Internal organization of medial rectus and inferior rectus muscle neurons in the C group of the oculomotor nucleus in monkey.

Mammalian extraocular muscles contain singly innervated twitch muscle fibers (SIF) and multiply innervated nontwitch muscle fibers (MIF). In monkey, MIF motoneurons lie around the periphery of oculomotor nuclei and have premotor inputs different from those of the motoneurons inside the nuclei. The most prominent MIF motoneuron group is the C group, which innervates the medial rectus (MR) and inferior rectus (IR) muscle. To explore the organization of both cell groups within the C group, we performed small injections of choleratoxin subunit B into the myotendinous junction of MR or IR in monkeys. In three animals the IR and MR myotendinous junction of one eye was injected simultaneously with different tracers (choleratoxin subunit B and wheat germ agglutinin). This revealed that both muscles were supplied by two different, nonoverlapping populations in the C group. The IR neurons lie adjacent to the dorsomedial border of the oculomotor nucleus, whereas MR neurons are located farther medially. A striking feature was the differing pattern of dendrite distribution of both cell groups. Whereas the dendrites of IR neurons spread into the supraoculomotor area bilaterally, those of the MR neurons were restricted to the ipsilateral side and sent a focused bundle dorsally to the preganglionic neurons of the Edinger-Westphal nucleus, which are involved in the "near response." In conclusion, MR and IR are innervated by independent neuron populations from the C group. Their dendritic branching pattern within the supraoculomotor area indicates a participation in the near response providing vergence but also reflects their differing functional roles.

Delineation of motoneuron subgroups supplying individual eye muscles in the human oculomotor nucleus.

The oculomotor nucleus (nIII) contains the motoneurons of medial, inferior, and superior recti (MR, IR, and SR), inferior oblique (IO), and levator palpebrae (LP) muscles. The delineation of motoneuron subgroups for each muscle is well-known in monkey, but not in human. We studied the transmitter inputs to human nIII and the trochlear nucleus (nIV), which innervates the superior oblique muscle (SO), to outline individual motoneuron subgroups. Parallel series of sections from human brainstems were immunostained for different markers: choline acetyltransferase combined with glutamate decarboxylase (GAD), calretinin (CR) or glycine receptor. The cytoarchitecture was visualized with cresyl violet, Gallyas staining and expression of non-phosphorylated neurofilaments. Apart from nIV, seven subgroups were delineated in nIII: the central caudal nucleus (CCN), a dorsolateral (DL), dorsomedial (DM), central (CEN), and ventral (VEN) group, the nucleus of Perlia (NP) and the non-preganglionic centrally projecting Edinger-Westphal nucleus (EWcp). DL, VEN, NP, and EWcp were characterized by a strong supply of GAD-positive terminals, in contrast to DM, CEN, and nIV. CR-positive terminals and fibers were confined to CCN, CEN, and NP. Based on location and histochemistry of the motoneuron subgroups in monkey, CEN is considered as the SR and IO motoneurons, DL and VEN as the B- and A-group of MR motoneurons, respectively, and DM as IR motoneurons. A good correlation between monkey and man is seen for the CR input, which labels only motoneurons of eye muscles participating in upgaze (SR, IO, and LP). The CCN contained LP motoneurons, and nIV those of SO. This study provides a map of the individual subgroups of motoneurons in human nIII for the first time, and suggests that NP may contain upgaze motoneurons. Surprisingly, a strong GABAergic input to human MR motoneurons was discovered, which is not seen in monkey and may indicate a functional oculomotor specialization.

Do palisade endings in extraocular muscles arise from neurons in the motor nuclei?

