Objective To study presynaptic alternations of cochlear ribbons arising from aminoglycoside ototoxic stimuli in C57BL/6J mice. Methods Animals were injected with low dose gentamicin (100 mg/kg/day) for 14 days, From t...Objective To study presynaptic alternations of cochlear ribbons arising from aminoglycoside ototoxic stimuli in C57BL/6J mice. Methods Animals were injected with low dose gentamicin (100 mg/kg/day) for 14 days, From the 14th to 28th days, the mice were maintained free of gentamicin treatment. Immunohisto-chemistry labeling was employed to trace RIBEYE, a major presynaptic componment of ribbon synapses. RIBEYE/CtBP2 expression levels were assessed and compared with hearing threshold shifts. Auditory func-tion was assessed by auditory brainstem responses. The stereocilia of outer hair cells (OHCs) and IHCs was examined by scanning electron microscopy (SEM). Results Hearing thresholds were elevated with peak hearing loss observed on the 7th day after gentamicin exposure, followed by improvement after the 7th day. RIBEYE/CtBP2 expression directly correlated with observed hearing threshold shifts. Strikingly, we did not see any obvious changes in stereocilia in both OHCs and IHCs until the 28th day. Mild changes in stereocil-ia were only observed in OHCs on the 28th day. Conclusions These findings indicate that presynapse co-chlear ribbons, rather than stereocilia, may be sensitive to aminoglycoside ototoxic exposure in mice cochle-ae. A pattern of RIBEYE/CtBP2 expression changes seems to parallel hearing threshold shifts and suggests presynaptic response properties to lower dosage of aminoglycoside ototoxic stimuli.展开更多
目的通过比较噪声暴露后基底膜不同区域内毛细胞(IHCs)带状突触损伤差异,探讨带状突触损伤易感性的相关因素。方法将28只C57BL/6J雄性小鼠随机分为噪声暴露组和对照组,每组14只。噪声暴露组小鼠给予强度103 dB SPL、频率2~20 kHz、持续...目的通过比较噪声暴露后基底膜不同区域内毛细胞(IHCs)带状突触损伤差异,探讨带状突触损伤易感性的相关因素。方法将28只C57BL/6J雄性小鼠随机分为噪声暴露组和对照组,每组14只。噪声暴露组小鼠给予强度103 dB SPL、频率2~20 kHz、持续2 h的宽带噪声暴露,对照组小鼠则饲养于安静环境中。噪声暴露前及噪声暴露后第一天进行ABR测试及毛细胞带状突触免疫荧光染色实验。使用全细胞膜片钳技术比较不同区域IHCs的钙离子流入。通过免疫荧光染色比较噪声暴露后的耳蜗基底膜顶回、中回、底回IHCs钙蛋白酶(Calpain)表达水平,并用蛋白质印迹实验验证钙蛋白酶对IHCs带状突触蛋白CtBP2的损伤作用。结果噪声暴露后一天,噪声暴露组在11.3、16.0、22.6、32.0 kHz的ABR阈值较对照组显著上升(均为P<0.001),中回、底回IHCs带状突触数量明显减少(P<0.05)。全细胞膜片钳实验结果表明耳蜗基底膜中回IHCs有较多的钙离子通道(P<0.01),但其单通道电流较小(P<0.01),顶回、中回IHCs钙离子通道开放率无显著差异(P>0.05)。噪声暴露后,耳蜗基底膜中回、底回IHCs的Calpain表达水平显著高于顶回(P<0.001),蛋白质印迹实验结果表明Calpain以钙离子依赖的方式降解带状突触蛋白CtBP2。结论钙蛋白酶是基底膜高频区内毛细胞带状突触噪声损伤易感的重要因素。展开更多
目的建立军事航空噪声性隐匿性听力损失(HHL)C57小鼠动物模型,并对其有效性进行评价,为军事航空噪声性HHL的研究提供稳定的小鼠动物模型构建方法。方法将听力正常的雄性C57BL/6J小鼠随机分成4组,分别给予不同强度的噪声暴露:105 dB 2 h,...目的建立军事航空噪声性隐匿性听力损失(HHL)C57小鼠动物模型,并对其有效性进行评价,为军事航空噪声性HHL的研究提供稳定的小鼠动物模型构建方法。方法将听力正常的雄性C57BL/6J小鼠随机分成4组,分别给予不同强度的噪声暴露:105 dB 2 h,110 dB 2 h,110 dB 4 h,115 dB 4 h。将噪声暴露前的小鼠作为对照组,根据噪声暴露后不同时间分为噪声暴露后1 d组(NE-1 d)、噪声暴露后7 d组(NE-7 d)、噪声暴露后14 d组(NE-14 d)和噪声暴露后28 d组(NE-28 d)。各实验组进行相应强度的军用直升机噪声暴露,并在各时间节点进行听性脑干反应(ABR)测试。筛选出符合条件的噪声暴露强度,并进行该条件噪声暴露后基底膜的免疫荧光染色和带状突触计数验证。结果105 dB 2 h军用直升机噪声暴露后C57小鼠听阈阈移不明显;110 dB 4 h和115 dB 4 h军用直升机噪声暴露后小鼠听阈发生永久性阈移;110 dB 2 h军用直升机噪声暴露后,C57小鼠听阈暂时性阈移,ABR波Ⅰ幅值降低,耳蜗带状突触数量降低,满足HHL的功能学和形态学要求。结论110 dB 2 h军用直升机噪声暴露可作为军事航空噪声性HHL C57小鼠模型的理想刺激参数。展开更多
