Joseph L. Demer

Human TUBB3 Mutations Perturb Microtubule Dynamics, Kinesin Interactions, and Axon Guidance

We report that eight heterozygous missense mutations in TUBB3, encoding the neuron-specific beta-tubulin isotype III, result in a spectrum of human nervous system disorders that we now call the TUBB3 syndromes. Each mutation causes the ocular motility disorder CFEOM3, whereas some also result in intellectual and behavioral impairments, facial paralysis, and/or later-onset axonal sensorimotor polyneuropathy. Neuroimaging reveals a spectrum of abnormalities including hypoplasia of oculomotor nerves and dysgenesis of the corpus callosum, anterior commissure, and corticospinal tracts. A knock-in disease mouse model reveals axon guidance defects without evidence of cortical cell migration abnormalities. We show that the disease-associated mutations can impair tubulin heterodimer formation in vitro, although folded mutant heterodimers can still polymerize into microtubules. Modeling each mutation in yeast tubulin demonstrates that all alter dynamic instability whereas a subset disrupts the interaction of microtubules with kinesin motors. These findings demonstrate that normal TUBB3 is required for axon guidance and maintenance in mammals.

Heterozygous mutations of the kinesin KIF21A in congenital fibrosis of the extraocular muscles type 1 (CFEOM1).

Congenital fibrosis of the extraocular muscles type 1 (CFEOM1; OMIM #135700) is an autosomal dominant strabismus disorder associated with defects of the oculomotor nerve. We show that individuals with CFEOM1 harbor heterozygous missense mutations in a kinesin motor protein encoded by KIF21A. We identified six different mutations in 44 of 45 probands. The primary mutational hotspots are in the stalk domain, highlighting an important new role for KIF21A and its stalk in the formation of the oculomotor axis.

Heterotopic Muscle Pulleys or Oblique Muscle Dysfunction

INTRODUCTION The description of connective tissue sleeves that function as pulleys for the rectus extraocular muscles (EOMs) suggests that abnormalities of EOM pulley position might provide a mechanical basis for some forms of incomitant strabismus. Pulleys determine the paths and thus the pulling directions of EOMs. METHODS High-resolution magnetic resonance images spanning the orbits were obtained in primary position, upgaze, and downgaze for each subject. Paths of the EOMs were measured with reference to the orbital center and permitted inference of pulley locations. RESULTS Data from 18 orbits of orthotropic subjects defined means and SDs of normal EOM pulley coordinates. Eight patients, aged 17 to 60 years, had heterotopic EOM pulleys, defined as displaced at least 2 SDs from normal. We found one to eight heterotopic pulleys (considering both orbits) in each of four patients who had been diagnosed with marked superior oblique (SO) overaction and mild to marked inferior oblique (IO) underaction. Each patient had superior mislocation of at least one lateral rectus pulley by 1.8 to 4.9 mm. Three patients diagnosed with mild to moderate IO overaction and mild to moderate SO underaction in only one orbit had one to three heterotopic EOM pulleys. Each of those patients had at least one lateral rectus pulley inferiorly dislocated by 1.9 to 4.9 mm. The final patient, who was diagnosed with mild IO underaction and normal SO function bilaterally, had bilateral superior mislocation of the medial rectus pulleys by greater than 2 mm. Computer simulations using the Orbit program (Eidactics, San Francisco) incorporating individually measured pulley positions reproduced the clinical patterns of incomitant strabismus in all cases without postulating abnormalities of oblique muscle innervation or contractility. CONCLUSION Heterotopic EOM pulleys can cause patterns of incomitant strabismus that have been attributed to oblique muscle dysfunction. Even isolated mislocations of less than 2 mm, coupled with smaller mislocations of the other pulleys, can produce the clinical appearance of bilateral oblique dysfunction. Pulley heterotopy should be considered in the differential diagnosis of incomitant strabismus and oblique dysfunction.

The Orbital Pulley System: A Revolution in Concepts of Orbital Anatomy

Abstract: Magnetic resonance imaging (MRI) now enables precise visualization of the mechanical state of the living human orbit. Resulting insights have motivated histological re‐examination of human and simian orbits, providing abundant consistent evidence for the active pulley hypothesis, a re‐formulation of ocular motor physiology. Each extraocular muscle (EOM) consists of a global layer (GL) contiguous with the tendon and inserting on the eyeball, and a similar‐sized orbital layer (OL) inserting on a connective tissue ring forming the EOM pulley. The pulley controls the EOM path and serves as the EOMs functional origin. Activity of the OL positions the pulley along each rectus EOM to assure that its pulling direction shifts by half the change in ocular orientation, the half‐angle behavior characteristic of a linear ocular motor plant. Half‐angle behavior is equivalent to Listings law of ocular torsion, and makes 3‐D ocular rotations effectively commutative. Pulleys are configured to maintain oblique EOM paths orthogonal to half‐angle behavior, and violate Listings law during the vestibulo‐ocular reflex. Rectus pulley positions shift during convergence, facilitating stereopsis.

