Biomedical subjects

M Eybalin

Publications and source records attributed to M Eybalin.

At least 19 recordsLinked to original sources

PubMed2006

Coxsackie adenovirus receptor and alpha nu beta3/alpha nu beta5 integrins in adenovirus gene transfer of rat cochlea.

This study was designed to determine whether Coxsackie adenovirus receptor (CAR) and alpha nu beta3/alpha nu beta5 integrin co-receptors are involved in adenovirus gene transfer in the rat cochlea. We find that CAR and integrin co-receptors are expressed in every cell subtype transduced by the adenoviral vector Ad5 DeltaE1-E3/cytomegalovirus/green fluorescent protein (GFP) on cochlear slices in vitro. The spiral ganglion neurons, which do not express CAR, were not transduced by the virus. Blocking these receptors by monoclonal antibodies decreased transgene expression, whereas disrupting tight junctions with ethylenediaminetetraacetic acid led to an increased transgene expression. However, sensory hair cells and strial cells also expressing CAR and alpha nu integrins were not transduced by the vector. GFP expression was also studied in vivo. Perilymphatic perfusion of adenovirus in vivo did not affect hearing and only cells lining the perilymphatic spaces were transduced. Endolymphatic perfusion resulted in low-frequency hearing loss and although some cells of the organ of Corti were efficiently transduced, the sensory and the strial cells were not. Transduced sensory and strial cells were occasionally observed in cochleas after single shot of adenovirus. Pretreatment with anti-CAR and anti-alpha nu antibodies decreases GFP expression in vivo, suggesting that the CAR/alpha nu integrin pathway is involved in adenovirus transduction in the cochlea.

Action Potentials↗

PubMed2002

Riluzole rescues cochlear sensory cells from acoustic trauma in the guinea-pig.

Acoustic trauma is the major cause of hearing loss in industrialised nations. We show in guinea-pigs that sound exposure (6 kHz, 120 dB sound pressure level for 30 min) leads to sensory cell death and subsequent permanent hearing loss. Ultrastructural analysis reveals that degeneration of the noise-damaged hair cells involved different mechanisms, including typical apoptosis, autolysis and, to a lesser extent, necrosis. Whatever the mechanisms, a common feature of noise damage to hair cells was mitochondrial alteration. Riluzole (2-amino-6-trifluoromethoxy benzothiazole) is a neuroprotective agent that prevents apoptosis- and necrosis-induced cell death. Perfusion of riluzole into the cochlea via an osmotic minipump prevents mitochondrial damage and subsequent translocation of cytochrome c, DNA fragmentation, and hair cell degeneration. This was confirmed by functional tests showing a clear dose-dependent reduction (ED(50)=16.8 microM) of permanent hearing loss and complete protection at 100 microM. Although less efficient than intracochlear perfusion, intraperitoneal injection of riluzole rescues the cochlea within a therapeutic window of 24 h after acoustic trauma.These results show that riluzole is able to prevent and rescue the cochlea from acoustic trauma. It may thus be an interesting molecule for the treatment of inner ear injuries.

Acoustic Stimulation↗

PubMed2001

Dopamine inhibition of auditory nerve activity in the adult mammalian cochlea.

Efferent feedback systems provide a means for modulating the input to the central nervous system. The lateral olivocochlear efferents modulate auditory nerve activity via synapses with afferent dendrites below sensory inner hair cells. We examined the effects of dopamine, one of the lateral olivocochlear neurotransmitters, by recording compound and single unit activity from the auditory nerve in adult guinea pigs. Intracochlear application of dopamine reduced the compound action potential (CAP) of the auditory nerve, increased the thresholds and decreased the spontaneous and driven discharge rates of the single unit fibres without changing their frequency-tuning properties. Surprisingly, dopamine antagonists SCH-23390 and eticlopride decreased CAP amplitude as did dopamine. In some units, both SCH-23390 and eticlopride increased the basal activity of auditory nerve fibres leading to an improvement of threshold sensitivity and a decrease of the maximum driven discharge rates to sound. In other units, the increase in firing rate was immediately followed by a marked reduction to values below predrug rates. Because CAP reflects the summed activity of auditory nerve fibres discharging in synchrony, both the decrease in sound-driven discharge rate and the postexcitatory reduction account for the reduction in CAP. Ultrastructural examination of the cochleas perfused with eticlopride showed that some of the afferent dendrites were swollen, suggesting that the marked reduction in firing rate may reflect early signs of excitotoxicity. Results suggest that dopamine may exert a tonic inhibition of the auditory nerve activity. Removal of this tonic inhibition results in the development of early signs of excitotoxicity.

