Human middle-ear muscles rarely contract in anticipation of acoustic impulses: Implications for hearing risk assessments.

The current study addressed the existence of an anticipatory middle-ear muscle contraction (MEMC) as a protective mechanism found in recent damage-risk criteria for impulse noise exposure. Specifically, the experiments reported here tested instances when an exposed individual was aware of and could anticipate the arrival of an acoustic impulse. In order to detect MEMCs in human subjects, a laser-Doppler vibrometer (LDV) was used to measure tympanic membrane (TM) motion in response to a probe tone. Here we directly measured the time course and relative magnitude changes of TM velocity in response to an acoustic reflex-eliciting (i.e. MEMC eliciting) impulse in 59 subjects with clinically assessable MEMCs. After verifying the presence of the MEMC, we used a classical conditioning paradigm pairing reflex-eliciting acoustic impulses (unconditioned stimulus, UCS) with various preceding stimuli (conditioned stimulus, CS). Changes in the time-course of the MEMC following conditioning were considered evidence of MEMC conditioning, and any indication of an MEMC prior to the onset of the acoustic elicitor was considered an anticipatory response. Nine subjects did not produce a MEMC measurable via LDV. For those subjects with an observable MEMC (n = 50), 48 subjects (96%) did not show evidence of an anticipatory response after conditioning, whereas only 2 subjects (4%) did. These findings reveal that MEMCs are not readily conditioned in most individuals, suggesting that anticipatory MEMCs are not prevalent within the general population. The prevalence of anticipatory MEMCs does not appear to be sufficient to justify inclusion as a protective mechanism in auditory injury risk assessments.

Wideband detection of middle ear muscle activation using swept-tone distortion product otoacoustic emissions.

The measurement of efferent-induced suppression of otoacoustic emissions (OAEs) using contralateral acoustic stimulation (CAS) is complicated by potential contamination by the middle ear muscle reflex (MEMR), particularly at moderate to high CAS levels. When logarithmically sweeping primaries are used to measure distortion product otoacoustic emissions, the level and phase of the primaries at the entrance of the ear canal may be monitored simultaneously along with the OAEs elicited by the swept-tones. A method of detecting MEMR activation using swept-tones is presented in which the differences in the primaries in the ear canal with and without CAS are examined, permitting evaluation of MEMR effects over a broad frequency range. A range of CAS levels above and below expected contralateral acoustic reflex thresholds permitted evaluation of conditions with and without MEMR activation.

Distribution of substance P and the calcitonin gene-related peptide in the human tensor tympani muscle.

Substance P-immunoreactive nerve fiber (SP-IR NF) and calcitonin gene-related peptide-immunoreactive nerve fiber (CGRP-IR NF) are important mediators of neurogenic inflammation and blood supply. SP-IR and CGRP-IR NFs in the tensor tympani muscle (TTM) of the human middle ear have yet to be described. In this study, the TTM, tympanic membrane, malleus in the middle ear and tensor veli palatini muscle (TVPM) were examined by whole-mount immunohistochemistry in tissue from Japanese subjects. Thirteen human cadavers (ranging in age from 46 to 90 years) were used in this study. SP-IR and CGRP-IR NFs were primarily found on vessels at the origin, insertion and belly of the surface of the TTM and on the internal surface of the tympanic membrane. These neural factors were also detected on the surface of the malleus and the insertion of the TVPM. Therefore, our results indicate that existence of the SP-IR and CGRP-IR NFs of the TTM and the TVPM may reflect muscle properties involved in pain or inflammation of the middle ear.

Tensor tympani motoneurons receive mostly excitatory synaptic inputs.