Effect of aging on human rectus extraocular muscle paths demonstrated by magnetic resonance imaging

PURPOSE To compare normal functional anatomy of rectus extraocular muscles (EOMs) and pulleys in normal older humans with previously reported findings in younger subjects. DESIGN Experimental study of the orbits of normal healthy older volunteers by magnetic resonance imaging (MRI). METHODS In planes perpendicular to the orbital axis, contiguous MRI images spanned the anteroposterior extents of 22 orbits in 12 older adults with an average age of 65.2 years (range, 56-74). Images were obtained in central gaze in all subjects and repeated in supraduction, infraduction, abduction, and adduction in some subjects. Mean EOM cross-sectional area centroids were normalized to an oculocentric coordinate system and plotted over the length of each EOM to determine paths. RESULTS Compared with images obtained using identical technique in 12 younger subjects (average age, 28.5 years, range 21-33), the horizontal rectus EOMs in the 12 older subjects were significantly displaced inferiorly throughout the anteroposterior extent of the orbit. The vertical rectus EOM was positioned identically to those of younger subjects. Inflections in EOM paths produced by the connective tissue pulleys could not be determined in most older subjects, because of difficulties in maintaining extreme eccentric gaze. For one subject who was able to do this, the anteroposterior location of the medial rectus pulley inferred from path inflection was similar to that of younger subjects. CONCLUSIONS The horizontal rectus EOMs are displaced inferiorly in the elderly relative to the globe center. This displacement presumably reflects an inferior location of the corresponding pulleys, partially converting horizontal rectus EOM force to depression. This may contribute to the observed impairment of elevation in older people and predispose them to a characteristic pattern of incomitant strabismus.

The human horizontal vestibulo-ocular reflex during combined linear and angular acceleration.

Abstract We employed binocular magnetic search coils to study the vestibulo-ocular reflex (VOR) and visually enhanced vestibulo-ocular reflex (VVOR) of 15 human subjects undergoing passive, whole-body rotations about a vertical (yaw) axis delivered as a series of pseudorandom transients and sinusoidal oscillations at frequencies from 0.8 to 2.0 Hz. Rotations were about a series of five axes ranging from 20 cm posterior to the eyes to 10 cm anterior to the eyes. Subjects were asked to regard visible or remembered targets 10 cm, 25 cm, and 600 cm distant from the right eye. During sinusoidal rotations, the gain and phase of the VOR and VVOR were found to be highly dependent on target distance and eccentricity of the rotational axis. For axes midway between or anterior to the eyes, sinusoidal gain decreased progressively with increasing target proximity, while, for axes posterior to the otolith organs, gain increased progressively with target proximity. These effects were large and highly significant. When targets were remote, rotational axis eccentricity nevertheless had a small but significant effect on sinusoidal gain. For sinusoidal rotational axes midway between or anterior to the eyes, a phase lead was present that increased with rotational frequency, while for axes posterior to the otolith organs phase lag increased with rotational frequency. Transient trials were analyzed during the first 25 ms and from 25 to 80 ms after the onset of the head rotation. During the initial 25 ms of transient head rotations, VOR and VVOR gains were not significantly influenced by rotational eccentricity or target distance. Later in the transient responses, 25–80 ms from movement onset, both target distance and eccentricity significantly influenced gain in a manner similar to the behavior during sinusoidal rotation. Vergence angle generally remained near the theoretically ideal value during illuminated test conditions (VVOR), while in darkness vergence often varied modestly from the ideal value. Regression analysis of instantaneous VOR gain as a function of vergence demonstrated only a weak correlation, indicating that instantaneous gain is not likely to be directly dependent on vergence. A model was proposed in which linear acceleration as sensed by the otoliths is scaled by target distance and summed with angular acceleration as sensed by the semicircular canals to control eye movements. The model was fit to the sinusoidal VOR data collected in darkness and was found to describe the major trends observed in the data. The results of the model suggest that a linear interaction exists between the canal and otolithic inputs to the VOR.

Extraocular connective tissue architecture

Extraocular muscle pulleys, now well known to be kinematically significant extraocular structures, have been noted in passing and described in fragments several times over the past two centuries. They were late to be fully appreciated because biomechanical modeling of the orbit was not available to derive their kinematic consequences, and because pulleys are distributed condensations of collagen, elastin and smooth muscle (SM) that are not sharply delineated. Might other mechanically significant distributed extraocular structures still be awaiting description?An imaging approach is useful for describing distributed structures, but does not seem suitable for assessing mechanical properties. However, an image that distinguished types and densities of constituent tissues could give strong hints about mechanical properties. Thus, we have developed methods for producing three dimensional (3D) images of extraocular tissues based on thin histochemically processed slices, which distinguish collagen, elastin, striated muscle and SM. Overall tissue distortions caused by embedding for sectioning, and individual-slice distortions caused by thin sectioning and subsequent histologic processing were corrected by ordered image warping with intrinsic fiducials. We describe an extraocular structure, partly included in Lockwoods ligament, which contains dense elastin and SM bands, and which might refine horizontal eye alignment as a function of vertical gaze, and torsion in down-gaze. This active structure might therefore be a factor in strabismus and a target of therapeutic intervention.

