Spiral Ganglion Nerve Research Articles

Autophagy-Mediated Synaptic Refinement and Auditory Neural Pruning Contribute to Ribbon Synaptic Maturity in the Developing Cochlea.

In rodents, massive initial synapses are formed in the auditory peripheral nervous system at the early postnatal stage, and one of the major phenomena is that the number of afferent synapses in the cochlea is significantly reduced in the duration of development. This raises the hypothesis that the number of cochlear ribbon synapses are dramatically changed with hearing development and maturation. In this study, several tracers identifying activities of autophagy were applied to estimate the level of autophagy activity in the process of ribbon synapse development in mice; further, changes in the synaptic number and spiral ganglion nerve (SGN) fibers were quantitatively measured. We found robust expression of LC3B and lysosomal-associated membrane protein 1 as well as LysoTracker in or near inner hair cells and cochlear ribbon synapses in the early stage of postnatal development. Moreover, we found a significant loss in the intensity of SGN fibers at ribbon synaptic development and hearing onset. Thus, this study demonstrates that ribbon synaptic refinement and SGN fibers pruning are closely associated with the morphological and functional maturation of ribbon synapses and that synaptic refinement and SGN fiber pruning are regulated by the robust activities of autophagy in the earlier stages of auditory development.

Transplantation and Tracking of the Human Umbilical Cord Mesenchymal Stem Cell Labeled with Superparamagnetic Iron Oxide in Deaf Pigs.

The purpose of this study was to establish a safe and effective approach to label the human umbilical cord mesenchymal stem cells (UC-MSCs) derived from the Wharton's Jelly with superparamagnetic iron oxide (SPIO) nanoparticles as a cell tracer. The cytotoxicity of the SPIO was screened in vitro by cytochemical experiments. The results showed the new infection protocol of SPIO-Lip2000 mixture had high efficiency and the optimal labeling concentration was a 50 μg/ml SPIO suspension. Transmission electron microscope (TEM) confirmed the distribution of the intracellular SPIO. We transplanted the labeled UC-MSCs into the sensorineural hearing loss (SNHL) minipigs at 1 week after noise exposure. Auditory brainstem response results demonstrated the transplantation of UC-MSCs was an efficient therapy for SNHL. The positive sediments in cochlear blood vessels, the bony wall of scala tympani, and spiral ganglion nerve fibers were found in the stem cell recipients' cochlea. We did not detect iron elements in the inner/outer hair cells' stereocilia, cuticular plate, or pillar cells from the basal to apex turns of the stem cell recipients' cochlea. In addition, TEM found SPIO in the medulla oblongata and the cerebrum in the SNHL minipigs after stem cell transplantation. In conclusion, we established a safe and effective approach to labeled human UC-MSCs derived from Wharton's Jelly by using SPIO nanoparticles as a cell tracer in vitro and in vivo. This protocol showed a wide promising application in stem cell therapy and tracing in vivo for experiments with large mammals. Anat Rec, 303:494-505, 2020. © 2019 American Association for Anatomy.

Spontaneous and Partial Repair of Ribbon Synapse in Cochlear Inner Hair Cells After Ototoxic Withdrawal.

Ototoxicity is one of the major causes of sensorineural deafness. However, it remains unclear whether sensorineural deafness is reversible after ototoxic withdrawal. Here, we report that the ribbon synapses between the inner hair cells (IHCs) and spiral ganglion nerve (SGN) fibers can be restored after ototoxic trauma. This corresponds with hearing restoration after ototoxic withdrawal. In this study, adult mice were injected daily with a low dose of gentamicin for 14 consecutive days. Immunostaining for RIBEYE/CtBP2 was used to estimate the number and size of synaptic ribbons in the cochlea. Hearing thresholds were assessed using auditory brainstem responses. Auditory temporal processing between IHCs and SGNs was evaluated by compound action potentials. We found automatic hearing restoration after ototoxicity withdrawal, which corresponded to the number and size recovery of synaptic ribbons, although both hearing and synaptic recovery were not complete. Thus, our study indicates that sensorineural deafness in mice can be reversible after ototoxic withdrawal due to an intrinsic repair of ribbon synapse in the cochlea.