Acoustic Stimulation↗

PubMed1999

Glutamate receptor phenotypes in the auditory brainstem and mid-brain of the developing rat.

Glutamate receptors mediate most excitatory synaptic transmission in the adult vertebrate brain, but their activation in developing neurons also influences developmental processes. However, little is known about the developmental regulation of the subunits composing these receptors. Here we have studied age-dependent changes in the expression of alpha-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) and N-methyl-D-aspartate (NMDA) receptor subunits in the cochlear nucleus complex (CN), the superior olivary complex (SOC), the nuclei of the lateral lemniscus, and the inferior colliculus of the developing rat. In the lateral superior olive, the medial nucleus of the trapezoid body, and the ventral nucleus of the lateral lemniscus, the distribution of AMPA receptor subunits changed drastically with age. While GluR1 and GluR2 subunits were highly expressed in the first 2 postnatal weeks, GluR4 staining was detectable only thereafter. GluR1 and GluR2 immunoreactivities rapidly decreased during the third postnatal week, with the GluR1 subunits disappearing from most neurons. In contrast, the adult pattern of the distribution of AMPA receptor subunits emerged gradually in most of the other auditory nuclei. Thus, progressive as well as regressive events characterized AMPA receptor development in some nuclei, while a monotonically maturation was seen in other regions. In contrast, the staining patterns of NMDA receptor subunits remained stable or only decreased during the same period. Although our data are not consistent with a generalized pattern of AMPA receptor development, the abundance of GluR1 subunits is a distinctive feature of early AMPA receptors. As similar AMPA receptors are present during plasticity periods throughout the brain, neurons undergoing synaptic and structural remodelling might have a particular need for these receptors.

Age Factors↗

PubMed1997

Protein kinase C may be involved in synaptic repair of auditory neuron dendrites after AMPA injury in the cochlea.

A suitable model of sudden deafness occurring after acoustic trauma or ischemia, is obtained in guinea pigs by an acute intracochlear perfusion of 200 microM alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), a glutamate analog. By overloading the AMPA/kainate receptors, located post-synaptically to inner hair cells (IHCs), it induces a massive swelling of primary auditory neuron dendrites, which disconnects the IHCs. This synaptic uncoupling and the resulting hearing loss are followed by a progressive regrowth of dendrites, which make new synapses with IHCs, leading to a functional recovery of auditory responses that is completed after 5 days. Knowing the role of protein kinase C in neuroplastic events, we studied the expression of its isoforms alpha,beta(I,II) and gamma, respectively pre- and post-synaptic, in auditory neurons at various times after AMPA administration. In untreated cochleas, we observed an expression of PKC alpha,beta(I,II) and gamma in cell bodies of primary auditory neurons. After the intracochlear administration of AMPA, both isozymes were transiently overexpressed, with a peak at 3-6 h, followed by a decrease after about 24 h. At this point in time immuno-electron microscopy revealed some regrowing dendrites immunoreactive for PKCgamma. Five days after AMPA, when the auditory responses were restored, PKCgamma levels were still elevated in ganglion cell bodies.

Animals↗

PubMed1997

Temporary sensory deprivation changes calcium-binding proteins levels in the auditory brainstem.