The tensor tympani is a middle ear muscle that contracts in two different situations: in response to sound or during voluntary movements. To gain insight into the inputs and neural regulation of the tensor tympani, we examined the ultrastructure of synaptic terminals on labeled tensor tympani motoneurons (TTMNs) using transmission electron microscopy. Our sample of six TTMNs received 79 synaptic terminals that formed 126 synpases. Two types of synapses are associated with round vesicles and form asymmetric junctions (excitatory morphology). One of these types has vesicles that are large and round (Lg Rnd) and the other has vesicles that are smaller and round (Sm Rnd) and also contains at least one dense core vesicle. A third synapse type has inhibitory morphology because it forms symmetric synapses with pleomorphic vesicles (Pleo). These synaptic terminals can be associated with TTMN spines. Two other types of synapse are found on TTMNs but they are uncommon. Synaptic terminals of all types form multiple synapses but those from a single terminal are always the same type. Terminals with Lg Rnd vesicles formed the most synpases per terminal (avg. 2.73). Together, the synaptic terminals with Lg Rnd and Sm Rnd vesicles account for 62% of the terminals on TTMNs, and they likely represent the pathways driving the contractions in response to sound or during voluntary movements. Having a high proportion of excitatory inputs, the TTMN innervation is like that of stapedius motoneurons but proportionately different from other types of motoneurons.

Auditory brainstem circuits that mediate the middle ear muscle reflex.

The middle ear muscle (MEM) reflex is one of two major descending systems to the auditory periphery. There are two middle ear muscles (MEMs): the stapedius and the tensor tympani. In man, the stapedius contracts in response to intense low frequency acoustic stimuli, exerting forces perpendicular to the stapes superstructure, increasing middle ear impedance and attenuating the intensity of sound energy reaching the inner ear (cochlea). The tensor tympani is believed to contract in response to self-generated noise (chewing, swallowing) and non-auditory stimuli. The MEM reflex pathways begin with sound presented to the ear. Transduction of sound occurs in the cochlea, resulting in an action potential that is transmitted along the auditory nerve to the cochlear nucleus in the brainstem (the first relay station for all ascending sound information originating in the ear). Unknown interneurons in the ventral cochlear nucleus project either directly or indirectly to MEM motoneurons located elsewhere in the brainstem. Motoneurons provide efferent innervation to the MEMs. Although the ascending and descending limbs of these reflex pathways have been well characterized, the identity of the reflex interneurons is not known, as are the source of modulatory inputs to these pathways. The aim of this article is to (a) provide an overview of MEM reflex anatomy and physiology, (b) present new data on MEM reflex anatomy and physiology from our laboratory and others, and (c) describe the clinical implications of our research.

A morphologic study of Fluorogold labeled tensor tympani motoneurons in mice.

The tensor tympani is one of two middle ear muscles that regulates the transmission of sound through the middle ear. Contraction of the tensor tympani in response to both auditory and non-auditory stimulation is mediated by the tensor tympani motoneurons (TTMNs). There are interesting differences among species in the acoustic thresholds for contraction of the middle ear muscles, which may be a reflection of underlying anatomical differences such as the number of TTMNs. However anatomical data for mice are lacking, even though the mouse is becoming the most common animal model for auditory and neuroscience research. We investigated the number and morphology of TTMNs in mice using Fluorogold, a retrograde neuronal tracer. After injections of Fluorogold into the tensor tympani muscle, a column of labeled TTMNs was identified ventro-lateral to the ipsilateral trigeminal nucleus. The labeled TTMNs were classified according to their morphological characteristics into three subtypes: "octopus-like", "fusiform" and "stellate", suggesting underlying differences in function. All three subtypes formed sparsely branched and radiating dendrites, some longer than 600 microm. Dendrites were longest and most numerous in the dorso-medial direction. In 18 cases, the mean number of mouse TTMNs was 51; the largest numbers were 70, 74 and 90 (n=3 injections). The mean size of mouse TTMNs was 13.0 microm (minor axis) and 23.5 microm (major axis). Compared with studies of TTMNs in larger species (cats and rats), mouse TTMNs are both fewer in number and smaller in size.

Neurochemistry of identified motoneurons of the tensor tympani muscle in rat middle ear.