The development of auditory synapses is a key process for the maturation of heating function. However, it is still on debate regaralng wnemer the development of auditory synapses is dominated by acquired sound stimula...The development of auditory synapses is a key process for the maturation of heating function. However, it is still on debate regaralng wnemer the development of auditory synapses is dominated by acquired sound stimulations. In this review, we summarize relevant publications in recent decades to address this issue. Most reported data suggest that extrinsic sound stimulations do affect, but not govern the development of periphery auditory synapses. Overall, periphery auditory synapses develop and mature according to its intrinsic mechanism to build up the synaptic connections between sensory neurons and/or interneurons.展开更多
目的探讨2型糖尿病对雄性小鼠听力脑干反应(ABR)和耳蜗带状突触的作用。方法将24只健康6周龄清洁级雄性C57BL/6J小鼠按体重随机分为2组,分别为对照[饲料含铁量为(8.26±2.67)mg/kg]组和糖尿病[饲料含铁量为(8.39±1.03)g/kg]组...目的探讨2型糖尿病对雄性小鼠听力脑干反应(ABR)和耳蜗带状突触的作用。方法将24只健康6周龄清洁级雄性C57BL/6J小鼠按体重随机分为2组,分别为对照[饲料含铁量为(8.26±2.67)mg/kg]组和糖尿病[饲料含铁量为(8.39±1.03)g/kg]组,每组12只。分别于喂养第0、4、8、12、16周监测血糖,第16周测定血红蛋白、血清铁、血清铁蛋白、总铁结合力、转铁蛋白饱和度,并进行葡萄糖耐量及胰岛素耐量测验,采用ABR检测。应用免疫荧光检测左侧耳蜗基底膜C末端结合蛋白2(Ct BP2)、囊泡谷氨酸转运体3(VGLUT3)、肌球蛋白Ⅶa(MyosinⅦa)在耳蜗内外毛细胞分布,分离右侧耳蜗进行HE染色。结果与对照组比较,糖尿病组小鼠血清铁、血清铁蛋白、转铁蛋白饱和度均升高,差异有统计学意义(P<0.05);而血红蛋白、总铁结合力均无明显改变。与对照组比较,糖尿病组小鼠血糖水平在喂养8周后明显升高,喂养16周时胰岛素抵抗指数(HOMA-IR)明显升高,差异均有统计学意义(P<0.05)。与对照组比较,糖尿病组小鼠血糖水平在葡萄糖耐量试验的60、120 min时较高,差异有统计学意义(P<0.05);而在15、30 min时仅略有升高。与对照组比较,胰岛素耐量实验各检测时间点糖尿病组小鼠血糖水平均较高,差异有统计学意义(P<0.05)。与对照组比较,糖尿病组小鼠在2、3 k Hz下的ABR阈值较高,差异均有统计学意义(P<0.05);而在1、4 k Hz下的ABR阈值仅略有升高。糖尿病可造成小鼠内毛细胞Ct BP2标记带状突触数量减少,但在耳蜗毛细胞中VGLUT3、MyosinⅦa免疫荧光阳性反应未受影响,另外,耳蜗Corti氏器、螺旋神经节细胞及神经丝未见损伤。结论膳食铁过量诱导的2型糖尿病小鼠可能出现内耳毛细胞带状突触数量减少,从而扰乱小鼠听力脑干反应,呈现高频段小鼠听力损伤。展开更多
Background Previous studies have suggested that primary degeneration of hair cells causes secondary degeneration of spiral ganglion neurons （SGNs）,but the effect of SGN degeneration on hair cells has not been studie...Background Previous studies have suggested that primary degeneration of hair cells causes secondary degeneration of spiral ganglion neurons （SGNs）,but the effect of SGN degeneration on hair cells has not been studied.In the adult mouse inner ear ouabain can selectively and permanently induce the degeneration of type 1 SGNs while leaving type 2 SGNs,efferent fibers,and sensory hair cells relatively intact.This study aimed to investigate the dynamic changes in hair cell ribbon synapse induced by loss of SGNs using ouabain application to the round window niche of adult mice.Methods In the analysis,24 CBA/CAJ mice aged 8-10 weeks,were used,of which 6 normal mice were used as the control group.After ouabain application in the round window niche 6 times in an hour,ABR threshold shifts at least 30 dB in the three experimental groups which had six mice for 1-week group,six for 1-month group,and six for 3-month group.All 24 animals underwent function test at 1 week and then immunostaining at 1 week,1 month,and 3 months.Results The loss of neurons was followed by degeneration of postsynaptic specializations at the afferent