Auditory brainstem neurons probably depend on afferent input to maintain calcium homeostasis within a narrow range. These neurons are endowed with high concentrations of the calcium-binding proteins parvalbumin, calretinin, and calbindin D28k that are presumed to buffer cytosolic calcium transients. To determine the effects of functional deafferentation on these proteins in the auditory brainstem of adult guinea pigs, we have manipulated the sensory input with an intracochlear perfusion of the glutamate agonist alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), which is known to transiently disconnect inner hair cells and primary auditory dendrites. Semiquantitative measures of immunostaining intensities showed rapid and reversible changes in calcium-binding protein levels. By 24 hours after AMPA treatment, calretinin immunostaining was reduced in deafferented neurons of the cochlear nuclei and their axons in the superior olivary nuclei. In contrast, calbindin D28k immunoreactivity levels by this time were higher in deafferented neurons of the medial nucleus of the trapezoid body and their axons in the lateral superior olivary nucleus (LSO). Parvalbumin immunostaining was also generally increased in deafferented neurons, but changes were less evident and more complex. The changes in all three immunoreactivities disappeared with the progressive restoration of afferent input. Normal levels were reestablished by 5 days after AMPA treatment, when afferent activity had almost completely recovered. These results show that calcium-binding protein immunostaining in auditory neurons is functionally responsive to afferent activity. The increased buffering capacity in deafferented neurons as shown by the rises in parvalbumin and calbindin D28k immunostaining may be part of mechanisms promoting neuronal survival after loss of sensory input. This input, on the other hand, may be necessary for maintaining the high calretinin levels normally present in cochlear nucleus neurons.

Animals↗

PubMed1997

Choline acetyltransferase, glutamate decarboxylase, tyrosine hydroxylase, calcitonin gene-related peptide and opioid peptides coexist in lateral efferent neurons of rat and guinea-pig.

The lateral efferent (olivocochlear) innervation of the cochlea originates in the brainstem lateral superior olive. It is likely to use acetylcholine, gamma-aminobutyric acid, dopamine and various neuropeptides as neurotransmitters and/or neuromodulators. In order to determine the different coexistence patterns of these molecules in lateral efferent perikarya, we have used double and triple immunofluorescence co-localization techniques to colocalize choline acetyltransferase, glutamate decarboxylase, tyrosine hydroxylase, calcitonin gene-related peptide and enkephalins in single sections of the lateral superior olive. We also used a non-radioactive in situ hybridization technique onto serial sections of this nucleus to confirm the immunofluorescence co-localization data at the mRNA level. Whatever the pair or triplet of primary antibodies tested was, a high ratio of coexistence was observed in the immunofluorescence experiments. In triple co-localization experiments, 90-93% of the choline acetyltransferase-like immunoreactive neurons were also immunoreactive to the two other antigens investigated. The in situ hybridization co-localization data, based on the use of biotin-labelled oligoprobes, qualitatively confirmed these immunofluorescence data. In conclusion, it can be postulated that acetylcholine, gamma-aminobutyric acid, dopamine, calcitonin gene-related peptide, enkephalins and dynorphins (whose coexistence with choline acetyltransferase and enkephalins has been previously described immunocytochemically) coexist in lateral efferent neurons. Based on these results, it is tempting to propose the lateral efferent innervation as a useful model with which the functional implications of the coexistence of neurotransmitters/neuromodulators can be investigated in vivo.

Animals↗

PubMed1996

Glutamine synthetase and glutamate metabolism in the guinea pig cochlea.

Glutamate is thought to act as a neurotransmitter of the sensory hair cells of the organ of Corti. Glutamine synthetase could be involved in a type of glutamate-glutamine cycle in the cochlea which could clear glutamate off the synaptic cleft and replenish the hair cell glutamate neurotransmitter store. Using both light and electron microscopic immunocytochemistry to localize this enzyme in the guinea pig cochlea, we have observed immunoreactive satellite glial cells surrounding parvalbumin-immunoreactive primary auditory neurons in the spiral ganglion. Glutamine synthetase was also detected in Schwann cells of the osseous spiral lamina which form the myelin sheath of nerve fibers. On the contrary, no immunoreactivity could be observed in the cochlear nerve and in the organ of Corti, although this organ contains structures able to take up glutamate. Although they confirm earlier works involving glutamine synthetase in the conversion of L-[3H]glutamate taken up by glial cells, our results suggest that the cochlear glutamate-glutamine cycle is not primarily involved in the recycling and replenishment of hair cell neurotransmitter glutamate. Alternatively, it is proposed that glutamine synthetase functions to limit the perilymphatic glutamate concentrations.

