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1.
Proc Natl Acad Sci U S A ; 119(37): e2207433119, 2022 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-36074819

RESUMO

A cardinal feature of the auditory pathway is frequency selectivity, represented in a tonotopic map from the cochlea to the cortex. The molecular determinants of the auditory frequency map are unknown. Here, we discovered that the transcription factor ISL1 regulates the molecular and cellular features of auditory neurons, including the formation of the spiral ganglion and peripheral and central processes that shape the tonotopic representation of the auditory map. We selectively knocked out Isl1 in auditory neurons using Neurod1Cre strategies. In the absence of Isl1, spiral ganglion neurons migrate into the central cochlea and beyond, and the cochlear wiring is profoundly reduced and disrupted. The central axons of Isl1 mutants lose their topographic projections and segregation at the cochlear nucleus. Transcriptome analysis of spiral ganglion neurons shows that Isl1 regulates neurogenesis, axonogenesis, migration, neurotransmission-related machinery, and synaptic communication patterns. We show that peripheral disorganization in the cochlea affects the physiological properties of hearing in the midbrain and auditory behavior. Surprisingly, auditory processing features are preserved despite the significant hearing impairment, revealing central auditory pathway resilience and plasticity in Isl1 mutant mice. Mutant mice have a reduced acoustic startle reflex, altered prepulse inhibition, and characteristics of compensatory neural hyperactivity centrally. Our findings show that ISL1 is one of the obligatory factors required to sculpt auditory structural and functional tonotopic maps. Still, upon Isl1 deletion, the ensuing central plasticity of the auditory pathway does not suffice to overcome developmentally induced peripheral dysfunction of the cochlea.


Assuntos
Vias Auditivas , Núcleo Coclear , Células Ciliadas Auditivas , Proteínas com Homeodomínio LIM , Neurogênese , Gânglio Espiral da Cóclea , Fatores de Transcrição , Animais , Vias Auditivas/embriologia , Cóclea/embriologia , Cóclea/inervação , Núcleo Coclear/embriologia , Células Ciliadas Auditivas/fisiologia , Proteínas com Homeodomínio LIM/genética , Proteínas com Homeodomínio LIM/fisiologia , Camundongos , Neurogênese/genética , Gânglio Espiral da Cóclea/enzimologia , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia
2.
Biomed Res Int ; 2015: 925890, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25918725

RESUMO

Rapamycin is an antifungal agent with immunosuppressive properties. Rapamycin inhibits the mammalian target of rapamycin (mTOR) by blocking the mTOR complex 1 (mTORC1). mTOR is an atypical serine/threonine protein kinase, which controls cell growth, cell proliferation, and cell metabolism. However, less is known about the mTOR pathway in the inner ear. First, we evaluated whether or not the two mTOR complexes (mTORC1 and mTORC2, resp.) are present in the mammalian cochlea. Next, tissue explants of 5-day-old rats were treated with increasing concentrations of rapamycin to explore the effects of rapamycin on auditory hair cells and spiral ganglion neurons. Auditory hair cell survival, spiral ganglion neuron number, length of neurites, and neuronal survival were analyzed in vitro. Our data indicates that both mTOR complexes are expressed in the mammalian cochlea. We observed that inhibition of mTOR by rapamycin results in a dose dependent damage of auditory hair cells. Moreover, spiral ganglion neurite number and length of neurites were significantly decreased in all concentrations used compared to control in a dose dependent manner. Our data indicate that the mTOR may play a role in the survival of hair cells and modulates spiral ganglion neuronal outgrowth and neurite formation.


Assuntos
Células Ciliadas Auditivas/enzimologia , Neuritos/enzimologia , Sirolimo/efeitos adversos , Gânglio Espiral da Cóclea/enzimologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Ciliadas Auditivas/patologia , Alvo Mecanístico do Complexo 1 de Rapamicina , Complexos Multiproteicos/metabolismo , Neuritos/patologia , Ratos , Ratos Wistar , Sirolimo/farmacologia , Gânglio Espiral da Cóclea/patologia , Serina-Treonina Quinases TOR/metabolismo
3.
PLoS One ; 10(3): e0121599, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25811375

RESUMO

The kinase Akt is a key downstream mediator of the phosphoinositide-3-kinase signaling pathway and participates in a variety of cellular processes. Akt comprises three isoforms each encoded by a separate gene. There is evidence to indicate that Akt is involved in the survival and protection of auditory hair cells in vitro. However, little is known about the physiological role of Akt in the inner ear-especially in the intact animal. To elucidate this issue, we first analyzed the mRNA expression of the three Akt isoforms in the inner ear of C57/BL6 mice by real-time PCR. Next, we tested the susceptibility to gentamicin-induced auditory hair cell loss in isoform-specific Akt knockout mice compared to wild-types (C57/BL6) in vitro. To analyze the effect of gene deletion in vivo, hearing and cochlear microanatomy were evaluated in Akt isoform knockout animals. In this study, we found that all three Akt isoforms are expressed in the cochlea. Our results further indicate that Akt2 and Akt3 enhance hair cell resistance to ototoxicity, while Akt1 does not. Finally, we determined that untreated Akt1 and Akt2/Akt3 double knockout mice display significant hearing loss, indicating a role for these isoforms in normal hearing. Taken together, our results indicate that each of the Akt isoforms plays a distinct role in the mammalian inner ear.


