroxida-tion, without clear effects on cellular antioxidant sys-tems. Furthermore, trials with selected antioxidantshave not improved the natural history of the dystrophicprocess (Bertorini et al., 1985; Edwards et al., 1984;Gamstorp et al., 1986; Jackson et al., 1989; Stern et al.,1982).

Despite the failure of the antioxidant therapies, thehypothesis that reactive oxygen species cause at leastsome of the pathological changes in the dystrophiesremains attractive. One consequence of the loss ofcalcium homeostasis is the increased production of freeradicals as a result of mitochondrial uncoupling, oractivation of cytosolic free radical generators (Hal-estrap et al., 1993; McCutchan et al., 1990; van deWater et al., 1994; Wrogemann and Pena, 1976). Al-tered redox homeostasis may then aggravate the loss ofcalcium handling capacity and recruit other pathwaysof muscle damage such as proteases (Jackson et al.,1984; Rodemann et al., 1982). Therefore, increasedproduction of reactive oxygen species in the musculardystrophies could be considered as a process of damageamplification, leading to the widespread necrosis char-


acteristic of the full-blown pathology. The extraocularmuscles appear to have an enhanced antioxidant capac-ity, which could fulfill a protective role under the settingof increased oxidative stress such as muscular dystro-phy (Ragusa et al., 1996). However, recent studiessuggest that free radical scavenging capacity, thoughsubstantial in the extraocular muscles, is not primarilyresponsible for their sparing in the mouse model ofdystrophin-deficiency (Ragusa et al., 1996, 1997;Wehling et al., 1998).

Unfortunately, most studies of the involvement ofreactive oxygen species in the pathological cascade ofevents in muscular dystrophy have been conducted latein the course of the disease, when there is considerableevidence of massive tissue damage. At this stage, theelevated indices of lipid peroxidation and altered levelsof some antioxidants may simply reflect the local activ-ity of inflammatory response cells that are recruited todamaged areas. In other words, the presence of reactiveoxygen species and associated oxidative damage areincidental and not primary steps of the dystrophicprocess; therefore, antioxidant therapies would im-prove indices of oxidative injury, but not the intrinsicpathological changes due to the muscular dystrophy.Recently, it has been suggested that reactive oxygenspecies play a role at a much earlier disease stage,before the appearance of frank muscle necrosis. Evi-dence of oxidative damage has been detected prior tothe full-blown muscle necrosis that characterizes mus-cular dystrophies; in this scenario, reactive oxygenspecies may participate as more or less specific signalsthat are integral to the sequence of events linking theprimary protein defect to overt muscle damage (Disat-nik et al., 1998). Such a role for free radicals remains tobe explored in the extraocular muscles of dystrophicpatients and animal models.

FUTURE DIRECTIONSThe devastating consequences of the various muscu-

lar dystrophies are even more apparent when a muscleor muscle group is spared. In the analysis of diseasemechanisms, study of the exceptional cell or tissueresponses may prove to be of considerable value inunderstanding the norm. The sparing of the extraocu-lar muscles has been noted as a typical clinical findingin DMD, LGMD, and CMD, pathologies due to a brokenlinkage between the cytoskeleton and the extracellularmatrix. Uncovering the cellular mechanisms that ex-plain the ‘‘natural’’ protection of the extraocular musclesin these diseases should contribute to knowledge ofboth pathogenesis and treatment. Three research direc-tions are particularly relevant: (1) characterize theDGC in the extraocular muscles, and determine whetheralternative ‘‘scaffolding’’ proteins are expressed; (2)evaluate whether the working conditions of the extra-ocular muscles do not induce mechanical damage asproposed for other skeletal muscles; (3) determine if theextraocular muscles adapt to or compensate for distur-bances in ionic and metabolic homeostasis that accom-pany the primary protein defect. Defining whether anyof these options explains the sparing of the extraocularmuscles by the muscular dystrophies should be animportant goal for the near future.

ACKNOWLEDGMENTSWe thank Beth Ann Benetz, M.A., C.R.A. Director,

Ophthalmic Photography for her skillful assistancewith the figures in this paper.