PURPOSE: The purpose of this study was to localize the cell bodies of palisade endings that are associated with the myotendinous junctions of the extraocular muscles. METHODS: Rhesus monkeys received tract-tracer injections (tetramethylrhodamine dextran [TMR-DA] or choleratoxin subunit B [CTB]) into the oculomotor and trochlear nuclei, which contain the motoneurons of extraocular muscles. All extraocular muscles were processed for the combined immunocytochemical detection of the tracer and SNAP-25 or synaptophysin for the visualization of the complete muscle innervation. RESULTS: In all muscles--except the lateral rectus--en plaque and en grappe motor endings, but also palisade endings, were anterogradely labeled. In addition a few tracer-labeled tendon organs were found. One group of tracer-negative nerve fibers was identified as thin tyrosine hydroxylase-positive sympathetic fibers, and a second less numerous group of tracer-negative fibers may originate from the trigeminal ganglia. No cellular or terminal tracer labeling was present within the mesencephalic trigeminal nucleus or the trigeminal ganglia. CONCLUSIONS: These results confirm those of earlier studies and furthermore suggest that the somata of palisade endings are located close to the extraocular motor nuclei--in this case, probably within the C and S groups around the periphery of the oculomotor nucleus. The multiple en grappe endings have also been shown to arise from these cells groups, but it is not possible to distinguish different populations in these experiments.

Parallel ascending vestibular pathways: anatomical localization and functional specialization.

Information from the vestibular nuclei ascending through the brainstem to the oculomotor and trochlear nuclei (NIII, NIV), the interstitial nucleus of Cajal (INC), the pretectum, or thalamus, is thought to be distributed in at least five different pathways. They include the medial longitudinal fasciculus (MLF), the ascending tract of Deiters (ATD), possibly the brachium conjuctivum (BC), the crossing ventral tegmental tracts (CVTT), and a recently observed ipsilateral pathway close to the medial lemniscus, possibly the equivalent of the ipsilateral vestibulo-thalamic tract (IVTT). This short review describes the location of these ascending tracts, their function with respect to ocular motor control and perception, and their clinical relevance. There is evidence that the MLF carries mainly information from the canals to NIII, NIV, INC, and possibly the thalamus, whereas otolith signals may ascend in the CVTT, along with excitatory anterior canal connections. The evidence for BC as a specific vestibulo-oculomotor pathway is weak and could be the result of the initial observations of CVTT. The ATD carries mostly ipsilateral otolithic information to the medial and inferior recti subgroups in NIII and the Edinger-Westphal nucleus. In the rostral pons an ipsilateral vestibular pathway was seen lying close to the medial lemniscus. This anatomical projection could be the equivalent of the IVTT, bypassing the ocular motor centers and projecting to the thalamus. The IVTT mediates perception of verticality and may be part of a fast three-neuron vestibulo-thalamo-cortical pathway, which provides the multisensory cortical system for spatial orientation and self-motion-perception with information about head acceleration.

Torsional deviations with voluntary saccades caused by a unilateral midbrain lesion.

Three dimensional eye rotations were measured using the magnetic search coil technique in a patient with a lesion of the right rostral interstitial nucleus of the medial longitudinal fasciculus (RIMLF) and in four control subjects. Up to 10° contralesional torsional deviations with each voluntary saccade were revealed, which also could be seen during bedside examination. There was no spontaneous nystagmus. Based on MRI criteria, the lesion involved the RIMLF but spared the interstitial nucleus of Cajal. To date, this deficit has not been described in patients. Our results support the hypothesis that the vertical-torsional saccade generator in humans is organised similarly as in monkeys: each RIMLF encodes torsional saccades in one direction, while both participate in vertical saccades.

Mapping the oculomotor system.

Over the last three decades and together with Bernard Cohen, Volker Henn, Ulrich Büttner, and Anja Horn, it has been possible to morphologically identify several functional cell groups in the oculomotor system: the medium-sized horizontal excitatory and inhibitory burst neurons (EBNs, IBNs) in the paramedian pontine reticular formation (PPRF), the more sparsely scattered vertical EBNs in the rostral interstitial nucleus of the MLF (RIMLF), and the typically elongated omnipause neurons (OPNs) in nucleus raphé interpositus--all essential for the generation of saccades. In contrast, the role of the central mesencephalic reticular formation (cMRF) in saccades is more complex, as is the morphological outlining of its borders. A detailed study of the extraocular motoneurons showed that they can be divided into two separate types: those for singly innervated (twitch) muscle fibres (SIFs) and those for multiply innervated (non-twitch) muscle fibres (MIFs). The two motoneuron types receive different premotor afferents, proving that MIF and SIF motoneurons have different functions. The cell groups were outlined by different tract tracing methods including rabies virus. The localization and histochemical characterization of all these functional cell groups in monkey formed the basis for the identification of the homologous groups in the human brainstem. Taken together these studies provide a neuroanatomical background for understanding clinical eye movement disorders.