Assuntos
Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/enzimologia , Audição/fisiologia , Mamíferos/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Animais Recém-Nascidos , Sobrevivência Celular , Suscetibilidade a Doenças , Potenciais Evocados Auditivos do Tronco Encefálico , Regulação Enzimológica da Expressão Gênica , Gentamicinas , Isoenzimas/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Proto-Oncogênicas c-akt/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Gânglio Espiral da Cóclea/enzimologia , Estria Vascular/enzimologia
4.
Cell Mol Neurobiol ; 34(7): 1011-21, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24935409

RESUMO

Matrix metalloproteinases (MMPs) play an important role in modeling of the extracellular matrix. There is increasing evidence that these proteases are important in neurite elongation and axonal guidance during development in the central nervous system and retina. Moreover, they are also expressed after acute injury and can be the key mediators of pathogenesis. However, the role of MMPs in the inner ear is largely unknown. Our group recently demonstrated that general inhibition of MMPs resulted in auditory hair cell loss in vitro. In the present study, we investigated the role of MMPs in inner ear spiral ganglion neuron (SGN) survival, neuritogenesis and neurite extension by blocking MMPs known to be involved in axonal guidance, neurite elongation, and apoptosis in other neuronal systems. Spiral ganglion (SG) explants from 5-day-old Wistar rats were treated with different concentrations of the general MMP inhibitor GM6001, a specific MMP-2 inhibitor, and a specific MMP-9 inhibitor, in vitro. The general inhibitor of MMPs and the specific inhibition of MMP-2 significantly reduced both the number of neurites that extended from SG explants, as well as the length of individual neurites. However, neither the general inhibitor of MMPs nor the specific inhibition of MMP-2 influenced SGN survival. Inhibition of MMP-9 had no influence on SGNs. The data suggest that MMPs, and more specifically MMP-2, influence the growth of developing afferent neurites in the mammalian inner ear in vivo.


Assuntos
Orelha Interna/citologia , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Neurônios/enzimologia , Gânglio Espiral da Cóclea/enzimologia , Animais , Contagem de Células , Sobrevivência Celular/efeitos dos fármacos , Células Labirínticas de Suporte/citologia , Células Labirínticas de Suporte/enzimologia , Metaloproteinase 9 da Matriz/genética , Neuritos/efeitos dos fármacos , Neuritos/enzimologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Gânglio Espiral da Cóclea/citologia , Gânglio Espiral da Cóclea/efeitos dos fármacos
5.
Kulak Burun Bogaz Ihtis Derg ; 23(2): 104-11, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23611324

RESUMO

OBJECTIVES: This study aims to investigate the effect of exogenous glucocorticoid exposure in the prenatal period on hearing and to evaluate the effectiveness of caffeic acid phenethyl ester (CAPE), an antioxidant, on the prevention of the inner ear injury. MATERIALS AND METHODS: Dexamethasone was given to half of twelve Sprague-Dawley pregnant rats and the distilled water was given to the remaining half. The real subjects were obtained by born of the offsprings. When the all subjects were two months of age, they were exposed to 110 dB noise during four hours as a stressor effect. These subjects were divided into three groups. Group 1: subjects to whose mothers were given distilled water; Group 2: subjects to whose mothers were given dexamethasone; Group 3: subjects to whose mothers were given dexamethasone and CAPE. RESULTS: While there was no statistical significance in hearing thresholds which exposed and not exposed to exogenous dexamethasone before noise exposure (p>0.05) between the groups, the elevation of hearing thresholds of subjects which exposed to exogenous dexamethasone was statistically significant after noise exposure (p<0.05). CONCLUSION: Prenatally exposure to exogenous glucocorticoids may cause the inner ear susceptible to the effect of noise, and CAPE is effective to prevent the possible damage.


Assuntos
Ácidos Cafeicos/administração & dosagem , Dexametasona/efeitos adversos , Orelha Interna/efeitos dos fármacos , Glucocorticoides/efeitos adversos , Audição/efeitos dos fármacos , Álcool Feniletílico/análogos & derivados , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Animais , Limiar Auditivo/efeitos dos fármacos , Caspases/análise , Dexametasona/administração & dosagem , Potenciais Evocados Auditivos do Tronco Encefálico/efeitos dos fármacos , Feminino , Glucocorticoides/administração & dosagem , Imuno-Histoquímica , Ruído/efeitos adversos , Órgão Espiral/enzimologia , Álcool Feniletílico/administração & dosagem , Gravidez , Ratos , Ratos Sprague-Dawley , Gânglio Espiral da Cóclea/enzimologia , Estresse Fisiológico , Estria Vascular/enzimologia
6.
Anat Rec (Hoboken) ; 296(2): 326-32, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23165776