REFERENCESAndrade FH, Krebs CR, Bonner PH, Porter JD. 1998. Extraocular

muscle is spared in merosin-deficient congenital muscular dystro-phy. Muscle Nerve Suppl. 7:S106 (Abstr).

Asmussen G, Gaunitz U. 1981. Mechanical properties of the isolatedinferior oblique muscle of the rabbit. Pflugers Arch 392:183–190.

Asmussen G, Beckers-Bleukx G, Marechal G. 1994. The force-velocityrelation of the rabbit inferior oblique muscle; influence of tempera-ture. Pflugers Arch 426:542–547.

Bahn RS, Heufelder AE. 1993. Pathogenesis of Graves’ ophthalmopa-thy. N Engl J Med 91:1411–1419.

Bertorini TE, Palmieri GM, Griffin J, Chesney C, Pifer D, Verling L,Airozo D, Fox IH. 1985. Chronic allopurinol and adenine therapy inDuchenne muscular dystrophy: effects on muscle function, nucleo-tide degradation, and muscle ATP and ADP content. Neurology35:61–65.

Blake DJ, Tinsley JM, Davies KE. 1996. Utrophin: a structural andfunctional comparison to dystrophin. Brain Pathol 6:37–47.

Briggs MH, Jacoby J, Davidowitz J, Schachat FH. 1988. Expression ofa novel combination of fast and slow troponin T isoforms in rabbitextraocular muscles. J Muscle Res Cell Motil 9:241–247.

Brueckner JK, Porter JD. 1998. Visual system maldevelopmentdisrupts extraocular muscle-specific myosin expresion. JAppl Physiol85:584–592.

Brueckner JK, Itkis O, Porter JD. 1996. Spatial and temporal patternsof myosin heavy chain expression in developing rat extraocularmuscle. J Muscle Res Cell Motil 17:297–312.

Brueckner JK, Ashby LP, Prichard JR, Porter JD. 1999. Vestibulo-ocular pathways modulate extraocular muscle myosin expressionpatterns. Cell Tissue Res 295:477–484.

Burden SJ. 1998. The formation of neuromuscular synapses. GenesDev 12:133–148.

Campbell KP. 1995. Three muscular dystrophies: loss of cytoskeleton-extracellular matrix linkage. Cell 80:675–679.

Celio MR, Heizmann CW. 1982. Calcium-binding protein parvalbuminis associated with fast contracting muscle fibres. Nature 297:504–506.

Clerk A, Morris GE, Dubowitz V, Davies KE, Sewry CA. 1993.Dystrophin-related protein, utrophin, in normal and dystrophichuman fetal skeletal muscle. Histochem J 25:554–561.

Close RI, Luff AR. 1974. Dynamic properties of inferior rectus muscleof the rat. J Physiol (London) 236:259–270.

Collins CC, O’Meara D, Scott AB. 1975. Muscle tension duringunrestrained human eye movements. J Physiol (London) 245:351–369.

Couly GF, Coltey PM, Le Douarin NM. 1992. The developmental fateof the cephalic mesoderm in quail-chick chimeras. Development114:1–15.

Crosbie RH, Lebakken CS, Holt KH, Venzke DP, Straub V, Lee JC,Grady RM, Chamberlain JS, Sanes JR, Campbell KP. 1999. Mem-brane targeting and stabilization of sarcospan is mediated by thesarcoglycan subcomplex. J Cell Biol 145:153–165.

Deconinck N, Tinsley J, De Backer F, Fisher R, Kahn D, Phelps S,Kavies K, Gillis J-M. 1997. Expression of truncated eutrophin leadsto major functional improvements in dystrophin-deficient muscles ofmice. Nature Med 3:1216–1221.

Di Constanzo A, Toreillo A, Mottola A, Di Iorio G, Bonavita V, TedeschiG. 1997. Relative sparing of extraocular muscles in myotonicdystrophy. Acta Neurol Scand 95:158–163.

Disatnik M-H, Dhawan J, Yu Y, Beal MF, Whirl MM, Franco AA,Rando TA. 1998. Evidence of oxidative stress in mdx mouse muscle:studies of the pre-necrotic state. J Neurol Sci 161:77–84.