Neuronal signalling expression profiles of motoneurons supplying multiply or singly innervated extraocular muscle fibres in monkey.

Motoneurons of the oculomotor nucleus subserving multiply innervated muscle fibres (MIF) receive different afferent inputs from the motoneurons subserving singly innervated muscle fibres (SIF). We asked whether MIF and SIF motoneurons have different neurotransmitter signalling expression profiles. Adult rhesus monkey extraocular muscles were injected with the retrograde tracer cholera toxin. Sections were then stained for various neurotransmitter-signalling markers. MIF motoneurons showed less glutamate receptor 4 (GluR4) and N-methyl-D-aspartate receptor 1 (NMDAR1) immunoreactivity, but showed similar amounts of glutamic acid decarboxylase (GAD) immunoreactive afferent terminals, compared to SIF motoneurons. This difference in excitatory neurotransmitter receptor expression may explain selective oculomotor deficits and allow development of selective pharmacotherapy in the future.

Brainstem circuits controlling lid-eye coordination in monkey.

In primate, the M-group is a cell cluster in the rostral mesencephalon which contains premotor neurons for the levator palpebrae (LP) and upward-pulling eye muscles. It is therefore thought to play a role in lid-eye coupling during vertical saccades. To further elucidate its role, the afferents to the M-group and LP motoneurons were studied in monkeys. Anterograde tracer injections were placed in one of the three eye-movement-related areas: 1. superior colliculus (SC), 2. interstitial nucleus of Cajal (INC), and 3. the omnipause neuron (OPN) region. Injections into the medial SC subtending upward saccades led to afferent labelling of the ipsilateral M-group and the adjacent rostral interstitial nucleus of the medial longitudinal fascicle (RIMLF), whereas only RIMLF was labelled after an injection into the lateral SC subtending downward saccades. Both RIMLF and M-group received bilateral projections from INC, but only RIMLF received glycineric inputs from the OPN region. This connectivity pattern supports the hypothesis that the M-group mediates lid-eye coupling during vertical upgaze, but is indirectly driven by collaterals of saccadic burst neurons in the RIMLF during lid saccades. A selective projection from the OPN area to the LP motoneurons, but not to other oculomotor neurons is reported here for the first time. The result is supported by the presence of glycinergic terminals only over LP motoneurons, and implies that a subset of OPNs may directly trigger saccade-related blinks.

Selective, circuit-wide sparing of floccular connections in hereditary olivopontine cerebellar atrophy with slow saccades.

We present a systems-oriented histopathologic analysis of the ocular motor control circuits in the cerebellum and brainstem from a patient with a hereditary form of olivopontine cerebellar atrophy of the Wadia type, which has a characteristic ocular motor presentation of slow saccades but relative preservation of smooth pursuit and gaze-holding. This differential pattern of clinical involvement is associated with a lobule-specific pattern of cerebellar degeneration. We asked whether these patterns of sparing and degeneration were consistent throughout the associated deep cerebellar and brainstem structures. Specimens were fixed in formalin, embedded in paraffin, and stained for various markers. We found that elements of the floccular and nodular pathways, controlling smooth pursuit and vestibular reflexes, were relatively spared, particularly those structures that are interconnected with the medial regions. Conversely, the elements of the dorsal vermis pathway controlling saccade adaptation were relatively involved. This subregional specificity of degeneration further defines possible areas of investigation for elucidating pathophysiology, testing biomarkers of disease, and developing areas for therapeutic intervention.

Horizontal saccadic palsy associated with gliosis of the brainstem midline.

