噪声性聋小鼠耳蜗螺旋神经节生长相关蛋白-43变化

目的通过噪声引起4周龄昆明小鼠出现暂时性阈移（tmporary threshold shift,TTS）和永久性阈移（permanent threshold shift,PTS）,观察听觉通路耳蜗螺旋神经节神经元（spiral ganglion of cochlea,CG）中生长相关蛋白（growth associated protein 43,GAP-43）变化,探讨听觉损伤后内耳突触修复的可能机制。方法采用32只昆明小鼠制作噪声性聋动物模型,进行听觉脑干诱发反应听力检测,用免疫组化法对耳蜗听觉通路中GAP-43在神经损伤刺激的表达进行检测。结果噪声性聋引起PTS后CG的GAP-43表达在损伤后第7天出现增加,第14天仍增加明显,而TTS组无明显变化。结论噪声性聋听觉通路神经性损伤作用后7天,GAP-43出现增高说明内耳开始出现突触修复。     

Mn-SOD在螺旋神经节区域分布差异与噪声性听力损伤的关系

目的探讨噪声暴露前后锰超氧化物歧化酶(Mn-SOD)在大鼠螺旋神经节不同区域表达分布的差异与噪声性聋高频听力易损性的相关性。方法 SD大鼠随机分为两组:实验组给予白噪声115dB2h3d造模,不处理者为对照组。ABR测听分析噪声损伤情况;免疫组织化学染色法分析噪声暴露前后Mn-SOD在螺旋神经节不同区域的表达水平;黄嘌呤氧化酶法检测Mn-SOD在螺旋神经节不同区域的活性及变化趋势。结果①ABR:大鼠噪声暴露后与暴露前相比,4、8、16、20、32kHzABR阈值均明显上移,以高频听阈变化更为显著;②免疫组织化学染色:在正常情况下,顶部螺旋神经节的Mn-SOD的阳性表达较底部明显;噪声暴露后,Mn-SOD在螺旋神经节的表达较对照组相应部位均显著增高;且顶部较底部的表达增强更为显著;③Mn-SOD活性测定:在正常情况下,顶部螺旋神经节的Mn-SOD活性与底部相比无统计学差异,噪声暴露后,Mn-SOD活性较对照组相应部位均有下降,且底部较顶部更为显著。结论 Mn-SOD在螺旋神经节顶、底部区域的表达分布差异,可能是噪声性聋高频听力易损性的分子机制之一。     

强噪声暴露下豚鼠耳蜗螺旋神经节细胞凋亡及Caspase-3的表达

目的探讨强噪声能否诱导豚鼠耳蜗螺旋神经节细胞（sprial ganglion cell,SGC）的凋亡及SGC的凋亡是否与凋亡蛋白酶Caspase-3信号转导有关。方法选用健康雄性白色豚鼠20只,随机分为4组,每组5只,其中3组为实验组,1组为对照组。将实验组动物暴露于4kHz、120dB SPL窄带噪声中4h,分别测试噪声刺激停止后1、4、14d及对照组豚鼠听性脑干反应（auditory brainstem response,ABR）。通过透射电镜和脱氧核糖核甘酸末端转移酶介导的缺口末端标记法（ternial-deoxynucleotidyl transferase mediated nick end labeing,TUNEL）,检测SGC凋亡细胞,免疫组织化学方法检查Caspase-3蛋白的表达。结果透射电镜观察发现实验组SGC出现了凋亡细胞的特征性改变。实验组SGC的TUNEL染色明显增强,Caspase-3的表达增高,与对照组比较差异均有显著统计学意义（P〈0.01）。结论强噪声可以诱导SGC凋亡;SGC凋亡与caspase-3的激活有关。     

听性稳态反应对噪声性聋客观评估的价值

目的 研究噪声性聋患者的听性稳态反应(ASSR)阈与纯音听阈的关系,为客观评估噪声性聋及伪聋提供依据.方法 对40例(80耳)噪声性聋患者及20例(40耳)正常人分别进行ASSR与纯音听阈测试,分别比较0.25、0.5、1、2、3、4、6 kHz各频率ASSR反应阈与纯音听阈的差值及相关性.结果 正常组在0.25、0.5 kHz的ASSR反应阈与纯音听阈相比差异无统计学意义(P＞0.05),在1、2、3、4、6 kHz的ASSR反应阈高于纯音听阈,二者相比差异有统计学意义(P＜0.05).噪声性聋组除2 kHz外其余各频率ASSR的反应阈与纯音听阈相比差异无统计学意义(P＞0.05),其ASSR反应阈与纯音听阈在0.25、0.5、1、2、3、4、6 kHz处的相关系数分别为0.55、0.62、0.55、0.49、0.54、0.73、0.73.结论 噪声性聋患者ASSR反应阈与纯音听阈显著相关,且随着听力损失的加重,ASSR反应阈越接近纯音听阈;ASSR可作为噪声性聋及伪聋患者的客观听力评估方法之一.     