Duclos F, Straub V, Moore SA, Venzke DP, Hrstka RF, Crosbie RH,Durbeej M, Lebakken CS, Ettinger AJ, van der Meulen J, Holt KH,Lim LE, Sanes JR, Davidson BL, Faulkner JA, Williamson R,Campbell KP. 1998. Progressive muscular dystrophy in a-sarcogly-can-deficient mice. J Cell Biol 142:1461–1471.

Duggan DJ, Hoffman EP. 1996. Autosomal recessive muscular dystro-phy and mutations of the sarcoglycan complex. Neuromusc Disord6:475–482.

Edwards RH, Jones DA, Jackson MJ. 1984. An approach to treatment


in muscular dystrophy with particular reference to agents influenc-ing free radical damage. Med Biol 62:143–147.

Emery AEH. 1987. Duchenne muscular dystrophy. Oxford: OxfordUniversity Press.

Emery AEH. 1994. Some unanswered questions in Duchenne muscu-lar dystrophy. Neuromusc. Disord 4:301–303.

Engel AG, Yamamoto M, Fischbeck K. 1994. Muscular dystrophies. In:Engel A, Franzini-Armstrong C, editors. Myology. New York: Mc-Graw-Hill, Inc. p 1133–1187.

Frueh BR, Hayes A, Lynch GS, Williams DA. 1994. Contractileproperties and temperature sensitivity of the extraocular muscles,the levator and superior rectus, of the rabbit. J Physiol (London)475:327–336.

Fuchs AF, Binder MD. 1983. Fatigue resistance of human extraocularmuscles. J Neurophysiol 49:28–34.

Gailly P, Hermans E, Octave JN, Gillis JM. 1993. Specific increase ofgenetic expression of parvalbumin in fast skeletal muscles of mdxmice. FEBS Lett 326:272–274.

Gamstorp I, Gustavson KH, Hellstrom O, Nordgren B. 1986. A trial ofselenium and vitamin E in boys with muscular dystrophy. J ChildNeurol 1:211–214.

Ginsberg J, Hamblet J, Menefee M. 1978. Ocular abnormality inmyotonic dystrophy. Ann Ophthalmol 10:1021–1028.

Guo W-XA, Nichol M, Merlie JP. 1996. Cloning and expression of fulllength mouse utrophin: the differential association of utrophin anddystrophin with AChR clusters. FEBS Lett398:259–264.

Hack AA, Ly CT, Jiang F, Clendenin CJ, Sigrist KS, Wollmann RL,McNally EM. 1998. g-Sarcoglycan deficiency leads to muscle mem-brane defects and apoptosis independent of dystrophin. J Cell Biol142:1279–1287.

Halestrap AP, Griffiths EJ, Connern CP. 1993. Mitochondrial calciumhandling and oxidative stress. Biochem Soc Trans 21:353–358.

Helliwell TR, Man NT, Morris GE, Davies KE. 1992. The dystrophin-related protein, utrophin, is expressed on the sarcolemma of regen-erating human skeletal muscle fibres in dystrophies and inflamma-tory myopathies. Neuromusc Disord 2:177–184.

Hennig R, Lømo T. 1985. Firing patterns of motor units in normal rats.Nature, 314: 164–166.

Hodges BL, Hayashi YK, Nonaka I, Wang W, Arahata K, Kaufman SJ.1997. Altered expression of the a7b1 integrin in human and murinemuscular dystrophies. J Cell Sci 110:2873–2881.

Hunter MI, Mohamed JB. 1986. Plasma antioxidants and lipidperoxidation products in Duchenne muscular dystrophy. Clin ChimActa 155:123–131.

Jackson MJ, Jones DA, Edwards RHT. 1984. Experimental skeletalmuscle damage: the nature of the calcium-activated degenerativeprocesses. Eur J Clin Invest 14:369–374.

Jackson MJ, Coakley J, Stokes M, Edwards RH, Oster O. 1989.Selenium metabolism and supplementation in patients with muscu-lar dystrophy. Neurology 39:655–659.