We studied premotor cell groups involved in the generation of saccades in a patient with a disturbance of voluntary horizontal gaze. The only neurological symptom found was a slowing of horizontal saccades, reported since birth and unaltered over his lifetime. We attribute this disorder, for the first time, to a fibrous gliosis of the brainstem midline, which may disrupt neuronal elements of the horizontal saccade generator crossing the brainstem midline, but it caused no obvious loss of omnipause-, excitatory burst-, and inhibitory burst neurons. No neuronal loss or demyelination, was apparent elsewhere in the brainstem; but there was evidence of an ependymal infection throughout the entire ventricular system. A diagnosis of Gaucher disease was made from the bone marrow of this patient shortly before his death, but for several reasons we considered this complication unlikely to be the cause of the saccadic disorder.

Perioculomotor cell groups in monkey and man defined by their histochemical and functional properties: reappraisal of the Edinger-Westphal nucleus.

The perioculomotor region contains several functional cell groups, including parasympathetic preganglionic neurons of the ciliary ganglion, motoneurons of multiply innervated muscle fibers (MIF) of extraocular muscles, and urocortin-positive neurons. In this study, midbrain sections of monkey and human were treated with antibodies against choline acetyltransferase (ChAT), cytochrome oxidase (CytOx), nonphosphorylated neurofilaments (NP-NF), chondroitin sulfate proteoglycan (CSPG), and urocortin (UCN) to identify them by their histochemical properties. To facilitate the comparison between species, a new nomenclature was introduced (see also May et al., 2007), which designates these perioculomotor cell populations (pIII) in terms of their function and histochemical properties. The name Edinger-Westphal nucleus (EW) is kept for the cytoarchitecturally defined cell group traditionally considered as the location of preganglionic neurons of the ciliary ganglion. In monkey, the EW contains ChAT-positive presumed preganglionic neurons, and is therefore termed EW(PG), but in contrast human EW consists of noncholinergic UCN-positive neurons, and is therefore termed EW(U). In human, the presumed preganglionic neurons were found dorsal to EW(U), as an inconspicuous group of ChAT- and CytOx-positive neurons. They were interspersed with prominent CSPG-positive cells, a pattern also present in monkey. For the first time, the MIF motoneurons could be identified around the medial aspect of the human oculomotor nucleus as a group of ChAT-positive neurons that lack CSPG-positive perineuronal nets. Moreover, the Perlia nucleus was found to share the histochemical properties of oculomotor twitch motoneurons. The present results form the basis for addressing the appropriate functional cell groups in correlative clinicopathological studies.

Torsional deviations with voluntary saccades caused by a unilateral midbrain lesion.

Three dimensional eye rotations were measured using the magnetic search coil technique in a patient with a lesion of the right rostral interstitial nucleus of the medial longitudinal fasciculus (RIMLF) and in four control subjects. Up to 10 degree contralesional torsional deviations with each voluntary saccade were revealed, which also could be seen during bedside examination. There was no spontaneous nystagmus. Based on MRI criteria, the lesion involved the RIMLF but spared the interstitial nucleus of Cajal. To date, this deficit has not been described in patients. Our results support the hypothesis that the vertical-torsional saccade generator in humans is organised similarly as in monkeys: each RIMLF encodes torsional saccades in one direction, while both participate in vertical saccades.

Anatomy of the oculomotor system.

The sensory and motor control of eye muscles are considered in this chapter. Eye muscles differ from skeletal muscles in several ways. One is the absence of muscle spindles and Golgi tendon organs in the eye muscles of some species, and their poor development in others. Second, eye muscles have an inner 'global layer', and the outer 'orbital layer', each containing different types of muscle fiber. Third, eye muscles contain not only twitch muscle fibers with a single endplate zone (SIFs), but also nontwitch muscle fibers with multiple endplate zones (MIFs), which are otherwise absent from mammalian muscles. There are cuffs of nerve terminals, called palisade endings, around the myotendinous junctions of global layer MIFs. Palisade endings are unique to eye muscles, and have been found in all mammalian species investigated up to now. The function of palisade endings is uncertain, but it is possible that they are 'sensory receptors'. Motoneurons innervating the eye muscles lie in the oculomotor, trochlear and abducens motor nuclei, and are contacted by several relatively independent premotor networks, which generate different types of eye movements such as saccades, vestibulo-ocular reflexes, optokinetic responses, smooth pursuit convergence or gaze-holding. In each motor nucleus, the motoneurons can be divided into two distinct sets: the first set innervating SIF muscle fibers and receiving inputs from all oculomotor premotor networks, and the second set innervating the MIFs and receiving premotor afferents from the gaze holding, convergence or smooth pursuit premotor networks, but not from the saccadic and vestibulo-oculomotor networks. We suggest that the SIF motoneurons and muscles are more suited to driving eye movements, and the MIF motoneurons and muscles to setting the tonic tension in eye muscles. Furthermore the 'palisade ending-MIF unit' may be part of a sensory feedback system in eye muscles, which should be considered in association with the causes and treatment of strabismus.