RESUMO

Mitochondrial degeneration in the inner ear is likely a contributing factor in age-related hearing loss and other otopathologies such as Meniere's disease. Most mitochondrial proteins are synthesized in the cytosol and imported through the mitochondrial membranes by translocators. The translocase of the outer membrane (Tom) is the universal entry gate for all proteins that are imported into mitochondria. Altered function of the translocator could alter protein transport into the mitochondria, and disrupt function. In this study, we determined the immunolocalization of Tom20, a major mitochondrial protein import receptor, in microdissected human cochlea frozen sections obtained from postmortem autopsy and celloidin-embedded archival specimens. We used affinity purified rabbit polyclonal antibodies against Tom20. We also determined the Tom20 immunolocalization in the mouse inner ear. In the human and mouse cochlea, Tom20 was ubiquitously distributed in the organ of Corti, allowing well-delineated visualization of inner and outer hair cells. Tom20 immunoreactivity localized in the cytoplasm of spiral ganglia neurons. In the inner ear of aged subjects with Meniere's disease, there was decreased expression of Tom20. These results suggest that Tom20 can be used in the inner ear as a marker for mitochondrial protein import.


Assuntos
Cóclea/enzimologia , Imuno-Histoquímica , Proteínas de Membrana Transportadoras/análise , Doença de Meniere/enzimologia , Mitocôndrias/enzimologia , Membranas Mitocondriais/enzimologia , Receptores de Superfície Celular/análise , Adolescente , Idoso , Idoso de 80 Anos ou mais , Animais , Biomarcadores/análise , Criança , Feminino , Fixadores , Formaldeído , Secções Congeladas , Células Ciliadas Auditivas/enzimologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos CBA , Microdissecção , Pessoa de Meia-Idade , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Ratos , Gânglio Espiral da Cóclea/enzimologia , Fixação de Tecidos/métodos
7.
Hear Res ; 283(1-2): 33-44, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22154930

RESUMO

Regrowth of peripheral spiral ganglion neuron (SGN) fibers is a primary objective in efforts to improve cochlear implant outcomes and to potentially reinnervate regenerated hair cells. Cyclic adenosine monophosphate (cAMP) regulates neurite growth and guidance via activation of protein kinase A (PKA) and Exchange Protein directly Activated by Cylic AMP (Epac). Here we explored the effects of cAMP signaling on SGN neurite length in vitro. We find that the cAMP analog, cpt-cAMP, exerts a biphasic effect on neurite length; increasing length at lower concentrations and reducing length at higher concentrations. This biphasic response occurs in cultures plated on laminin, fibronectin, or tenascin C suggesting that it is not substrate dependent. cpt-cAMP also reduces SGN neurite branching. The Epac-specific agonist, 8-pCPT-2'-O-Me-cAMP, does not alter SGN neurite length. Constitutively active PKA isoforms strongly inhibit SGN neurite length similar to higher levels of cAMP. Chronic membrane depolarization activates PKA in SGNs and also inhibits SGN neurite length. However, inhibition of PKA fails to rescue neurite length in depolarized cultures implying that activation of PKA is not necessary for the inhibition of SGN neurite length by chronic depolarization. Expression of constitutively active phosphatidylinositol 3-kinase, but not c-Jun N-terminal kinase, isoforms partially rescues SGN neurite length in the presence of activated PKA. Taken together, these results suggest that activation of cAMP/PKA represents a potential strategy to enhance SGN fiber elongation following deafness; however such therapies will likely require careful titration so as to promote rather than inhibit nerve fiber regeneration.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Regeneração Nervosa , Neurônios/enzimologia , Gânglio Espiral da Cóclea/enzimologia , Animais , Animais Recém-Nascidos , Colforsina/farmacologia , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Proteínas Quinases Dependentes de AMP Cíclico/genética , Relação Dose-Resposta a Droga , Ativação Enzimática , Ativadores de Enzimas/farmacologia , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Potenciais da Membrana , Regeneração Nervosa/efeitos dos fármacos , Neuritos/enzimologia , Neurônios/efeitos dos fármacos , Fosfatidilinositol 3-Quinase/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Ratos , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais , Gânglio Espiral da Cóclea/efeitos dos fármacos , Tionucleotídeos/farmacologia , Técnicas de Cultura de Tecidos , Transfecção
8.
Biochem Biophys Res Commun ; 415(1): 94-8, 2011 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-22020098

RESUMO

Expression of antioxidant enzymes is regulated by transcription factor NF-E2-related factor (Nrf2) and induced by oxidative stress. Reactive oxygen species contribute to the formation of several types of cochlear injuries, including age-related hearing loss and gentamicin ototoxicity. In this study, we examined the roles of Nrf2 in age-related hearing loss and gentamicin ototoxicity by measuring auditory brainstem response thresholds in Nrf2-knockout mice. Although Nrf2-knockout mice maintained normal auditory thresholds at 3 months of age, their hearing ability was significantly more impaired than that of age-matched wild-type mice at 6 and 11 months of age. Additionally, the numbers of hair cells and spiral ganglion cells were remarkably reduced in Nrf2-knockout mice at 11 months of age. To examine the importance of Nrf2 in protecting against gentamicin-induced ototoxicity, 3-day-old mouse organ of Corti explants were cultured with gentamicin. Hair cell loss caused by gentamicin treatment was enhanced in the Nrf2-deficient tissues. Furthermore, the expressions of some Nrf2-target genes were activated by gentamicin treatment in wild-type mice but not in Nrf2-knockout mice. The present findings indicate that Nrf2 protects the inner ear against age-related hearing injuries and gentamicin ototoxicity by up-regulating antioxidant enzymes and detoxifying proteins.