Jacoby J, Ko K. 1993. Sarcoplasmic reticulum fast Ca21-pump andmyosin heavy chain expression in extraocular muscles. InvestOphthalmol Vis Sci 34:2848–2858.

Kaminski H, Ruff R. 1997. Ocular muscle involvement by myastheniagravis. Ann Neurol 41:419–420.

Kaminski HJ, Al-Hakim M, Leigh RJ, Katirji MB, Ruff RL. 1992.Extraocular muscles are spared in advanced Duchenne dystrophy.Ann Neurol 32:586–588.

Kaminski HJ, Fenstermaker RA, Abdul-Karim FW, Clayman J, RuffRL. 1993. Acetylcholine receptor subunit gene expression in thymictissue of myasthenic and non-myasthenics. Muscle Nerve 16:1332–1337.

Kar NC, Pearson CM. 1979. Catalase, superoxide dismutase, glutathi-one reductase and thiobarbituric acid-reactive products in normaland dystrophic human muscle. Clin Chim Acta 94:277–280.

Karpati G, Carpenter S. 1986. Small-caliber skeletal muscle fibers donot suffer deleterious consequences of dystrophic gene expression.Am J Med Genet 25:653–658.

Karpati G, Carpenter S, Prescott S. 1988. Small-caliber skeletalmuscle fibers do not suffer necrosis in mdx mouse dystrophy. MuscleNerve 11:795–803.

Karpati G, Carpenter S, Morris GE, Davies KE, Guerin C, Holland P.1993. Localization and quantitation of the chromosome 6-encodeddystrophin-related protein in normal and pathological humanmuscle. J Neuropathol Exp Neurol 52:119–128.

Katoh A, Kitazawa H, Itohara S, Nagao S. 1998. Dynamic characteris-tics and adaptability of mouse vestibulo-ocular and optokineticresponse eye movements and the role of the flocculo-olivary systemrevealed by chemical lesions. Proc Natl Acad Sci USA 95:7705–7710.

Khurana TS, Hoffman EP, Kunkel LM. 1990. Identification of a

chromosome 6-encoded dystrophin-related protein. J Biol Chem265:16717–16720.

Khurana TS, Watkins SC, Chafey P, Chelly J, Tome FMS, Fardeau M,Kaplan J-C, Kunkel LM. 1991. Immunolocalization and developmen-tal expression of dystrophin related proteins in skeletal muscle.Neuromusc Disord 1:185–194.

Khurana TS, Prendergast RA, Alameddine HS, Tome FMS, FardeauM, Arahata K, Sugita H, Kunkel LM. 1995. Absence of extraocularmuscle pathology in Duchenne’s muscular dystrophy: role for cal-cium homeostasis in extraocular muscle sparing. J Exp Med 182:467–475.

King WM, Fuchs AF, Magnin M. 1981. Vertical eye movement-relatedresponses of neurons in midbrain near interstitial nucleus of Cajal.J Neurophysiol 46:549–562.

Kuwabara T, Lessel S. 1976. Electron microscopic study of extraocularmuscles in myotonic dystrophy. Am J Ophthalmol 82:303–309.

Law DJ, Allen DL, Tidball JG. 1994. Talin, vinculin and DRP(utrophin) concentrations are increased at mdx myotendinous junc-tions following onset of necrosis. J Cell Sci 107:1477–1483.

Leberer E, Hartner K-T, Pette D. 1988. Postnatal development of Ca21

sequestration by the sarcoplasmic reticulum of fast and slowmuscles in normal and dystrophic mice. Eur J Biochem 174:247–253.

Lefaucheur JP, Pastoret C, SebilleA. 1995. Phenotype of dystrophinopa-thy in old mdx mice. Anat Rec 242:70–76.

Love DR, Hill DF, Dickson G, Spurr NK, Byth BC, Marsden RF, WalshFS, Edwards YH, Davies KE. 1989. An autosomal transcript inskeletal muscle with homology to dystrophin. Nature 339:55–58.