Animals↗

PubMed1996

Distribution of calcium-binding protein immunoreactivities in the guinea pig auditory brainstem.

This study was intended to provide an overview of the distribution of calcium-binding proteins in the rodent auditory brainstem. We based our observations on immunohistochemical material obtained in the guinea pig, a species widely used in auditory research in which a mapping of calcium-binding proteins in the auditory brainstem is still missing. Differences in the amounts of these proteins throughout the auditory brainstem were further analyzed semiquantitatively. Parvalbumin was present in most neurons and their axon terminals throughout the ascending auditory brainstem. Nuclei that surround the main relay nuclei of the ascending auditory pathway lacked labeling. Calretinin staining was prominent in spherical and globular cells of the cochlear nucleus, in their axon terminals in the superior olivary complex, and in principal cells of the medial superior olive. Measures of optical densities showed that auditory neurons involved in sound localization had the highest calretinin labeling levels. Calbindin D-28k was present in cartwheel cells of the dorsal cochlear nucleus, in almost all neurons of the medial nucleus of the trapezoid body, and in globular cells in the ventral nucleus of the lateral lemniscus. The labeling patterns for calretinin and calbindin D-28k were non-overlapping throughout the auditory brainstem. This was also evident in the ventral nucleus of the lateral lemniscus where calbindin D-28k-immunoreactive terminals were found in the medial portion, while the calretinin-immunoreactive terminals were observed in the lateral portion. This study presents the first direct and comprehensive comparison of these three calcium-binding proteins in the auditory brainstem of a rodent. Each antibody yields a unique staining pattern that provides a basis for further defining neuronal populations. In addition, since their axons are also selectively stained, auditory nuclei can further be compartmentalized based on different terminal fields. These immunoreactivities have provided clues to the complex structure of the auditory brainstem.

Animals↗

PubMed1996

Pre- and postsynaptic M3 muscarinic receptor mRNAs in the rodent peripheral auditory system.

The medial and lateral efferent innervations originate from distinct parts of the superior olivary complex. Both use acetylcholine, respectively, to modulate the activity of outer hair cells (OHC), and spiral ganglion neurons (SGN) which are postsynaptic to the inner hair cells (IHC). Besides predominantly activating nicotinic receptors, acetylcholine recognizes muscarinic M3 receptors, whose the role(s) and cellular localization(s) are not yet firmly established. We used reverse transcription and polymerase chain reaction to amplify the M3 receptor cDNA in the rat and guinea pig organ of Corti and spiral ganglion. Then, we localized the M3 receptor mRNAs in cochleas and superior olivary complex of both species. The M3 receptor cDNA was amplified from samples of brain, organ of Corti and spiral ganglion. Indeed, its corresponding mRNA was localized in SGNs, OHCs and IHCs. However, in the apical turns, OHCs were often found unlabeled. In the superior olivary complex, M3 mRNAs were colocalized with choline acetyltransferase mRNAs in neurons of the lateral superior olive and ventral nucleus of the trapezoid body. These results suggest that the M3 receptor-induced inositol phosphate formation described in previous studies [21] takes place in both postsynaptic (SGNs, OHCs) and presynaptic components of efferent cochlear synapses, and in cells that are not contacted by efferents in the adult cochlea (IHCs).

Animals↗

PubMed1995

Expression of mGluR1 alpha mRNA receptor in rat and guinea pig cochlear neurons.

Glutamate or a parent substance is thought to be the afferent neurotransmitter in the auditory system. In situ hybridization showed that mGluR1 alpha mRNA was expressed by type I and type II spiral ganglion neurons in the cochlea. The glial cells surrounding the type I spiral ganglion neurons lacked such expression. The hybridization signal was low compared to that reported for non-NMDA receptors, suggesting that mGluR1 alpha receptors, as is the case for NMDA receptors, play a minor role in auditory transmission. The uniform expression of mGluR1 alpha mRNAs along the cochlear spiral suggests their co-expression in spiral ganglion neurons with NMDA and non-NMDA receptors and thus functional cooperation.