Assuntos
Envelhecimento , Antibacterianos/efeitos adversos , Orelha Interna/enzimologia , Gentamicinas/efeitos adversos , Perda Auditiva/induzido quimicamente , Perda Auditiva/genética , Fator 2 Relacionado a NF-E2/fisiologia , Animais , Orelha Interna/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento , Células Ciliadas Auditivas/efeitos dos fármacos , Células Ciliadas Auditivas/enzimologia , Heme Oxigenase-1/genética , Camundongos , Camundongos Knockout , NAD(P)H Desidrogenase (Quinona)/genética , Fator 2 Relacionado a NF-E2/genética , Elementos de Resposta , Gânglio Espiral da Cóclea/efeitos dos fármacos , Gânglio Espiral da Cóclea/enzimologia , Superóxido Dismutase/genética , Superóxido Dismutase-1
9.
Hear Res ; 278(1-2): 77-85, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21554942

RESUMO

Jun N-terminal kinase (JNK) is a multifunctional protein kinase crucial for neuronal apoptosis as well as neurite growth. We have previously shown that JNK activity is correlated with spiral ganglion neuron (SGN) apoptosis following hair cell loss in rats (Alam et al., 2007) implying that JNK inhibition may have therapeutic potential to protect SGNs in deaf individuals. Here we investigated the role of JNK in neurite outgrowth from cultured neonatal rat and mouse SGNs. We show that JNK is required for initial growth of neurites and for continued extension of already established neurites. The effect of JNK inhibition on neurite growth is rapid and is also rapidly reversible after washout of the inhibitor. Using phosphoJNK immunoreactivity as an indicator, we show that JNK is activated in growth cones within 30 min after transfer to medium lacking neurotrophic stimuli (5 K medium) but activation in the nucleus and soma requires hours. By transfecting epitope-tagged JNK1, JNK2, or JNK3 isoforms into SGNs, we found that all are present in the nucleus and cytoplasm and that there is no preferential redistribution to the nucleus after transfer to 5 K medium. Cotransfection of dominant-negative (dn) JNK1 and JNK2 into SGNs reduced neurite growth, although transfection of dnJNK1 or dnJNK2 alone had no significant effect. SGNs cultured from JNK3(-/-) mice showed reduced neurite growth that was further reduced by transfection of dnJNK1 and dnJNK2. This indicates that all three JNK isoforms promote SGN neurite growth although there may be functional redundancy between JNK1 and JNK2.


Assuntos
Sistema de Sinalização das MAP Quinases , Neuritos/enzimologia , Neuritos/ultraestrutura , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/inervação , Animais , Células Cultivadas , Ativação Enzimática , Cinética , MAP Quinase Quinase Quinases/metabolismo , Camundongos , Camundongos Knockout , Proteína Quinase 10 Ativada por Mitógeno/deficiência , Proteína Quinase 10 Ativada por Mitógeno/genética , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Proteína Quinase 8 Ativada por Mitógeno/deficiência , Proteína Quinase 8 Ativada por Mitógeno/genética , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Proteína Quinase 9 Ativada por Mitógeno/deficiência , Proteína Quinase 9 Ativada por Mitógeno/genética , Proteína Quinase 9 Ativada por Mitógeno/metabolismo , Neurônios/enzimologia , Neurônios/ultraestrutura , Fosforilação , Ratos , Gânglio Espiral da Cóclea/ultraestrutura , Frações Subcelulares/enzimologia , Transfecção
10.
Proc Natl Acad Sci U S A ; 107(29): 13051-6, 2010 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-20616061

RESUMO

A significantly increased risk for dominant sensorineural deafness in patients who have Hirschsprung disease (HSCR) caused by endothelin receptor type B and SOX10 has been reported. Despite the fact that c-RET is the most frequent causal gene of HSCR, it has not been determined whether impairments of c-Ret and c-RET cause congenital deafness in mice and humans. Here, we show that impaired phosphorylation of c-Ret at tyrosine 1062 causes HSCR-linked syndromic congenital deafness in c-Ret knockin (KI) mice. The deafness involves neurodegeneration of spiral ganglion neurons (SGNs) with not only impaired phosphorylation of Akt and NF-kappaB but decreased expression of calbindin D28k in inner ears. The congenital deafness involving neurodegeneration of SGNs in c-Ret KI mice was rescued by introducing constitutively activated RET. Taken together with our results for three patients with congenital deafness with c-RET-mediated severe HSCR, our results indicate that c-Ret and c-RET are a deafness-related molecule in mice and humans.