Luff AR. 1981. Dynamic properties of the inferior rectus, extensordigitorum longus, diaphragm and soleus muscles of the mouse. JPhysiol (London) 313:161–171.

Lynch GS, Frueh BR, Williams DA. 1994. Contractile properties ofsingle skinned fibres from the extraocular muscles, the levator andsuperior rectus, of the rabbit. J Physiol (London) 475:337–346.

Matsumura K, Campbell KP. 1994. Dystrophin-glycoprotein complex:its role in the molecular pathogenesis of muscular dystrophies.Muscle Nerve 17:2–15.

Matsumura K, Ervasti JM, Ohlendieck K, Kahl SD, Campbell KP.1992. Association of dystrophin-related protein with dystrophinassociated proteins in mdx mouse muscle. Nature 360:588–591.

McArdle A, Helliwell TR, Beckett GJ, Catapano M, Davis A, JacksonMJ. 1998. Effect of propylthiouracil-induced hypothyroidism on theonset of skeletal muscle necrosis in dystrophin-deficient mdx mice.Clin Sci 95:83–89.

McCutchan HJ, Schwappach JR, Enquist EG, Walden DL, Terada LS,Reiss OK, Leff JA, Repine JE. 1990. Xanthine oxidase-derived H2O2contributes to reperfusion injury of ischemic skeletal muscle. Am JPhysiol 258:H1415-H1419.

Mechler F, Imre S, Dioszeghy P. 1984. Lipid peroxidation and superox-ide dismutase activity in muscle and erythrocytes in Duchennemuscular dystrophy. J Neurol Sci 63:279–283.

Mendell JR, Sahenk Z, Prior TW. 1995. The childhood musculardystrophies: diseases sharing a common pathogenesis of membraneinstability. J Child Neurol 10:150–159.

Mitsumoto H, Chad DA, Pioro EP. 1998. Amyotrophic lateral sclerosis.Philadelphia: F.A. Davis.

Morris GE. 1998. Dystrophin is replaced by utrophin in frog heart:implications for muscular dystrophy. Neuromusc Disord 7:493–498.

Pachter BR, Davidowitz J, Breinin GM. 1973. A light and electronmicroscopic study in serial sections of dystrophic extraocular musclefibers. Invest Ophthalmol 12:917–923.

Pastoret C, Sebille A. 1995. mdx Mice show progressive weakness andmuscle deterioration with age. J Neurol Sci 129:97–105.

Pegoraro E, Marks H, Garcia CA, Crawford T, Mancias P, ConnollyAM, Fanin M, Martinello F, Trevisan CP, Angelini C, Stella A,Scavina M, Munk RL, Servidei S, Bonnemann CC, Bertorini T,Acsadi G, Thompson CE, Gagnon D, Hoganson G, Carver V,Zimmerman RA, Hoffman EP. 1998. Laminin a2 muscular dystro-phy. Genotype/phenotype studies of 22 patients. Neurology 51:101–110.

Petrof BJ, Shrager JB, Stedman HH, Kelly AM, Sweeney HL. 1993.Dystrophin protects the sarcolemma from stresses developed duringmuscle contraction. Proc Natl Acad Sci USA 90:3710–3714.

Pons F, Robert A, Marini JF, Leger JJ. 1994. Does utrophin expressionin muscles of mdx mice during postnatal development functionallycompensate for dystrophin deficiency. J Neurol Sci 122:162–170.

Porter, J.D. 1998. Extraocular muscle sparing in muscular dystrophy:a critical evaluation of potential protective mechanisms. Neuro-musc. Disord., 8: 198–203.

Porter JD, Baker RS. 1992. Prenatal morphogenesis of primate


extraocular muscle: neuromuscular junction formation and fibertype differentiation. Invest. Ophthalmol. Vis. Sci., 33: 657–670.

Porter JD, Baker RS. 1996. Muscles of a different ‘‘color’’: the unusualproperties or the extraocular muscles may predispose or protectthem in neurogenic and myogenic disease. Neurology 46: 30–37.

Porter JD, Karathanasis P. 1998. Extraocular muscle in merosin-deficient muscular dystrophy: cation homeostasis is maintained butis not mechanistic in muscle sparing. Cell Tissue Res 292:495–501.