Loss of central auditory processing in a mouse model of Canavan disease.

Canavan Disease (CD) is a leukodystrophy caused by homozygous null mutations in the gene encoding aspartoacylase (ASPA). ASPA-deficiency is characterized by severe psychomotor retardation, and excessive levels of the ASPA substrate N-acetylaspartate (NAA). ASPA is an oligodendrocyte marker and it is believed that CD has a central etiology. However, ASPA is also expressed by Schwann cells and ASPA-deficiency in the periphery might therefore contribute to the complex CD pathology. In this study, we assessed peripheral and central auditory function in the AspalacZ/lacZ rodent model of CD using auditory brainstem response (ABR). Increased ABR thresholds and the virtual loss of waveform peaks 4 and 5 from AspalacZ/lacZ mice, indicated altered central auditory processing in mutant mice compared with Aspawt/wt controls and altered central auditory processing. Analysis of ABR latencies recorded from AspalacZ/lacZ mice revealed that the speed of nerve conduction was unchanged in the peripheral part of the auditory pathway, and impaired in the CNS. Histological analyses confirmed that ASPA was expressed in oligodendrocytes and Schwann cells of the auditory system. In keeping with our physiological results, the cellular organization of the cochlea, including the organ of Corti, was preserved and the spiral ganglion nerve fibres were normal in ASPA-deficient mice. In contrast, we detected substantial hypomyelination in the central auditory system of AspalacZ/lacZ mice. In summary, our data suggest that the lack of ASPA in the CNS is responsible for the observed hearing deficits, while ASPA-deficiency in the cochlear nerve fibres is tolerated both morphologically and functionally.

The Role of Mobile Calcium Buffers in Synaptic Transmission at the Inner Hair Cell Ribbon Synapse

Temporally precise sound encoding at the inner hair cell (IHC) ribbon synapse is tightly regulated by calcium. The mobile calcium buffers calbindin, parvalbumin alpha and calretinin might contribute to shaping the presynaptic Ca2+ signals. We investigated the function of these calcium binding proteins in IHC synaptic transmission by examining the auditory phenotype of double and triple buffer knockout mice. Our results show that buffer deficiency does not significantly alter hearing thresholds; however, we observed a slight increase in peak and steady-state sound-driven spike rates of spiral ganglion nerve fibers in knockout mice. The presynaptic function of IHCs was first studied by perforated patch-clamp recordings of Ca2+ currents and exocytic membrane capacitance increments. The absence of mobile calcium buffering proteins augmented sustained exocytosis in IHCs while leaving the amplitude and kinetics of exocytosis of the readily-releasable pool unchanged. Further, Ca2+-dependent inactivation of calcium currents was stronger in IHCs of triple buffer knockout mice. In ruptured patch experiments we then tried to restore the calcium buffer capacity by adding exogeneous buffers. We estimated the concentration of endogenous buffers in IHCs to be equivalent to 0.5-1 mM BAPTA, which agrees well with previous estimates obtained by quantitative immunogold electron microscopy (Hackney et al., 2005). Our results demonstrate that calbindin, parvalbumin alpha and calretinin are involved in the regulation of synaptic transmission at the IHC ribbon synapse; however they do not seem to be essential for hearing, at least not during transient sound stimulation.

From High-Tech to Biotech Using Stem Cell and Gene Therapy to Treat Hearing Loss

From High-Tech to Biotech Using Stem Cell and Gene Therapy to Treat Hearing Loss

Expression of myelin basic protein in the human auditory nerve—An immunohistochemical and comparative study

The aim of this study is to analyse the expression and distribution of myelin basic protein (MBP or Myelin A1 protein) in the human spiral ganglion and auditory nerve. Cryostat sections were made from freshly fixed human cochlear specimens removed at surgery in patients with life-threatening petro-clival meningiomas compressing the brain stem. The sections were subjected to immunohistochemistry using antibodies against MBP, S-100 and Tubulin. The immunoreaction was documented using laser confocal microscopy. Type I spiral ganglion nerve somata (SGN) were surrounded by so-called "satellite glial cells" (SGCs) that lacked expression of MBP consistent with earlier light and electron microscopic findings indicating that these cells are non-myelinating. S-100 labeling showed that the SGCs form a continuous network in the apical region. The pattern of myelination in human spiral ganglion is different from that in other species' spiral ganglion. The striking differences in myelin outline should be investigated further in combination with its influence on signal coding and preservation properties in man.