The objective of the present study was to identify efferent and afferent transmitters of motoneurons of the tensor tympani muscle (MoTTM) to gain more insight into the neuronal regulation of the muscle. To identify MoTTM, we injected the fluorescent neuronal tracer Fluoro-Gold (FG) into the muscle after preparation of the middle ear in adult rats. Upon terminal uptake and retrograde neuronal transport, we observed FG in neurons located lateral and ventrolateral to the motor trigeminal nucleus ipsilateral to the injection site. Immunohistochemical studies of these motoneurons showed that apparently all contained choline acetyltransferase, demonstrating their motoneuronal character. Different portions of these cell bodies were immunoreactive to bombesin (33%), cholecystokinin (37%), endorphin (100%), leu-enkephalin (25%) or neuronal nitric oxide synthase (32%). MoTTM containing calcitonin gene-related peptide, tyrosine hydroxylase, substance P, neuropeptide Y or serotonin were not found. While calcitonin gene-related peptide was not detected in the region under study, nerve fibers immunoreactive to tyrosine hydroxylase, substance P, neuropeptide Y or serotonin were observed in close spatial relationship to MoTTM, suggesting that these neurons are under aminergic and neuropeptidergic influence. Our results demonstrating the neurochemistry of motoneuron input and output of the rat tensor tympany muscle may prove useful also for the general understanding of motoneuron function and regulation.

Neurons in the cochlear nuclei controlling the tensor tympani muscle in the rat: a study using pseudorabies virus.

The middle ear muscle reflex has been implicated in modulation of auditory input and protection of the inner ear from acoustic trauma. However, the identification of neurons in the cochlear nuclei participating in this reflex has not been fully elucidated. In the present study, we injected the retrograde transynaptic tracer pseudorabies virus into single tensor tympani (TT) muscles, and identified transynaptically labeled cochlear nucleus neurons at multiple survival times. Motoneurons controlling TT were located ventral to the ipsilateral motor trigeminal nucleus and extended rostrally towards the medial aspect of the lateral lemniscus. Transynaptically labeled neurons were observed bilaterally in the dorsal and dorso-medial parts of ventral cochlear nuclei as early as 48 h after virus injection, and had morphological features of radiate multipolar cells. After >or=69 h, labeled cells of different types were observed in all cochlear nuclei. At those times, labeling was also detected bilaterally in the medial nucleus of the trapezoid body and periolivary cell groups in the superior olivary complex. Based on the temporal course of viral replication, our data strongly suggest the presence of a direct projection of neurons from the ventral cochlear nuclei bilaterally to the TT motoneuron pool in rats. The influence of neurons in the cochlear nuclei upon TT activity through direct and indirect pathways may account for multifunctional roles of this muscle in auditory functions.

The extraocular motor nuclei: organization and functional neuroanatomy.

The organization of the motoneuron subgroups in the brainstem controlling each extraocular eye muscle is highly stable through the vertebrate species. The subgroups are topographically organized in the oculomotor nucleus (III) and are usually considered to form the final common pathway for eye muscle control. Eye muscles contain a unique type of slow non-twitch, fatigue-resistant muscle fiber, the multiply innervated muscle fibers (MIFs). The recent identification the MIF motoneurons shows that they too have topographic organization, but very different from the classical singly innervated muscle fiber (SIF) motoneurons. The MIF motoneurons lie around the periphery of the oculomotor nucleus (III), trochlear nucleus (IV), and abducens nucleus (VI), slightly separated from the SIF subgroups. The location of four different types of neurons in VI are described and illustrated: (1) SIF motoneurons, (2) MIF motoneurons, (3) internuclear neurons, and (4) the paramedian tract neurons which project to the flocculus. Afferents to the motoneurons arise from the vestibular nuclei, the oculomotor and abducens internuclear neurons, the mesencephalic and pontine burst neurons, the interstitial nucleus of Cajal, nucleus prepositus hypoglossi, the supraoculomotor area and the central mesencephalic reticular formation and the pretectum. The MIF and SIF motoneurons have different histochemical properties and different afferent inputs. The hypothesis that SIFs participate in moving the eye and MIFs determine the alignment seems possible but is not compatible with the concept of a final common pathway.

Is there a double innervation of the tensor tympani muscle in humans?