脑源性神经营养因子对耳蜗螺旋神经节的保护作用

目的 观察腺疾病携带的脑源性神经营养因（brain derived neurotrophic factor,BDNF）在豚鼠耳蜗中的表达，及噪声损伤后螺旋神经节的保护作用。方法 27只白色纯豚鼠，暴露于135dBSLP，4kHz的窄带噪声4h。7d后，12只经圆注入腺病毒携带BDNF（adenoviral-mediated BDNF,ad-BDNF），12只经圆窗注入ad-LacZ,3只注入人工外淋巴液。分别于1、4、8周后取材，石蜡包埋中轴切片后，用免疫组化方法（ABC法）检测BDNF的表达。于光镜下计数螺旋神经节细胞。结果 在耳蜗各回中BDNF均有表达，4、8周组动脉较1周组织动物表达弱。在8周，注入ad-BNDF组较adLacZ组和人工淋巴组螺旋神经发生退行性病变的数目少，螺旋神经节细胞计数结果经统计学t检验，P＜0．01，差异有显著性。结论 腺病毒携带的神经营养因子在耳蜗中能高效表达，在噪声损伤情况下腺病毒携带的脑源性神经营养因子对螺旋神经节有保护作用。该研究为基因治疗感音神经性聋提供了坚实的实验基础。     

复方丹参联合星状神经节阻滞治疗噪声性聋

目前临床治疗噪声性聋主要是吸氧,应用扩张血管、活血化淤、降低血黏度、溶解小血栓等药物,并给予多种维生素和能量合剂。曾有用丹参注射液、高压氧治疗噪声性聋的报道;也有用星状神经节阻滞(steuate ganglion block,SGB)术治疗突发性聋的。近年来我们采用复方丹参注射液联合SGB治疗噪声性聋,取得了一定效果,现报告如下。     

冲击波对豚鼠螺旋神经节细胞谷氨酸免疫反应及听阈的影响

豚鼠经平均压力峰值１８４ｄＢ（ＳＰＬ）的冲击波一次暴露后，分别测８、２４、７２小时、７天的螺旋神经节（ＳＧ）细胞谷氨酸免疫反应（Ｇｌｕ－ＩＲ）阳性产物的光密度值，均较对照组明显增高（Ｐ＜０．０１）。以８小时者光密度最大，７天者最小。免疫透射电镜发现，爆震后８小时豚鼠ＳＧⅠ型、Ⅱ型细胞浆内ＬＵ－ＩＲ阳性产物明显增多。不同时间组ＡＢＲ阈移与Ｇｌｕ－ＩＲ阳性产物光密度的变化率成正相关（ｒ＝０．９１９，Ｐ＜０．０５）。提示冲击波所致听力损伤与ＳＧ细胞内Ｇｌｕ变化有关。     

中-低强度噪声暴露对耳蜗带状突触的影响

哺乳动物耳蜗是编码声音信号的器官,其中外毛细胞(Outer Hair Cells, OHCs)、内毛细胞(Inner Hair Cells, IHCs)与螺旋神经节神经元(Spiral Ganglion Neurons,SGNs)是将声波振动转化成神经信号的三个关键结构,其对声音暴露、耳毒性物质、衰老以及遗传缺陷等因素都很敏感。高强度的噪声暴露可以损伤耳蜗毛细胞等结构进而导致耳聋,最新研究表明,中-低强度噪声暴露可导致暂时性听阈升高,然而这种暴露常常并未引起耳蜗毛细胞的缺失或损伤;进一步的研究则发现内毛细胞和螺旋神经纤维之间的带状突触很容易受到噪声损伤,从而产生不可逆转病变,进而导致各种形式听功能的异常表现。内毛细胞带状突触(Ribbon Synapses)是听觉通路中第一个兴奋性传入突触,该结构对于声音的编码具有决定性作用。不同强度和频率的声刺激会引起带状突触在数量、结构、形态及功能方面发生相应的改变。本文通过对中-低等强度噪声暴露对小鼠耳蜗带状突触的影响以及它所引起的听觉障碍做一简要综述,希望对感音神经性耳聋患者的疾病预防有一定的指导。     