Assuntos
Perda Auditiva/complicações , Perda Auditiva/enzimologia , Doença de Hirschsprung/complicações , Doença de Hirschsprung/enzimologia , Proteínas Proto-Oncogênicas c-ret/metabolismo , Substituição de Aminoácidos/genética , Animais , Surdez/complicações , Surdez/enzimologia , Ativação Enzimática , Técnicas de Introdução de Genes , Perda Auditiva/congênito , Imuno-Histoquímica , Camundongos , Mutação/genética , NF-kappa B/metabolismo , Degeneração Neural/enzimologia , Degeneração Neural/patologia , Neurônios/enzimologia , Neurônios/patologia , Neurônios/ultraestrutura , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-ret/genética , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/patologia , Gânglio Espiral da Cóclea/ultraestrutura
11.
Auris Nasus Larynx ; 37(3): 274-80, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19900774

RESUMO

OBJECTIVE: Fabry disease is characterized by genetic alpha-galactosidase A deficiency, resulting in accumulation of glycolipids (GL-3) and tissue damage. Hearing loss is also common and attributed to GL-3 accumulation in the inner ear. The only reported histological studies dealt with murine and human specimens. Accordingly, histopathological studies of the cochlea were performed on an alpha-galactosidase A deficient murine model of Fabry disease, using C57BL6/J mice as the controls. METHODS: The hearing ability was evaluated using the ABR threshold, while cochlear specimens were observed light microscopically and ultrathin temporal bone sections by TEM. RESULTS: HE staining showed no accumulation of GL-3 or abnormal cochlear morphology in the alpha-galactosidase A deficient mice, but toluidine blue staining and TEM revealed GL-3 accumulation in the stria vascularis and kidney. No GL-3 accumulation was detected in the C57BL6/J controls by either HE staining or TEM. The alpha-galactosidase A deficient mice and the controls showed no clear differences in the ABR threshold (hearing acuity), but for older animals the threshold was higher in the C57BL6/J controls. CONCLUSION: In summary, although the alpha-galactosidase A deficient mice showed no clear hearing loss, GL-3 accumulation was demonstrated in the cochlea.


Assuntos
Cóclea/enzimologia , Cóclea/patologia , Doença de Fabry/enzimologia , Doença de Fabry/metabolismo , Doença de Fabry/patologia , Animais , Modelos Animais de Doenças , Potenciais Evocados Auditivos do Tronco Encefálico/fisiologia , Doença de Fabry/genética , Perda Auditiva Neurossensorial/diagnóstico , Camundongos , Camundongos Endogâmicos C57BL , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/patologia
12.
Hear Res ; 253(1-2): 116-24, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19376215

RESUMO

Manganese superoxide dismutase 2 (SOD2) is a key metabolic anti-oxidant enzyme for detoxifying free radicals inside mitochondria. This study documents a gradient in expression of SOD2 by spiral ganglion cells in basal versus apical turn of cochlea that is consistent with differential vulnerability of high frequency hearing to free radical damage. Immunohistochemical methods were used to identify distribution of SOD2 in temporal bone sections from mice, rats, macaques, and humans. In mice and rats, both the proportion of SOD2 immunopositive type 1 spiral ganglion cells and the intensity of immunoreactivity were elevated near cochlear apex. In macaques and humans, the proportion of SO2 immunopositive spiral ganglion cells was equal across cochlear turn, but the intensity of immunoreactivity remained highest for ganglion cells near cochlear apex. Strong SOD2 immunoreactivity was also observed in human type 1 spiral ganglion cells. The average area density of SOD2 immunoreactivity in ganglion cells for each species and cochlear turn showed an allometric relationship with body weight, which is consistent with a conserved basal metabolic characteristic. These findings suggest that spiral ganglion cell responses to ROS exposure may vary along cochlear spiral with lower response capacity at cochlear base contributing to cumulative susceptibility to high frequency hearing loss.


Assuntos
Gânglio Espiral da Cóclea/enzimologia , Superóxido Dismutase/metabolismo , Animais , Pré-Escolar , Feminino , Humanos , Imuno-Histoquímica , Macaca , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Modelos Biológicos , Ratos , Ratos Long-Evans , Ratos Wistar , Especificidade da Espécie , Gânglio Espiral da Cóclea/citologia , Gânglio Espiral da Cóclea/metabolismo , Superóxidos/metabolismo , Distribuição Tecidual
13.
J Assoc Res Otolaryngol ; 10(1): 37-49, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19082858

RESUMO

Processing of sound in the cochlea involves both afferent and efferent innervation. The Na,K-ATPase (NKA) is essential for cells that maintain hyperpolarized membrane potentials and sodium and potassium concentration gradients. Heterogeneity of NKA subunit expression is one mechanism that tailors physiology to particular cellular demands. Therefore, to provide insight into molecular differences that distinguish the various innervation pathways in the cochlea, we performed a variety of double labeling experiments with antibodies against three of the alpha isoforms of the NKA (NKA alpha 1-3) and markers identifying particular subsets of neurons or supporting cells in whole mount preparations of the organ of Corti and spiral ganglion. We found that the NKA alpha 3 is abundantly expressed within the membranes of the spiral ganglion somata, the type I afferent terminals contacting the inner hair cells, and the medial efferent terminals contacting the outer hair cells. We also found expression of the NKA alpha 1 in the supporting cells that neighbor the inner hair cells and express the glutamate transporter GLAST. These findings suggest that both the NKA alpha 1 and NKA alpha 3 are poised to play an essential role in the regulation of the type I afferent synapses, the medial efferent synapses, and also glutamate transport from the afferent-inner hair cell synapse.