Histochemical differences between motoneurons supplying multiply and singly innervated extraocular muscle fibers.

The extraocular muscle fibers of vertebrates can be classified into two categories: singly innervated fibers (SIFs) and multiply innervated fibers (MIFs). In monkeys, the motoneurons of SIFs lie within the oculomotor, trochlear, and abducens nucleus, whereas the motoneurons of MIFs appear in separate subgroups in the periphery of the classical nuclei borders. In the present study, we investigated the histochemical properties of SIF and MIF motoneurons by using combined tract-tracing and immunofluorescence techniques. In monkeys, SIF and MIF motoneurons of extraocular muscles were identified by tracer injections into the belly or the distal myotendinous junction of the medial or lateral rectus muscle. Alternatively, the motoneurons were identified by choline acetyltransferase immunostaining. These techniques were combined with the detection of histochemical markers for perineuronal nets, nonphosphorylated neurofilaments, parvalbumin, or cytochrome oxidase. The experiments revealed that the MIF motoneurons in the periphery of the motonuclei do not contain nonphosphorylated neurofilaments or parvalbumin and lack perineuronal nets. In contrast, SIF motoneurons express all markers at high intensity. Cytochrome oxidase immunostaining was found in both motoneuron populations. An additional population of motoneurons with "MIF properties" was identified within the boundaries of the abducens nucleus, which could represent the motoneurons innervating MIFs in the orbital layer of lateral rectus muscle. Our data provide evidence that SIF and MIF motoneurons, which can be correlated with twitch motoneurons and presumed non-twitch motoneurons, differ in their histochemical properties. The absence of perineuronal nets, nonphosphorylated neurofilaments, and parvalbumin may help to identify the homologous MIF motoneurons in other species, including humans.

Spatial properties of central vestibular neurons of monkeys after bilateral lateral canal nerve section.

Thirty-seven neurons were recorded in the superior vestibular nucleus (SVN) of two cynomolgus monkeys 1-2 yr after bilateral lateral canal nerve section to test whether the central neurons had spatially adapted for the loss of lateral canal input. The absence of lateral canal function was verified with eye movement recordings. The relation of unit activity to the vertical canals was determined by oscillating the animals about a horizontal axis with the head in various orientations relative to the axis of rotation. Animals were also oscillated about a vertical axis while upright or tilted in pitch. In the second test, the vertical canals are maximally activated when the animals are tilted back about -50 degrees from the spatial upright and the lateral canals when the animals are tilted forward about 30 degrees . We reasoned that if central compensation occurred, the head orientation at which the response of the vertical canal-related neurons was maximal should be shifted toward the plane of the lateral canals. No lateral canal-related units were found after nerve section, and vertical canal-related units were found only in SVN not in the rostral medial vestibular nucleus. SVN canal-related units were maximally activated when the head was tilted back at -47 +/- 17 and -50 +/- 12 degrees (means +/- SD) in the two animals, close to the predicted orientation of the vertical canals. This indicated that spatial adaptation of vertical canal-related vestibular neurons had not occurred. There were substantial neck and/or otolith-related inputs activating the vertical canal-related neurons in the nerve-sectioned animals, which could have contributed to oculomotor compensation after nerve section.

Proprioception and palisade endings in extraocular eye muscles.