The middle ear muscles and their function have not yet been fully explored. The statement of Lawrence, for example, that the tensor tympani muscle of humans might have a dual innervation has never been proven or disproven. The question is of great interest; in our opinion, it represents one of the key questions in the putative afferent feedback loop of the middle ear muscles in humans. A light microscopic study was performed on 16 tensor tympani muscles taken from 11 cadavers. Six muscles were taken out in toto and stained according to the modified method of Sihler. The remaining 10 muscles were dehydrated and embedded in paraffin. In 5 of these muscles, complete transverse serial sections were made on a microtome at 7 microm and alternately stained by silver impregnation, S-100 protein immunohistochemistry, and ferric oxide. In the remaining 5 muscles, complete longitudinal serial sections were made at 10 microm. These sections were alternately stained by the methods of Cason and Maskar. Neither the surgical microscopic investigation nor the light microscopic investigation revealed any innervation to the human tensor tympani muscle other than the one arising from the mandibular branch of the trigeminal nerve. Our findings, apart from the fact that they clearly refute an unproven hypothesis, might represent another small step toward understanding the innervation of the tensor tympani muscle.

Acoustic tensor tympani response and vestibular-evoked myogenic potential.

OBJECTIVE: To investigate the acoustic response properties and the vestibular-evoked myogenic potential (VEMP) in various lesions. STUDY DESIGN: Retrospective study of the clinical records of patients. METHODS: Neurotological tests including acoustic response and VEMP were performed and analyzed in 62 patients with facial palsy, otosclerosis, ossicular chain interruption, sensorineural hearing loss, or acoustic tumor. RESULTS: Inverted acoustic responses were observed in 25 of 38 (65.8%) patients with facial palsy, in 5 of 6 (83.3%) patients with acoustic tumor, and in all patients with otosclerosis, ossicular chain interruption, or sensorineural hearing loss. These inverted responses were obtained only when ipsilateral stimulation was used. The thresholds of the inverted responses were statistically significantly higher than those of the normal response. CONCLUSIONS: The vibration of the eardrum is thought to stimulate the ipsilateral trigeminal nerve, leading to contraction of the tensor tympani muscle. The stapedius response had an inhibitory effect on the inverted response. Vibration of the stapes footplate (which requires a normal middle ear conduction system) is necessary to induce the VEMP, whereas the functioning of the facial and cochlear nerves is independent of the VEMP response.

["Irritative" pathology of the external auditory canal and decreased sound sensation]. / Patología "irritativa" del conducto auditivo externo y disminución de la sensación sonora.

We have not found in the literature an explanation for the intermittent and transient decreased lesser hearing sensation in patients with dysesthesia of the external auditory canal (EAC). In this paper we offer a possible explanation for it. Our hypothesis is that the stimulation of the sensory fibers of the trigeminal and facial nerves in the EAC is able to increase the stiffness of the ossicular chain by means of a reflex stimulation of malleus and stapes muscles. This intermittent and transient increase of the ossicular stiffness could explain the intermittent and transient decrease of hearing sensation in these patients.

Serotoninergic innervation of stapedial and tensor tympani motoneurons.

Retrograde tracing and neurotransmitter immunohistochemistry were combined to determine whether serotonin neurons innervated stapedial and tensor tympani motoneurons. With high-power light microscopy, putative axo-somatic and axo-dendritic contacts were observed between serotonin-positive endings and both stapedial and tensor tympani motoneurons, indicating that serotonin neurons terminate on brainstem motoneurons innervating the middle-ear muscles. With this connection, the serotonin system may directly modulate middle-ear muscle activity.

Cholera toxin B-HRP and wheat germ agglutinin-HRP tracing of tensor tympani muscle motoneurons and processes in rabbits.

The brain stem position, organization and number of motoneurons innervating the rabbit tensor tympani muscle (TTM) were determined by retrograde axonal transport of cholera toxin B/horseradish peroxidase conjugate (CTB-HRP) and wheat germ agglutinin HRP conjugate (WGA-HRP) tracers. The synaptic input to the TTM motoneurons was examined with WGA-HRP. The results show the motoneurons of the TTM to be localized in a cluster ventro-lateral to the outer margin of the ipsilateral trigeminal motor nucleus (VMN) and dorso-lateral to the superior olive. The number of labeled cells was greater in the combined CTB-HRP/WGA-HRP injected cases. The TTM motoneurons were triangular and elongated in shape and smaller than those of the VMN. An extensive network of dendritic branches was present ventro-laterally in the vicinity of the superior olive. Similar, but less extensive collections of dendritic processes were observed to course dorso-medially, rostrally and caudally. Axons were observed to project first dorsally or laterally, towards the trigeminal motor root, then after a sharp turn coursed ventrally within the trigeminal motor root (VMR). Transneuronal transport of the WGA-HRP was not accomplished in any preparation, suggesting among other things, system or species differences in the effectiveness of the WGA-HRP conjugate as a transynaptic tracer. It is concluded that the TTM acoustic reflex in rabbits and other mammals, its threshold, prolonged contraction capacity, and its influence on middle ear sound transmission may be related to its demonstrated extensive synaptic field in the reflex chain, particularly in the area of the superior olive, while its many other physiological functions may be made possible by the number, location, and multi-dimensional orientation of its motoneurons and dendrites.