Assuntos
Órgão Espiral/enzimologia , ATPase Trocadora de Sódio-Potássio/biossíntese , Gânglio Espiral da Cóclea/enzimologia , Animais , Transporte Biológico Ativo/fisiologia , Transportador 1 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/metabolismo , Células Ciliadas Auditivas Internas/enzimologia , Imuno-Histoquímica , Isoenzimas/biossíntese , Células Labirínticas de Suporte/enzimologia , Neurônios Aferentes/enzimologia , Neurônios Eferentes/enzimologia , Órgão Espiral/citologia , Terminações Pré-Sinápticas/enzimologia , Ratos , Gânglio Espiral da Cóclea/citologia , Sinapses/fisiologia
14.
Artigo em Chinês | MEDLINE | ID: mdl-18717313

RESUMO

OBJECTIVE: To study the differences of regulation of sodium salicylate on the auditory brain stem response (ABR) threshold and expression of glutamic acid decarboxylase (GAD) protein in spiral ganglion of juvenile and adult guinea pigs. METHODS: Fourty juvenile guinea pigs which were born just four days and fourty adult guinea pigs which were born thirty days were selected. They were divided four groups (group A; group B; group C; group D). ABR threshold was detected before administration, after administration for 15 days and after administration stopped for 30 days. The protein expression of GAD were measured after administration for 15 days and after administration stopped for 30 days by the method of immunohistochemistry. RESULTS: ABR threshold of juvenile sodium salicylate groups (group C) was increased remarkably than that of before administration and the control after administration for 15 days (P < 0.001). ABR threshold of group C was returned to the level of that of before administration and after administration stopped for 30 days. ABR threshold of adult sodium salicylate groups (group D) was increased remarkably than that of before administration and the control after administration for 15 days (P < 0.001). ABR threshold of group D was kept the high level after administration stopped for 30 days. The protein expression of GAD of sodium salicylate groups (group C and D) was decreased than that of the control after administration for 15 days. The protein expression of group C was more visible regression than that of group D (t = 4.7, P < 0.001). The protein expression of group C was returned the level of before administration after administration stopped for 30 days, but the protein expression of group D was kept the high level. CONCLUSIONS: The results suggest that sodium salicylate can regulate differently ABR threshold and expression of GAD protein in spiral ganglion of juvenile and adult guinea pigs. The effects of sodium salicylate on ABR threshold and expression of GAD protein in spiral ganglion of juvenile pigs are more noticeable than that of adult guinea pigs, but these changes are easier to return the normal than that of adult guinea pigs.


Assuntos
Limiar Auditivo/efeitos dos fármacos , Potenciais Evocados Auditivos do Tronco Encefálico/efeitos dos fármacos , Glutamato Descarboxilase/metabolismo , Salicilato de Sódio/farmacologia , Gânglio Espiral da Cóclea/efeitos dos fármacos , Animais , Potenciais Evocados Auditivos do Tronco Encefálico/fisiologia , Cobaias , Gânglio Espiral da Cóclea/enzimologia
15.
J Neurotrauma ; 25(1): 72-7, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18355160

RESUMO

Noise exposure is known to induce cell death signaling in the cochlea. Since c-Jun N-terminal kinase (JNK) signaling is known to induce both cell survival and apoptosis, the present study focused on early changes (within 24 h) after impulse noise exposure, inquiring whether cell death is always related to phosphorylation of JNK in the inner ear. Anesthetized adult albino rats were exposed to a single impulse noise exposure (194 kPa) and sacrificed 3 or 24 h later. Paraffin-embedded sections were examined for positive staining of phosphorylated JNK and the presence of cells with fragmented DNA (TUNEL staining). Positive TUNEL staining was observed at the spiral limbus and in the stria vascularis at 24 h following impulse noise exposure, but no correlation with JNK activation was found at these locations. In the hearing organ (organ of Corti) and in the lateral wall, TUNEL-reactive cells were observed at 24 h following trauma. This was preceded by p-JNK staining at 3 h, indicating JNK-activated cell death in these regions. Finally, p-JNK reactivity was observed in the spiral ganglion with no correlation to TUNEL staining within the time frame of this study. These results suggest that JNK activation following impulse noise exposure may not always be related to cell death, and conversely, some cells may die through JNK-independent signaling.