噪声对豚鼠耳蜗外淋巴Ca2+的影响及天麻素对噪声性聋的防护作用

目的探讨噪声暴露对耳蜗外淋巴Ca2 含量的影响,并观察天麻素对噪声性聋的防护作用。方法将42只健康豚鼠随机分为3组,8只为对照组,未作噪声暴露,17只为噪声暴露组,17只为噪声 天麻素组,用天麻素预防,用微量进样-火焰原子吸收光谱法测定耳蜗外淋巴Ca2 含量,检测噪声暴露前、后听性脑干反应(ABR),并与对照组比较。结果噪声暴露组耳蜗外淋巴平均Ca2 含量(113.67±42.22μg/ml)显著低于对照组(196.91±47.10μg/ml)(P<0.01),噪声 天麻素组耳蜗外淋巴平均Ca2 含量(151.15±60.65μg/ml)与对照组差异无统计学意义(P>0.05);噪声暴露后,噪声 天麻素组、噪声暴露组ABR阈值均显著升高(P<0.01),但噪声暴露组(88.53±5.08dBSPL)高于噪声 天麻素组(83.68±4.93dBSPL),差异有统计学意义(P<0.05)。结论噪声暴露可降低耳蜗外淋巴Ca2 含量,后者是噪声致聋的重要发病机理之一;天麻素能减轻噪声暴露引起的外淋巴Ca2 下降,对噪声性聋可能具有一定的预防作用。     

100dB SPL白噪声暴露对小鼠耳蜗听神经髓鞘的影响

目的观察中等强度噪声暴露是否对成年C57BL/6J小鼠耳蜗听神经造成损害。方法选取24只听阈正常的6周龄雄性C57BL/6J小鼠随机分为4组(6只/每组),实验组按照暴露后时间分别为暴露后即刻(P0)、暴露后7天(P7)和暴露后14天(P14)组,另外一组未经噪声暴露,设为对照组;实验组小鼠用100dB SPL宽频带白噪声暴露2小时,在噪声暴露后即刻、7天和14天分别检测小鼠ABR阈值,之后取小鼠耳蜗行基底膜免疫荧光观察,并进一步采用透射电镜观察小鼠听神经髓鞘结构的变化。结果 1)听力学表现:P0组C57BL/6J小鼠各个频率的ABR阈值均明显高于对照组(P<0.05),以8kHz、16kHz及32kHz最为显著(P<0.01);P7时各频率阈值基本恢复,仅在32kHz显著高于对照组(P<0.05);P14时小鼠ABR阈值与对照组相比已无显著差异(P>0.05)。2)形态学表现:经共聚焦显微镜观察发现P0时听神经髓鞘信号偶有轻微减弱,而各组标本中髓鞘信号的数量和密度无明显差异。冰冻切片显示各实验组与对照组相比,髓鞘信号均有效表达,信号强度均匀、致密连贯,条索样结构清晰,数量亦无明显缺失。透射电镜下各时间点听神经髓鞘成像清晰,排列整齐,且板层样结构致密,边缘光滑,与对照组相比髓鞘形态上并无明显差异(P>0.05)。结论 100dB SPL宽频带白噪声暴露后可造成小鼠出现暂时性阈移(TTS);此种噪声暴露下耳蜗听神经髓鞘并未观察到明显的形态学改变。     

Inhibition of the c-Jun N-terminal kinase signaling pathway influences neurite outgrowth of spiral ganglion neurons in vitro

OBJECTIVES: Inhibitors of the c-Jun N-terminal kinase (JNK) signaling pathway have been demonstrated to protect hair cells of the auditory system and different types of neurons from various insults, and their use for future therapeutic applications has been proposed. In the study, we evaluated the effects of inhibition of the JNK pathway on process outgrowth from spiral ganglion neurons. METHODS: Spiral ganglion explants from rats (postnatal days 3-5) that were cultured on laminin were treated with neurotrophin-3 and/or the JNK signaling pathway inhibitor CEP-11004. Both neurite length and number of the explants were evaluated and statistically analyzed by analysis of variance. RESULTS: Inhibition of the JNK signaling pathway reduced process outgrowth from spiral ganglion explants. The reduction, both in length and number of neurites, was reversed by the application of neurotrophin-3. CONCLUSIONS: The results indicate that an intact JNK signaling pathway is important for process outgrowth of spiral ganglion neurons. However, neurotrophin-3 stimulates process extension by a JNK independent pathway. Our results demonstrate that inhibition of the JNK pathway can have adverse effects on the extension of spiral ganglion neurons, but that the negative effects can be ameliorated by appropriate treatment.     