Modeling the electrode–neuron interface of cochlear implants: Effects of neural survival, electrode placement, and the partial tripolar configuration

The partial tripolar electrode configuration is a relatively novel stimulation strategy that can generate more spatially focused electric fields than the commonly used monopolar configuration. Focused stimulation strategies should improve spectral resolution in cochlear implant users, but may also be more sensitive to local irregularities in the electrode–neuron interface. In this study, we develop a practical computer model of cochlear implant stimulation that can simulate neural activation in a simplified cochlear geometry and we relate the resulting patterns of neural activity to basic psychophysical measures. We examine how two types of local irregularities in the electrode–neuron interface, variations in spiral ganglion nerve density and electrode position within the scala tympani, affect the simulated neural activation patterns and how these patterns change with electrode configuration. The model shows that higher partial tripolar fractions activate more spatially restricted populations of neurons at all current levels and require higher current levels to excite a given number of neurons. We find that threshold levels are more sensitive at high partial tripolar fractions to both types of irregularities, but these effects are not independent. In particular, at close electrode–neuron distances, activation is typically more spatially localized which leads to a greater influence of neural dead regions.

Spontaneous association of glial cells with regrowing neurites in mixed cultures of dissociated spiral ganglia

Evidence from developmental and regeneration studies of the cochlea and other tissues gives reason to hypothesize a role for nonneural cells in the growth and regeneration of cochlear spiral ganglion nerve fibers. We examined the spontaneous associations of regrowing neurites and nonneural cells in mixed cultures of dissociated newborn mouse spiral ganglia. After 7 days in vitro, nonneural cells formed a confluent layer in the culture well. Regrowing neurites grew atop this layer, forming non-uniform patterns that were similar to those formed by endogenously expressed laminin-1, entactin and integrin β4, but not fibronectin or tenascin. In cultures grown for 42 h and maintained in three different growth media, all regrowing neurites were preferentially associated with spindle-shaped nonneural cells. The spindle-shaped cells incorporated bromodeoxyuridine in culture and were immunoreactive for the proteins S100, laminin-1, laminin-2, SRY-related high-mobility-group box 10 transcription factor (Sox10), neurotrophin receptor (P75) and connexin29 but negative for fibronectin and glial fibrillary acidic protein. These cells existed in the culture within a much larger, general population of fibronectin positive cells. Immunolabeling of fixed cochleas from neonatal mice localized Sox10, P75 and connexin29, to peripheral nerve bundles. The observed expressions of protein markers and the bipolar, spindle shape of the neurite-associated cells indicate that they are derived in vitro from the original Schwann or satellite cells in the ganglion or spiral lamina. The spontaneous and preferential association of neurites in culture with mitotic Schwann cells highlights the potential contribution neurite–Schwann cell interactions may have in promoting the growth and regrowth of damaged spiral ganglion neurons in the cochlea.

Pathologic changes in the inner ear of audiogenic seizure-susceptible mice treated with 6-aminonicotinamide

The inner ears were studied histologically to determine a possible morphological basis for our earlier finding of the abolition of sound-induced seizures in inbred audiogenic seizure-susceptible mice treated with 6-aminonicotinamide (6-AN). A single intraperitoneal injection of 20 mg/kg of 6-AN resulted in elimination of sound-induced seizures and produced severe degenerative lesions in the cochlear duct of the seizure-susceptible mice. Similar cochlear lesions were produced in a control mouse strain (CF#1). The earliest lesions, first sought and clearly evident at 24 hours after the injection, were localized in the spiral ligament, the epithelium of the external spiral sulcus, and the lateral portions of the papilla acoustica. The acute degeneration was at its highest 3–5 days after the injection, when the medial portions of the papilla were also involved. The damage to the spiral organ (Corti) seen at that time is incompatible with a normal sensory input. The degree of damage is sufficient to explain the loss of, or marked restriction in, response to auditory stimuli, and the abolition of audiogenic seizures. In all animals treated with 6-AN, the lower basal turn and the upper second turn were more extensively damaged than the rest of the cochlea. However, in the longest surviving 6-AN-treated mice not even a remnant of the spiral organ could be detected. The spiral ganglion nerve cells were not damaged in the acute stage of 6-AN intoxication. The atrophy of the spiral ganglion which appeared in the late stages was, like the degeneration of the spiral organ, most severe in the lower basal and upper second turns. The changes in the spiral ganglion were interpreted as being secondary to the acute destruction of the spiral organ.