Assuntos
Cóclea/enzimologia , Células Ciliadas Auditivas/enzimologia , Perda Auditiva Provocada por Ruído/enzimologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Ruído/efeitos adversos , Estimulação Acústica , Animais , Biomarcadores/metabolismo , Morte Celular/fisiologia , Cóclea/patologia , Cóclea/fisiopatologia , Fragmentação do DNA , Ativação Enzimática/fisiologia , Feminino , Células Ciliadas Auditivas/patologia , Perda Auditiva Provocada por Ruído/fisiopatologia , Marcação In Situ das Extremidades Cortadas , Fosforilação , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/patologia , Gânglio Espiral da Cóclea/fisiopatologia , Fatores de Tempo
16.
Hear Res ; 235(1-2): 8-14, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17980526

RESUMO

Isolating cells from the cochlea to perform molecular biology assessment presents a challenge, because it is not possible to dissect pure cell pools by conventional methods. Thus, we set out to demonstrate that laser microdissection and pressure catapulting (LMPC) is superior to conventional manual cochlea dissection for this purpose. Spiral ganglions (SG) were isolated from neonatal rat cochleae by manual dissection and LMPC. Also, modioli were manually dissected. Total RNA was isolated from all three cell pools. In order to demonstrate contamination of the dissected cell pool, we determined the expression of type II iodothyronine deiodinase (D2), claudin 11 (Cld-11), neurofilament light chain (NF-L) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) transcripts by RT-PCR. The results showed that LMPC is not only a suitable method for selectively dissecting cochlear tissues, but in addition the molecular markers confirmed pure spiral ganglion cell pools without indication for any contamination by other cells. This indicates that LMPC is capable of providing a pure SG cell pool in contrast to conventional manual dissection. Therefore, LMPC presents a new technique for cochlear tissue separation improving the validity of molecular biological studies of the inner ear.


Assuntos
Separação Celular/métodos , Cóclea/química , Lasers , Microdissecção , RNA Mensageiro/análise , Gânglio Espiral da Cóclea/química , Animais , Animais Recém-Nascidos , Claudinas , Cóclea/citologia , Cóclea/enzimologia , Secções Congeladas , Gliceraldeído-3-Fosfato Desidrogenases/análise , Iodeto Peroxidase/análise , Proteínas do Tecido Nervoso/análise , Proteínas de Neurofilamentos/análise , Fenótipo , Pressão , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Gânglio Espiral da Cóclea/enzimologia , Iodotironina Desiodinase Tipo II
17.
Hear Res ; 209(1-2): 76-85, 2005 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16055286

RESUMO

Mice, in which the genetics can be manipulated and the life span is relatively short, enable evaluation of the effects of specific gene expression on cochlear degeneration over time. Antioxidant enzymes such as Cu/Zn superoxide dismutase (SOD1) protect cells from toxic, reactive oxygen species and may be involved in age-related degeneration. The effects of SOD1 deletion and over-expression on the cochlea were examined in Sod1-null mice, Sod1 transgenic mice and in age- and genetics-matched controls. Auditory brainstem responses (ABR) were measured and cochleae were histologically examined. The absence of SOD1 resulted in hearing loss at an earlier age than in wildtype or heterozygous mice. The cochleae of the null mice had severe spiral ganglion cell degeneration at 7-9 months of age. The stria vascularis in the aged, null mice was thinner than in the heterozygous or wildtype mice. Over-expression of SOD1 did not protect against hearing loss except at 24 months of age. In conclusion, SOD1 seems important for survival of cochlear neurons and the stria vascularis, however even half the amount is sufficient and an over abundance does not provide much protection from age-related hearing loss.


Assuntos
Envelhecimento/metabolismo , Perda Auditiva/enzimologia , Superóxido Dismutase/deficiência , Superóxido Dismutase/genética , Envelhecimento/patologia , Análise de Variância , Animais , Tronco Encefálico/fisiologia , Cóclea/enzimologia , Cóclea/metabolismo , Cóclea/patologia , Feminino , Expressão Gênica , Perda Auditiva/patologia , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Órgão Espiral/enzimologia , Órgão Espiral/metabolismo , Órgão Espiral/patologia , Presbiacusia/fisiopatologia , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/metabolismo , Gânglio Espiral da Cóclea/patologia , Estria Vascular/enzimologia , Estria Vascular/metabolismo , Estria Vascular/patologia , Superóxido Dismutase-1
18.
Hear Res ; 202(1-2): 21-7, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15811695

RESUMO

The aim of this study was to investigate the involvement of oxidative stress and apoptosis in an animal model of Meniere's disease. Endolymphatic hydrops (ELH) is generally accepted as the decisive histological characteristic of Meniere's disease. Closure of the endolymphatic duct (Kimura's method) was used to induce endolymphatic hydrops in guinea pigs. Sham-operated animals served as controls. After 4 weeks the animals operated showed a significant elevation of the hearing thresholds as measured by audiometric brainstem responses (ABR) pre- and postoperatively. Immediately after the second ABR measurement, the animals were sacrificed for further immunohistological examinations of the inner ear with specific antibodies to active caspase-3 (cas-3) as a marker for apoptosis and antibodies to 8-isoprostane (8-iso) and nitrotyrosine (NT) as indicators of oxidative stress. Compared with the sham-operated controls, hydropic cochleae showed strong immunostaining for both oxidative stress markers in spiral ganglion cells, in the blood-vessels and fibrocytes of the lateral wall, as well as in supporting cells of the organ of Corti. Activation of cas-3 in spiral ganglion cells and the lateral wall was found exclusively in hydropic cochleae. Our findings suggest that oxidative stress is involved in the development of endolymphatic hydrops and may lead to cellular damage which induces apoptosis by activation of cas-3. Apoptotic cell death might contribute to the sensorineural hearing loss found in later stages of Meniere's disease.