Magnetic Resonance Imaging of the Functional Anatomy of the Inferior Rectus Muscle in Superior Oblique Muscle Palsy

PURPOSE Biomechanical modeling consistently indicates that superior oblique (SO) muscle weakness alone is insufficient to explain the large hypertropia often observed in SO muscle palsy. Magnetic resonance imaging (MRI) was used to investigate if any size or contractility changes in the inferior rectus (IR) muscle may contribute. DESIGN Prospective, case-control study. PARTICIPANTS Seventeen patients with unilateral SO muscle palsy and 18 orthotropic subjects. METHODS Surface coils were used to obtain sets of contiguous, 2-mm-thick, high-resolution, coronal MRI views in different gazes. Cross-sectional areas of the IR and SO muscles were determined in supraduction and infraduction for evaluation of size and contractility. Diagnosis of SO muscle palsy was based on clinical presentations, subnormal contractility, and SO muscle size less than the normal 95% confidence limit. MAIN OUTCOME MEASURES Cross-sectional areas of the IR and SO muscles. RESULTS Patients had 15.9+/-7.2 prism diopters (Delta; mean+/-standard deviation) of central gaze hypertropia and exhibited ipsilesional SO muscle atrophy and subnormal contractility. Mean ipsilesional, contralesional, and normal IR muscle cross-sections were 28.5+/-3.5 mm(2), 31.9+/-3.8 mm(2), and 31.8+/-5.8 mm(2), whereas mean contractility was 16.5+/-3.8 mm(2), 20.5+/-4.1 mm(2), and 16.6+/-4.8 mm(2), respectively. Ipsilesional IR muscle cross-section and contractility was significantly less than contralesional cross-section and contractility (P<0.01). CONCLUSIONS In SO muscle palsy, the contralesional IR muscle is larger and more contractile than the ipsilesional IR muscle, reflecting likely neurally mediated changes that augment the relatively small hypertropia resulting from SO muscle weakness alone. Recession of the hyperfunctioning contralesional IR muscle recession in SO muscle palsy is a physiologic therapy.

Diffusion tensor MRI shows abnormal brainstem crossing fibers associated with ROBO3 mutations

Horizontal gaze palsy with progressive scoliosis (HGPPS) is caused by mutations in the ROBO3 gene, critical for the crossing of long ascending medial lemniscal and descending corticospinal tracts in the medulla. Diffusion tensor imaging in a patient with HGGPS revealed the absence of major pontine crossing fiber tracts and no decussation of the superior cerebellar peduncles. Mutations in the ROBO3 gene lead to a widespread lack of crossing fibers throughout the brainstem.

Surgical implications of the rectus extraocular muscle pulleys

PURPOSE Magnetic resonance imaging (MRI) shows that the paths of rectus extraocular muscle bellies remain fixed in the orbit during large ocular rotations, and across large surgical transpositions of their insertions. This stability of muscle paths is due to their passage through pulleys which are coupled to the orbit and located in a coronal plane anterior to the muscle bellies near the equator of the globe. Autopsy studies have shown the pulleys to be fibroelastic sleeves consisting of dense bands of collagen and elastin, suspended from the orbit and adjacent extraocular muscle sleeves by bands of similar composition. Immunohistochemical studies have revealed substantial smooth muscle in the pulley suspensions and in posterior Tenons fascia. The pulleys function as mechanical origins of the rectus extraocular muscles in the sense of determining extraocular muscle pulling directusons. This study was conducted to determine the theoretical effects of the pulleys on the outcome of rectus transposition surgery. METHODS The functional and anatomical evidence for the existence of the rectus extraocular muscle pulleys was reviewed. In two patients, binocular alignment data were collected using the Hess screen test before and after vertical rectus transposition surgeries for lateral rectus paralysis. Paths of the rectus extraocular muscles were determined using high resolution MRI. The OrbitTM 1.5 extraocular biosimulation program was employed to compute theoretical binocular alignment and muscle paths, under alternative conditions including or omitting the pulleys. RESULTS Pulleys are required to account for observed paths of rectus extraocular muscles following transposition surgery. In the absence of pulleys, transposition of the superior and inferior rectus muscles to the lateral rectus insertion for abducens paralysis would result in bizarre ocular misalignments not observed clinically. CONCLUSIONS The human orbit contains specialized musculofibroelastic tissues in and just posterior to Tenons fascia, which serve as pulleys, determining actions of rectus extraocular muscles. These pulleys are located in a roughly coronal plane just posterior to the equator of the globe. Unimpaired pulley function is essential to effective muscle transposition surgery.