CYTOLOGICAL AND ELECTRON MICROSCOPIC STUDIES ON THE SPIRAL GANGLION CELLS OF THE ADULT GUINEA PIGS AND RABBITS

Although several kinds of electron-microscopic studies have so far been made on various ganglion cells, it seems that concerning spiral ganglion cells there has been only one brief report of YAMAMOTO et al. (1963). The present study was made on the spiral ganglion cells of matured guinea-pig which are used most frequently for our experimental researches. At the same time rabbit's spiral ganglion cells were observed, too.Material and methods: Fifteen healthy matured guinea-pigs and four healthy rabbits were etherized. Modioli previously taken after destroying osseus labyrinth were fixed in LEVI's solution, REGAUD's solution and formol-alcohol. For the study of the GOLGI apparatus, KOLATCHEV's method was adopted. Decalcification was performed in the 5% trichloracetic acid about 5 days and 3 to 4μ paraffined serial sections were made after dehydration. For the electron-microscopy, modioli were fixed in 1% osmium tetroxide adjusted to pH 7.4 with phosphate buffer, dehydrated through a series of alcohol, and then embedded in styrene and n-butyl methacrylate (1:1), epon and new epoxy resine‘Epok 533’recommended by KUSHIDA. Thin sections were cut on a PORTER-BLUM microtome with glass knives, stained with uranyl acetate, and examined in a HITACHI's HU-10 electron microscope and a NIPPON DENSHI's JEMT-6 electron microscope at magnification of 1, 000-10, 000.1. The spiral ganglion consists of comparatively light bipolar nerve cells which are almost of the same size. The nucleus is round or oval in shape and is sometimes located to one side in the cells. It contains one or several nucleoli which are often found closely attached to the nuclear membrane. Occasionally a small number of comparatively dark cells are recognized. They have many small vacuoles, their cytoplasm has got dark stained and their nucleus has often become atrophic. This fact, however, is considered to be an indication of one stage of the above-mentioned nerve cell function.A great number of filamentous and bacilliform mitochondria are recognized scatteringly and a comparatively simply formed GOLGI apparatus is found around nucleus. NISSL substances are often observed diffusely in the cytoplasm. In spiral ganglion nerve cell, somewhat light yellowed gross pigment granules are often recognized. They are of more or less angular pseudomorph and not homogeneous, and there are some of their portions which are blackened by the fixative which contains OsO4. Furthermore occasions not frequently arise when several pigment granules are placed in the form of‘rosette’on one side in the cell. In fact, they are well stained with iron-hematoxylin, slightly stained with azocarmine and positive to PAS reaction. Besides this gross pigment granules, many small pigment granules which are positive to PAS reaction are found plentfully in the cytoplasm. It is often difficult to distinguish themselves from mitochondria as they are so well stained to the iron-hematoxylin.The spiral ganglion cell is called‘markhaltige Ganglienzellen’(SCHARF 1958) and its cell body is covered with myelin sheath. In many cases, only one satellite cell envelops around a nerve cell body in a section.2. Electron microscopic study was made on the above-mentioned nerve cell and the following results were obtained.a) No remarkable density difference is observed between the cytoplasm and the nucleus of the spiral ganglion cell, which is different from the spinal ganglion cell (KUROSUMI and AKIYAMA 1958). Nuclear membrane pores are confirmed on the nuclear membrane. Very often the outer membrane of the nucleus swells into cytoplasm. The nucleolus chiefly consists of nucleolonema and often finds itself close to the nuclear membrane. Besides, chromatin aggregates are seen in the nucleus.GOLGI apparatus consists of vesicles, vacuoles and lamellae, and quite often vacuoles and lamellae grow much larger.