Animals↗

PubMed1995

Molecular cloning and expression of alpha parvalbumin in the guinea pig cochlea.

We have cloned and sequenced an alpha parvalbumin cDNA from the guinea pig cochlea. The deduced amino acid sequence shows greater identity with the rabbit sequence (86.3%) than with other mammalian sequences (< 82%). Using in situ hybridization and immunohistochemistry, alpha parvalbumin mRNA and protein were found in primary auditory neurons and inner hair cells, in agreement with RT-PCR data showing alpha parvalbumin mRNA expression in the spiral ganglion and the organ or Corti.

Amino Acid Sequence↗

PubMed1995

Synaptic connections and putative functions of the dopaminergic innervation of the guinea pig cochlea.

We report our findings in the guinea pig involving dopamine in postsynaptic regulation of the activity of glutamatergic inner hair cells (IHCs) and in protection of primary auditory neurons during transient ischemia. Seven days after intracochlear perfusion of 6-hydroxydopamine, no immunoreactivity to tyrosine hydroxylase (TH) was demonstrable within the organ of Corti. TH and aromatic amino acid decarboxylase were immunolocalized at an ultrastructural level within lateral olivocochlear varicosities synapsing with radial auditory dendrites postsynaptic to the IHCs. The D2 agonist piribedil induced a dose-dependent decrease in the amplitude of the compound action potential of the auditory nerve. Piribedil also prevented appearance of ischemia-induced swelling of the radial dendrites.

Animals↗

PubMed1995

Effects of a dopaminergic agonist in the guinea pig cochlea.

This study investigates the role of dopamine, a putative lateral efferent neurotransmitter/modulator, in cochlear physiology and physiopathology. Cochlear potentials were recorded in guinea pigs after intracochlear perfusion of increasing doses (0.1-1 mM) of piribedil, an agonist of the D2/D3 receptors. A dose-dependent reduction in the amplitude of auditory nerve compound action potential (CAP) was observed, predominantly at high-intensity tone-burst stimulations, and without significant effect on CAP threshold. There was no variation of cochlear microphonic and summating potential. When 1 mM piribedil was perfused into the cochlea during continuous 130 dB SPL pure tone exposure (6 kHz, 15 min), CAP threshold shifts were significantly less than in control animals with artificial perilymph-perfused cochleas. No dendritic damage was observed, although there was evident hair cell damage. Similarly, radial dendrites were clearly protected against ischemia-induced damage when 1 mM piribedil was applied prior to a 10-min ischemia. These results suggest that dopamine modulates the activity of radial afferent fibers via D2/D3 receptors. The protective effect of piribedil during acoustic trauma or ischemia suggests that this modulation corresponds to a prevention of excitotoxicity due to dysfunction of inner hair cell neurotransmission.

Acoustic Stimulation↗

PubMed1995

Synaptic regeneration and functional recovery after excitotoxic injury in the guinea pig cochlea.

Cochlear ischemia and acoustic trauma result in an immediate hearing loss accompanied by the complete disruption of the terminal dendrites of primary auditory neurons postsynaptic to the sensory inner hair cells (IHCs). This synaptic uncoupling, due to an acute glutamate (IHC neurotransmitter) excitotoxicity process, can be mimicked by glutamate agonists. Thus, we have followed over a 5-day period the responses of guinea-pig cochleas to a local application of 200 microM AMPA. This application immediately results in a destruction of all postsynaptic endings of the auditory nerve, resulting in a total loss of cochlear potentials. Twenty-four hours after this excitotoxic injury, the inner hair cells were contacted by repaired postsynaptic dendrites and the cochlear potential had partially recovered. This process of neo-synaptogenesis was completed and the potentials were fully restored at 5 days post exposure. As shown by in situ hybridization, an up-regulation of NMDA and metabotropic glutamate receptors in the primary auditory neurons occurred during this process of recovery. This process of neo-synaptogenesis and functional recovery probably accounts for restoring hearing after temporary losses due to excitotoxic-related pathologies.