synapse with hair cells.One week after ouabain treatment,the nerve endings of type 1 SGNs and postsynaptic densities,as measured by Na/K ATPase and PSD-95,were affected but not entirely missing,but their partial loss had consequences for synaptic ribbons that form the presynaptic specialization at the synapse between hair cells and primary afferent neurons.Ribbon numbers in inner hair cells decreased （some of them broken and the ribbon number much decreased）,and the arrangement of the synaptic ribbons had undergone a dynamic reorganization：ribbons with or without associated postsynaptic densities moved from their normal location in the basal membrane of the cell to a more apical location and the neural endings alone were also found at more apical locations without associated ribbons.After 1 month,when the neural postsynaptic densities had completed their degeneration,most ribbons were lost and 展开更多
目的利用三维建模的方法对耳蜗内毛细胞带状突触进行计数分析,解决由于内毛细胞带状突触数量少、位置深在造成的计数困难,为内毛细胞带状突触的可塑性研究提供有效可靠的计数方法。方法取小鼠耳蜗基底膜,使用免疫荧光双标记的方法对突触...目的利用三维建模的方法对耳蜗内毛细胞带状突触进行计数分析,解决由于内毛细胞带状突触数量少、位置深在造成的计数困难,为内毛细胞带状突触的可塑性研究提供有效可靠的计数方法。方法取小鼠耳蜗基底膜,使用免疫荧光双标记的方法对突触前 RIBEYE 和突触后膜 GluR2&3 进行标记,激光共聚焦显微镜进行光学连续切片,每个荧光色对代表一个突触的存在。使用 3ds max 进行三维建模,对内毛细胞带状突触进行计数。结果耳蜗基底膜内毛细胞的带状突触显示清晰,每个内毛细胞的带状突触数量为(16.10±1.03)个。结论利用免疫荧光双标,激光共聚焦显微镜光学连续切片,3ds max 三维建模所得的内毛细胞带状突触数量准确,方法简单可行,是一种对内毛细胞带状突触进行计数的可靠方法。展开更多
基金supported by grants from the National Basic Research Program of China(973Program)(2012CB9679002012CB967901)+3 种基金Beijing Natural Science Foundation(5122040)supported by grants from the China Postdoctoral Science Foundation(20100377920100470103)the National Natural Science Foundation of China(NSFC)(31040 038)
文摘Objective To study presynaptic alternations of cochlear ribbons arising from aminoglycoside ototoxic stimuli in C57BL/6J mice. Methods Animals were injected with low dose gentamicin (100 mg/kg/day) for 14 days, From the 14th to 28th days, the mice were maintained free of gentamicin treatment. Immunohisto-chemistry labeling was employed to trace RIBEYE, a major presynaptic componment of ribbon synapses. RIBEYE/CtBP2 expression levels were assessed and compared with hearing threshold shifts. Auditory func-tion was assessed by auditory brainstem responses. The stereocilia of outer hair cells (OHCs) and IHCs was examined by scanning electron microscopy (SEM). Results Hearing thresholds were elevated with peak hearing loss observed on the 7th day after gentamicin exposure, followed by improvement after the 7th day. RIBEYE/CtBP2 expression directly correlated with observed hearing threshold shifts. Strikingly, we did not see any obvious changes in stereocilia in both OHCs and IHCs until the 28th day. Mild changes in stereocil-ia were only observed in OHCs on the 28th day. Conclusions These findings indicate that presynapse co-chlear ribbons, rather than stereocilia, may be sensitive to aminoglycoside ototoxic exposure in mice cochle-ae. A pattern of RIBEYE/CtBP2 expression changes seems to parallel hearing threshold shifts and suggests presynaptic response properties to lower dosage of aminoglycoside ototoxic stimuli.