Assuntos
Caspases/metabolismo , Cóclea/metabolismo , Doenças Cocleares/metabolismo , Dinoprosta/análogos & derivados , Hidropisia Endolinfática/metabolismo , Estresse Oxidativo , Tirosina/análogos & derivados , Animais , Apoptose , Audiometria , Caspase 3 , Cóclea/enzimologia , Cóclea/fisiopatologia , Doenças Cocleares/enzimologia , Doenças Cocleares/patologia , Doenças Cocleares/fisiopatologia , Dinoprosta/metabolismo , Modelos Animais de Doenças , Hidropisia Endolinfática/enzimologia , Hidropisia Endolinfática/patologia , Hidropisia Endolinfática/fisiopatologia , Ativação Enzimática , Potenciais Evocados Auditivos do Tronco Encefálico , Cobaias , Imuno-Histoquímica/métodos , Doença de Meniere/metabolismo , Gânglio Espiral da Cóclea/enzimologia , Gânglio Espiral da Cóclea/patologia , Coloração e Rotulagem , Tirosina/metabolismo
19.
Acta Otolaryngol Suppl ; (554): 12-6, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15513504

RESUMO

Boettcher cells lie on the basilar membrane beneath Claudius cells. The cells are considered supporting cells for the organ of Corti, and present only in the lower turn of the cochlea, which responds to high-frequency sound. Boettcher cells interdigitate with each other, and project microvilli into the intercellular space. Their structural specialization suggests that Boettcher cells may play a significant role in the function of the cochlea. Nitric oxide synthase (NOS) has previously been detected in substructures of the cochlea. In the cochlea, it is believed that nitric oxide plays an important role in neurotransmission, blood flow regulation, and induction of cytotoxicity under pathological conditions. Findings concerning detection of NOS on Boettcher cells are rare. We demonstrated here the localization of NOS on Boettcher cells of the rat by immunohistochemistry using polyclonal antibody to NOS. On observation with the light microscope using DAB staining, positive immunostaining to NOS was observed in Boettcher cells. In immunoelectron micrographs, NOS was detected abundantly in the cytoplasm of the interdigitations. This suggests that the interdigitations may play significant roles by using NOS. It follows from this that the nitric oxide (NO) on Boettcher cells may influences neighboring Boettcher cells. The ultrastructure of Boettcher cells suggests that they may be active cells, which perform both secretory and absorptive functions.


Assuntos
Cóclea/citologia , Cóclea/enzimologia , Óxido Nítrico Sintase/metabolismo , Animais , Membrana Basilar/citologia , Membrana Basilar/enzimologia , Cóclea/ultraestrutura , Sequestradores de Radicais Livres/metabolismo , Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/enzimologia , Masculino , Microscopia Eletrônica de Transmissão , Microscopia Imunoeletrônica , Neurotransmissores/metabolismo , Neurotransmissores/fisiologia , Óxido Nítrico/fisiologia , Óxido Nítrico Sintase/imunologia , Órgão Espiral/citologia , Órgão Espiral/enzimologia , Ratos , Gânglio Espiral da Cóclea/citologia , Gânglio Espiral da Cóclea/enzimologia , Estria Vascular/citologia , Estria Vascular/enzimologia , Vasoconstrição/fisiologia
20.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 29(6): 682-5, 2004 Dec.
Artigo em Chinês | MEDLINE | ID: mdl-16114557

RESUMO

OBJECTIVE: To investigate the influence of EGCG on H2O2-induced gene expression of manganese superoxide dismutase (MnSOD or SOD2) in cultured spiral ganglion cells (SGCs) in vitro. METHODS: SGCs were cultured in vitro, and H2O2 and/or EGCG in different concentrations were used. Semi-quantitative RT-PCR was applied to observe MnSOD gene expression in SGCs after H2O2 and EGCG treatment. RESULTS: The expression of MnSOD gene was up-regulated with the increase of the concentration of H2O2 in cultured SGCs, and MnSOD gene expression was significantly up-regulated at a dose of H2O2 > or =100 micromol/L. However, this up-regulation was suppressed after simultaneously treated with 100 microg/ml EGCG. CONCLUSION: EGCG suppresses H2O2-induced up-regulation of MnSOD gene expression in cultured SGCs by getting rid of oxygen free radicals, reinforcing the activity of antioxidant enzymes such as MnSOD, and protects cultured SGCs from H2O2-induced oxidizing damage.


Assuntos
Catequina/análogos & derivados , Peróxido de Hidrogênio/farmacologia , Gânglio Espiral da Cóclea/enzimologia , Superóxido Dismutase/biossíntese , Animais , Catequina/farmacologia , Células Cultivadas , Cóclea/citologia , Cóclea/inervação , Feminino , Masculino , Camundongos , Gânglio Espiral da Cóclea/citologia , Superóxido Dismutase/genética
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