Animals↗

PubMed1994

Choline-acetyltransferase-like immunoreactivity in the organ of Corti of the rat during postnatal development.

The mammalian cochlea receives efferent innervation from neurons located in the superior olivary complex. This efferent olivocochlear innervation is divided in two separate systems, lateral and medial, which mainly innervate afferent dendrites connected to inner hair cells and the cell body of outer hair cells, respectively. Besides other substances, lateral and medial efferent terminals of the adult cochlea use acetylcholine (ACh) as a neurotransmitter. In this study, we have used immunocytochemistry to detect the presence of choline acetyltransferase (ChAT), the synthesizing enzyme of ACh, in efferent olivocochlear terminals during the development of the rat. The appearance and distribution of immunoreactivity to ChAT has been studied in developing rat cochleas from birth (postnatal day 1, P1) to adulthood. Attention was paid to the temporal relationships between the expression of ChAT, the presence of other putative neuroactive substances, the onset of hearing and other developmental phenomena. Our results indicate that ChAT-like immunoreactivity is already present at birth (P1) in the region of inner hair cells, that it appears at P3 in the outer hair cell area and that it reaches an adult pattern of distribution by P15. ACh may thus be present early in the developing cochlea, before the onset of hearing, as it also occurs with other putative transmitters/modulators such as enkephalins, CGRP or GABA. It is suggested that ACh could be involved in the modulation of sound-evoked potentials as soon as they appear, and in the regulation of other developmental phenomena such as neurite outgrowth or synaptogenesis.

Aging↗

PubMed1994

Excitatory amino acid antagonists protect cochlear auditory neurons from excitotoxicity.

Since ischemic damage in the brain is linked to glutamate excitotoxicity, the effects of an acute exposure to glutamate, alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) or N-methyl-D aspartate (NMDA) on the radial dendrites were compared with those occurring after a severe cochlear ischemia. Glutamate and AMPA, but not NMDA, produced a drastic swelling restricted to the radial dendrites below the inner hair cells (IHCs). At a concentration of 20 microM AMPA, a full electrophysiological recovery could be observed in some cochleas after washing the drug out. A prior perfusion of 6-7-dinitroquinoxaline-2,3-dione (DNQX, 50 microM) prevented the 25 microM AMPA-induced dendritic swelling. No protective effect of D-2-amino-5-phosphonopentanoate (D-AP5) could be observed. In the same way, ischemia (5-40 minutes) resulted in a clear swelling of the radial dendrites. While D-AP5 had no protective effects, 50 microM DNQX protected most of the radial dendrites from the ischemia-induced swelling, excepting those contacting the modiolar side of the IHCs. Finally, 50 microM DNQX + 50 microM D-AP5 resulted in a nearly complete protection of all the radial dendrites. Altogether, these results suggest that the acute swelling of radial dendrites primarily occurs via AMPA/kainate receptors. However, in radial dendrites contacting the inner hair cells on their modiolar side, NMDA receptors may be also involved.

2-Amino-5-phosphonovalerate↗

PubMed1994

Immunocytochemical detection of choline acetyltransferase in the human organ of Corti.

In the mammalian cochlea acetylcholine has been considered a major neurotransmitter of the lateral and medial efferent fibers. The aims of the present study were to investigate the expression of ChAT in the human cochlea and to develop a new method for immunohistochemical investigations in the human cochlea both at the light and electronmicroscopic level. We thus examined the ChAT-like immunoreactivity in the human inner ear using light and electron microscopy with a pre-embedding technique. Our present results agree with the previously published data acquired in rodent species. The ChAT-like immunostaining could be found in the inner spiral fibers, the inner spiral bundle, tunnel crossing fibers and at the base of the outer hair cells. No staining was noted in the negative controls experiments, while rat cochleas used as positive controls showed the usual ChAT-like immunostaining as described above. The main difference between human and rat cochleas was that the efferent nerve supply seems to be less pronounced in the human cochleas.

Animals↗