The protective effect of olfactory ensheathing cells on post-injury spiral ganglion cells

Conclusions: Transplantation of OECs into the cochlea may protect and increase the survival of SGCs.

Objective: To investigate the protective effect of the transplantation of olfactory ensheathing cells (OECs) on injured spiral ganglion cells (SGCs) in rats.

Methods: OECs were transplanted into the cochlea in rats with SGCs that were injured by kanamycin sulfate (KM). An equal volume of D-Hanks was injected into the cochlea of control rats. Auditory brainstem responses (ABRs) were recorded from the rats in both groups to monitor changes in hearing thresholds. Immunofluorescence was employed to examine the density and morphology of SGCs to assess the ototoxic condition of the cochlea.

Results: There was no significant difference in the ABR threshold at each frequency between the control and experimental groups. Notably, in the experimental group, a number of Hoechst 3334-labeled nuclei were detected from the apex to the basal turn of the cochlea, demonstrating that the OECs were successfully transplanted and survived in the cochlea. In the experimental group, most of the SGCs were tightly arranged, and the nuclear membrane, chromatin, and nucleolus were all clear. The SGCs in the control group were loosely arranged, and only a few normal SGCs were observed in this group.     

Qualitative and quantitative observations of spiral ganglion development in the rat

The postnatal development of the spiral ganglion cells in the rat was studied from birth until the adult stage. At birth, a single population of ganglion cells is present. Some of them are surrounded by one or two layers of satellite cell processes. With maturation, the satellite cell processes increase in number around the cell body and its processes. At the end of the first postnatal week, two important events occur. The first is the appearance of myelin lamellae between the 4th and the 6th postnatal day in both ganglion cell processes, and between the 6th and the 8th day in the cell body. The second event is the appearance of a new type of cell (the Type II spiral ganglion cell) on the 6th to the 8th day postpartum. At this stage, the Type II cell is mainly characterized by densely packed neurofilamentous structures in the cytoplasm. Comparison between the myelination of the cell body and its processes reveals three main differences! There is a time lag of approximately 2 days between the onset of myelination in the cell body and in its processes. The kinetics of myelination are different in the cell processes and in the cell body. The myelination of the cell body starts slowly, whereas it is very fast in the processes. Later, the kinetics of myelination decrease in the processes, and increase in the cell body. At all stages including the adult, the fibers have a myelin sheath composed of more lamellae than the cell body. These observations are discussed with respect to development in other species.     

Salicylate enhances expression and function of NMDA receptors in cochlear spiral ganglion neurons

Objective To study the effect of salicylate on the expression and function of NMDA receptors in spiral ganglion neurons (SGNs). Methods The mRNA of NR1 subunit of NMDA receptor in modiolus tissues were detected by Real time fluorescence quantitative PCR (FQ-PCR). NMDA receptor whole-cell currents were recorded using patch clamp in acute isolated SGNs. Results Compared with the control group, salicylate significantly increased the mRNA level of NR1 subunit in SGNs. NMDA of concentrations ranging from 0.1 mM to 10 mM evoked no current in SGNs. NMDA (0.1mM and 0.5 mM) applied with salicylate (5 mM), however, induced inward currents (212.6±15.2pA, n=5; 607.9±44.3pA, n=5) in a dose-dependent manner, which could be inhibited by APV. Salicylate alone did not produce any current in SGNs. Conclusion Salicylate increases the expression of NMDA receptors and facilitates the currents mediated by NMDA receptors in SGNs.     

Responses from AVCN units in the cat before and after inducement of an acute noise trauma

Acoustically evoked responses of single units in the anteroventral part of the cochlear nucleus (AVCN) in the cat were studied together with compound eighth-nerve action potentials (AP). Halfway through the experiments the cats were exposed for half an hour to pink noise at 105 dB SPL, producing an average threshold shift of 30 dB (maximum 50 dB) in the 2-6 kHz region. The effect of noise exposure was studied in two ways. On the one hand we compared the results for one population of units measured before the noise exposure with those found for another population measured afterwards. On the other hand we compared the results before and after the noise exposure for one unit that could be kept under observation during the noise exposure. After the noise exposure spontaneous activity and phase-locking of the responses of the units to the stimulus waveform were not significantly different from the pre-exposure findings. Response latency tended to increase. Therefore, the decrease of latency found for APs must be due to a shift to higher frequencies of the population of units contributing to the AP. The sharply tuned tip segments of tuning curves shift to higher levels whereas the low-frequency tails remain at about the same level. Q10 decreases by at most 50%, which was also found for AP tuning curves. Response spectra for clicks and noise (reverse correlation function) did not show a significant decrease of frequency selectivity. Units with CF less than 3 kHz may show a shift of CF to a lower frequency by 10-20%. After inducement of the noise trauma the sharply tuned tip segment of a tuning curve may not be found and CF may be assigned to a local minimum in the low-frequency tail of the tuning curve.     