Porter JD, Karathanasis P. 1999. The development of extraocularmuscle calcium homeostasis parallels visuomotor system matura-tion. Biochem Biophys Res Commun 257:678–683.

Porter JD, Edney DP, McMahon EJ, Burns LA. 1988. Extraocularmyotoxicity of the retrobulbar anesthetic bupivacaine hydrochlo-ride. Invest Ophthalmol Vis Sci 29:163–174.

Porter JD, Baker RS, Ragusa RJ, Brueckner JK. 1995. Extraocularmuscles: basic and clinical aspects of structure and function. SurvOphthalmol 39:451–484.

Porter JD, Karathanasis P, Bonner PH, Brueckner JK. 1997. Theoculomotor periphery: the clinician’s focus is no longer a basicscience stepchild. Curr Opin Neurobiol 7:880–887.

Porter JD, Rafael JA, Ragusa RJ, Brueckner JK, Trickett JI, DaviesKE. 1998. The sparing of extraocular muscle in dystrophinopathy islost in mice lacking utrophin and dystrophin. J Cell Sci 111:1801–1811.

Ragusa RJ, Chow CK, St. Clair DK, Porter JD. 1996. Extraocular, limband diaphragm muscle group-specific antioxidant enzyme activitypatterns in control and mdx mice. J Neurol Sci.139:180–186.

Ragusa RJ, Chow CK, Porter JD. 1997. Oxidative stress as a potentialpathogenic mechanism in an animal model of Duchenne musculardystrophy. Neuromusc Disord 7:379–386.

Robinson DA. 1970. Oculomotor unit behavior in the monkey. JNeurophysiol 33:393–404.

Robinson DA. 1981. Control of eye movements. In: Brooks VB, editor.Handbook of physiology, Section 1: the nervous system. Bethesda,MD: American Physiological Society. p 1275–1320.

Rodemann HP, Waxman L, Goldberg AL. 1982. The stimulation ofprotein degradation in muscle by Ca11 is mediated by prostaglandinE

2and does not require the calcium-activated protease. J Biol

Chem.257:8716–8723.Rome LC. 1998. Some advances in integrative muscle physiology.

Comp Biochem Physiol B120:51–72.Sartore S, Mascarello F, Rowlerson A, Gorza L, Ausoni S, Vianello M,

Schiaffino S. 1987. Fibre types in extraocular muscles: a new myosinisoform in the fast fibres. J Muscle Res Cell Motil 8:161–172.

Scelsa S, Simpson D, Reichler B, Dai M. 1996. Extraocular muscleinvolvement in Becker muscular dystrophy. Neurology 46:564–566.

Schachat FH, Diamond MS, Brandt PW. 1987. Effect of differenttroponin T-tropomyosin combinations on thin filament activation. JMol Biol 198:551–554.

Settles DL, Cihak RA, Erickson HP. 1996. Tenascin-C expression indystrophin-related muscular dystrophy. Muscle Nerve 19:147–154.

Sewry CA, Taylor J, Anderson LVB, Ozawa E, Pogue R, Piccolo G,Bushby K, Dubowitz V, Muntoni F. 1996. Abnormalities in a-, b- andg-sarcoglycan in patients with limb-girdle muscular dystrophy.Neuromusc Disord 6:467–474.

Sohal GS, Ali AA, Ali MM. 1998. Ventral neural tube cells differentiateinto craniofacial skeletal muscles. Biochem Biophys Res Commun252:675–678.

Spencer RF, Porter JD. 1988. Structural organization of the extraocu-lar muscles. In: Buttner-Ennever JA, editor. Neuroanatomy of theoculomotor system. New York: Elsevier Science Publishers. p 33–79.

Stern LZ, Ringel SP, Ziter FA, Menander-Huber KB, Ionasescu V,

Pellegrino RJ, Snyder RD. 1982. Drug trial of superoxide dismutasein Duchenne’s muscular dystrophy. Arch Neurol 39:342–346.

Straub V, Campbell KP. 1997. Muscular dystrophies and the dystro-phin-glycoprotein complex. Curr Opin Neurol 10:168–175.