Palisade endings occur only in extraocular muscles, and their function is unknown. They form a cuff of nerve terminals around the tips of muscle fibers. We describe here the advantages of using antibodies to a synaptosomal-associated protein (SNAP-25) to study properties of palisade endings in man, monkey, and rat. The stain can be combined readily with other immunofluorescence procedures, and results suggest that the synapses of palisade endings do not bind alpha-bungarotoxin (i.e., are not motor), nor do they contain substance P. These double-labeling data support the hypothesis that palisade endings are non-nociceptive sensory receptors, and could serve a proprioceptive function. With SNAP-25 immunolabeling, palisade endings were identified in the rat for the first time. Thus, palisade endings appear to be present in all vertebrate extraocular muscles studied to date. Their apparent universality, which contrasts with the more variable manifestation of extraocular muscle spindles and Golgi tendon organs, would be expected if proprioceptive feedback is necessary to the function of the ocular motor system, and if palisade endings are the critical proprioceptive structure.

Palisade endings in extraocular eye muscles revealed by SNAP-25 immunoreactivity.

Palisade endings form a cuff of nerve terminals around the tip of muscle fibres. They are found only in extraocular muscles, but no definite evidence for their role in eye movements has been established. Palisade endings have been reported in all species so far investigated except the rat. In this study we demonstrate that antibodies against SNAP-25, the synaptosomal associated protein of 25 kDa, reliably visualize the complete motor, sensory and autonomic innervation of the extraocular muscles in human, monkey and rat. The SNAP-25 antibody can be combined with other immunofluorescence procedures, and is used here to study properties of palisade endings. With SNAP-25 immunolabelling putative palisade endings are identified in the rat for the first time. They are not well branched, but fulfil several criteria of palisade endings, being associated with non-twitch fibres as shown by double labelling with 'myosin heavy chain slow-twitch' antibodies. The putative palisade endings of the rat lack alpha-bungarotoxin binding, which implies that these synapses are sensory. If palisade endings are sensory then they could function as an eye muscle proprioceptor. They seem to be a general feature of all vertebrate eye muscles, unlike the other two extraocular proprioceptors, muscle spindles and Golgi tendon organs, the presence of which varies widely between species.

Twitch and nontwitch motoneuron subgroups in the oculomotor nucleus of monkeys receive different afferent projections.

Motoneurons in the primate oculomotor nucleus can be divided into two categories, those supplying twitch muscle fibers and those supplying nontwitch muscle fibers. Recent studies have shown that twitch motoneurons lie within the classical oculomotor nucleus (nIII), and nontwitch motoneurons lie around the borders. Nontwitch motoneurons of medial and inferior rectus are in the C group dorsomedial to nIII, whereas those of inferior oblique and superior rectus lie near the midline are in the S group. In this anatomical study, afferents to the twitch and nontwitch subgroups of nIII have been anterogradely labeled by injections of tritiated leucine into three areas and compared. 1) Abducens nucleus injections gave rise to silver grain deposits over all medial rectus subgroups, both twitch and nontwitch. 2) Laterally placed vestibular complex injections that included the central superior vestibular nucleus labeled projections only in twitch motoneuron subgroups. However, injections into the parvocellular medial vestibular nucleus (mvp), or Y group, resulted in labeled terminals over both twitch and nontwitch motoneurons. 3) Pretectal injections that included the nucleus of the optic tract (NOT), and the olivary pretectal nucleus (OLN), labeled terminals only over nontwitch motoneurons, in the contralateral C group and in the S group. Our study demonstrates that twitch and nontwitch motoneuron subgroups do not receive identical afferent inputs. They can be controlled either in parallel, or independently, suggesting that they have basically different functions. We propose that twitch motoneurons primarily drive eye movements and nontwitch motoneurons the tonic muscle activity, as in gaze holding and vergence, possibly involving a proprioceptive feedback system.

The neuroanatomical basis of oculomotor disorders: the dual motor control of extraocular muscles and its possible role in proprioception.

Current investigations show that two separate sets of motoneurons control the extraocular eye muscles, and that is there is a dual final common pathway. We propose that one set of motoneurons are the major source of tension generating eye movements, whereas the other may participate in a proprioceptive system concerned more with the exact alignment and stabilization of the eyes. In this article we discuss the structures that may participate in the proprioceptive circuits; and consider several recent publications in the light of this sensory feedback hypothesis, emphasizing the relevance to eye movement disorders.