Src家族调节Nay1．1在耳蜗螺旋神经节神经元的表达

目的：研究蛋白酪氨酸激酶Src家族对耳蜗螺旋神经节神经元的电压门控钠离子通道基因和蛋白表达的影响。方法：采用RT—PCR和Western blot技术分别观察在Src家族抑制剂的作用下,Nav1．1mRNA和蛋白的表达变化情况。结果：Src家族抑制剂PP2（10μmol／L）和SU6656（2μmol／L）作用后,螺旋神经节神经元Nav1．1mRNA的表达量分别减少至26％±0．8％和36％±1．5％（P〈0．05）,蛋白的表达量分别减少至39％±12．5％和53％±1．7％（P〈0．05）。结论：抑制蛋白酪氨酸激酶Src家族可下调Nav1．1mRNA和蛋白在螺旋神经节神经元的表达水平。     

The spiral ganglion and hair cells of Bronx waltzer mice

Inner hair cells and spiral ganglion cells were counted in a mutant mouse strain (Bronx waltzer) reported to have very few of these cells (6) in order to determine if the remaining ganglion cells would be predominantly type II cells. These cell counts indicate a 50% reduction of spiral ganglion cells in Bronx waltzer cochleas compared to normal mice. Averaged throughout the cochlea about 11% of the remaining cells are type II cells while in normal mice this percentage is 5%. In some regions however, as many as 20% of the remaining cells are type II cells. Counts of IHC in surface preparations reveal 37 normal looking IHC (about 5% of the normal population) in each of two Bronx waltzer mouse cochleas. There were also about 120 shrunken IHC in each cochlea, representing 17% of the normal cell population. While there appears to be an increased proportion of type II cells in the Bronx waltzer spiral ganglion there are also many more type I cells than might be expected from the small number of IHC.     

The influence of moderate-intensity noise on the compound action potential evoked by tone bursts in the guinea pig, Cavia porcellus

Noise-induced changes in the compound action potential (CAP) evoked by tone bursts in the frequency range 0.5-24 kHz were studied in 15 pigmented guinea pigs by means of chronically implanted electrodes positioned near the round window. The animals were exposed for 120 h to continuous pink noise at the intensities 80, 90 and 100 dB SPL. During the exposure period, all the animals exhibited an exponential rise in CAP threshold, leveling out after 24-72 h (asymptotic threshold shift, ATS). The largest threshold shifts were recorded during exposure to 100 dB SPL, for frequencies in the range 8-12 kHz. In the recovery phase, after the end of noise exposure, the threshold to tones at all frequencies tested fell exponentially, reaching the original level in about 72 h in all cases.     

The spiral ganglion and cochlear nuclei of deafness mice

Deafness mice (dn/dn) never develop hearing, but, except for the associated physical defects, have no other known abnormalities. For this study, cochleas and brains of five adult homozygotes (dn/dn) and five adult heterozygotes (+/dn), matched by weight and sex, were prepared for serial section light microscopy. In the homozygotes, the organ of Corti was totally degenerated basally, gradually improving toward the apex where supporting cells, border cells and pillar cells were present; however, the stria vascularis was dystrophic in the apex. The saccular macula was atrophied or dystrophic in seven of the ten homozygote ears. The homozygotes had only 23% of the number of spiral ganglion cells found in the heterozygotes, but they appeared robust. In six of the ears of each group there was clumping of apical spiral ganglion neurons. In the homozygotes, the volume of Rosenthal's canal was 121%, that of the dorsal nuclei was 90%, and that of the ventral cochlear nuclei was 63% of the comparable volumes in heterozygotes. The globular cells of the homozygote ventral cochlear nucleus were 72% of the size of those of heterozygotes. These quantitative morphological abnormalities of the homozygote spiral ganglion and cochlear nuclei may result from organ of Corti atrophy.