Straub V, Rafael JA, Chamberlain JS, Campbell KP. 1997. Animalmodels for muscular dystrophy show different patterns of sarcolem-mal disruption. J Cell Biol 139:375–385.

Takemitsu M, Ishiura S, Koga R, Kamakura K, Arahata K, Nonaka I,Sugita H. 1991. Dystrophin-related protein in the fetal and dener-vated skeletal muscles of normal and mdx mice. Biochem BiophysRes Commun 180:1179–1186.

Tanaka J, Ishiguro I, Efuchi C, Saito K, Ozawa E. 1991. Expression ofa dystrophin-related protein associated with the skeletal muscle cellmembrane. Histochemistry 96:1–5.

Taylor J, Muntoni F, Dubowitz V, Sewry CA. 1997. The abnormalexpression of utrophin in Duchenne and Becker muscular dystrophyis age related. Neuropathol Appl Neurobiol 23:399–405.

Tinsley JM, Davies KE. 1993. Utrophin: a potential replacement fordystrophin? Neuromusc Disord 3:537–539.

Tinsley JM, Blake DJ, Roche A, Fairbrother U, Riss J, Byth BC,Knight AE, Kendrick-Jones J, Suthers GK, Love DR, Edwards YH,Davies KE. 1992. Primary structure of dystrophin-related protein.Nature 360: 591–593.

Tinsley JM, Potter AC, Phelps SR, Fisher R, Trickett JI, Davies KE.1996. Amelioration of the dystrophic phenotype of mdx mice using atruncated utrophin transgene. Nature 384:349–353.

Tullis A, Block BA. 1996. Expression of sarcoplasmic reticulumCa21-ATPase isoforms in marlin and swordfish muscle and heatercells. Am J Physiol 271:R262-R275.

Turner PR, Westwood T, Regen CM, Steinhardt RA. 1988. Increasedprotein degradation results from elevated free calcium levels foundin muscle from mdx mice. Nature 335:735–738.

Turner PR, Fong P, Denetclaw WF, Steinhardt RA. 1991. Increasedcalcium influx in dystrophic muscle. J Cell Biol 115:1701–1712.

van de Water B, Zoeteweij JP, de Bont HJGM, Mulder GJ, NagelkerkeJF. 1994. Role of mitochondrial Ca21 in the oxidative stress-induceddissipation of the mitochondrial membrane potential. J Biol Chem269:14546–14552.

Voit T. 1998. Congenital muscular dystrophies: 1997 update. BrainDev 20:65–74.

Wahl CM, Noden DM, Baker R. 1994. Developmental relationsbetween sixth nerve motor neurons and their targets in the chickembryos. Dev Dyn 201:191–202.

Webster C, Silberstein L, Hays AP, Blau HM. 1988. Fast muscle fibersare preferentially affected in Duchenne muscular dystrophy. Cell 52:503–513.

Wehling M, Stull JT, McCabe TJ, Tidball JG. 1998. Sparing of mdxextraocular muscles from dystrophic pathology is not attributable tonormalized concentration or distribution of neuronal nitric oxidesynthase. Neuromusc Disord 8:22–29.

Weller B, Karpati G, Carpenter S. 1990. Dystrophin-deficient mdxmuscle fibers are preferentially vulnerable to necrosis induced byexperimental lengthening contractions. J Neurol Sci 100:9–13.

Wieczorek DF, Periasamy M, Butler-Browne GS, Whalen RG, Nadal-Ginard B. 1985. Co-expression of multiple myosin heavy chaingenes, in addition to a tissue-specific one, in extraocular muscula-ture. J Cell Biol 101:618–629.

Winder SJ, Gibson TJ, Kendrick-Jones J. 1995. Dystrophin andutrophin: the missing links!. FEBS Lett 369:27–33.

Wrogemann K, Pena SDJ. 1976. MItochondrial calcium overload: ageneral mechanism for cell-necrosis in muscle diseases. Lancet1:672–674.


Documents

Eye muscle sparing by the muscular dystrophies: Lessons to be learned?