The innervation of the middle ear muscles of the rat.

The innervation of the tensor tympani muscle and the stapedius muscle in the rat was studied. This was done by acetylcholinesterase in toto staining of the tympanic bullae and of muscles dissected separately, acetylcholinesterase staining of serial cross-sections of the muscles, silver impregnation of serial sections of complete tympanic bullae, serial semithin sections stained according to Laczko & Levai and electron microscopy of both muscles. The gross innervation of the muscles and the relation to other nerves in the bulla are described. It is shown that both muscles are innervated by very thin nerve fibres which form a well-organised elaborate network in the muscles, with very short branches that connect with motor endplates. Electron microscopically there are indications that the endplates in the stapedius muscle seem to enable faster activation of the muscle fibres than those of tensor tympani muscle. No morphological evidence for any sensory innervation of the muscles could be detected in the muscles themselves, in the connective tissue related to the muscles, or in the contents of the bulla tympanica. It is postulated that the afferent input of the acoustic middle ear muscle reflex is sound alone and that sensory information from the muscles themselves or from other structures in the tympanic bulla do not contribute to the reflex.

Electrophysiological and HRP studies of the direct afferent inputs from the cochlear nuclei to the tensor tympani muscle motoneurons in the cat.

The direct fiber connections from the cochlear nuclei to the tensor tympani muscle (TTM) motoneurons were investigated by means of electrophysiological and horseradish peroxidase (HRP) methods, using cats. When HRP was injected into motoneuron region of the TTM, HRP-labelled cells were found bilaterally in the dorsal cochlear nucleus (DCN) and ventral cochlear nucleus (VCN). When the electrical stimulus was applied to the cochlear nucleus, the TTM motoneurons fired spikes monosynaptically with s short latency. These histological and electrophysiological results indicate the existence of direct fiber connections from the bilateral cochlear nuclei to the TTM motoneurons.

[Reaction of the tympanic tensor muscle--elicited by nasally applied trigeminal stimulants]. / Reaktion des M. tensor tympani--ausgelöst durch nasal applizierte Trigeminusreizstoffe.

Computerised evaluation of tensor muscle reaction was carried out by using a biosignal analysing unit triggered by nasal inhalation. The trigeminus nerve was stimulated by application of 3-molar acetylacetic acid into the nasal respiratory air, inducing a contraction of the tympanic muscle, followed by a change in impedance. This change in impedance of the tympanic membrane ossicle system was recorded and printed out on a display. In this manner evidence was obtained of a tensor muscle reaction induced by the third branch of the trigeminal nerve as efference, and demonstrated for the first time. This reflex arc had long been considered as being of negligible clinical importance before its stimulation and measurement had become possible. It is a generally accepted theory that the reflex arc of the m. tensor tympani is linked to the formatio reticularis which assesses the sensory afferences. For this reason, the reflex arc habituates rapidly, and continuous stimulation is no longer possible.

Tensor tympani reflex pathways studied with retrograde horseradish peroxidase and transneuronal viral tracing techniques.

The neural pathway involved in activation of the tensor tympani (TT) muscle was studied in the rat using retrograde HRP and transneuronal viral tracing techniques. The pool of TT motoneurons labeled with HRP was located ipsilaterally under the anterior third of the trigeminal motor nucleus and extended rostrally towards the lateral lemniscus. The origin of the inputs to these motoneurons was then determined using transneuronal viral transport: presumably transneuronally infected neurons appeared bilaterally in the vicinity of the superior olivary complex, mainly in between the two nuclei of the trapezoid body. The present data are consistent with previous conclusions based on lesion experiments that the TT reflex loop is made up of a chain of 4 neurons.