文摘目的建立军事航空噪声性隐匿性听力损失(HHL)C57小鼠动物模型,并对其有效性进行评价,为军事航空噪声性HHL的研究提供稳定的小鼠动物模型构建方法。方法将听力正常的雄性C57BL/6J小鼠随机分成4组,分别给予不同强度的噪声暴露:105 dB 2 h,110 dB 2 h,110 dB 4 h,115 dB 4 h。将噪声暴露前的小鼠作为对照组,根据噪声暴露后不同时间分为噪声暴露后1 d组(NE-1 d)、噪声暴露后7 d组(NE-7 d)、噪声暴露后14 d组(NE-14 d)和噪声暴露后28 d组(NE-28 d)。各实验组进行相应强度的军用直升机噪声暴露,并在各时间节点进行听性脑干反应(ABR)测试。筛选出符合条件的噪声暴露强度,并进行该条件噪声暴露后基底膜的免疫荧光染色和带状突触计数验证。结果105 dB 2 h军用直升机噪声暴露后C57小鼠听阈阈移不明显;110 dB 4 h和115 dB 4 h军用直升机噪声暴露后小鼠听阈发生永久性阈移;110 dB 2 h军用直升机噪声暴露后,C57小鼠听阈暂时性阈移,ABR波Ⅰ幅值降低,耳蜗带状突触数量降低,满足HHL的功能学和形态学要求。结论110 dB 2 h军用直升机噪声暴露可作为军事航空噪声性HHL C57小鼠模型的理想刺激参数。
基金supported by grants from the National Basic Research Program of China(973Program)(2012CB9679002012CB967901)+3 种基金Beijing Natural Science Foundation(5122040)supported by grants from the China Postdoctoral Science Foundation(20100377920100470103)the National Natural Science Foundation of China(NSFC)(31040038)
文摘The development of auditory synapses is a key process for the maturation of heating function. However, it is still on debate regaralng wnemer the development of auditory synapses is dominated by acquired sound stimulations. In this review, we summarize relevant publications in recent decades to address this issue. Most reported data suggest that extrinsic sound stimulations do affect, but not govern the development of periphery auditory synapses. Overall, periphery auditory synapses develop and mature according to its intrinsic mechanism to build up the synaptic connections between sensory neurons and/or interneurons.
文摘目的探讨2型糖尿病对雄性小鼠听力脑干反应(ABR)和耳蜗带状突触的作用。方法将24只健康6周龄清洁级雄性C57BL/6J小鼠按体重随机分为2组,分别为对照[饲料含铁量为(8.26±2.67)mg/kg]组和糖尿病[饲料含铁量为(8.39±1.03)g/kg]组,每组12只。分别于喂养第0、4、8、12、16周监测血糖,第16周测定血红蛋白、血清铁、血清铁蛋白、总铁结合力、转铁蛋白饱和度,并进行葡萄糖耐量及胰岛素耐量测验,采用ABR检测。应用免疫荧光检测左侧耳蜗基底膜C末端结合蛋白2(Ct BP2)、囊泡谷氨酸转运体3(VGLUT3)、肌球蛋白Ⅶa(MyosinⅦa)在耳蜗内外毛细胞分布,分离右侧耳蜗进行HE染色。结果与对照组比较,糖尿病组小鼠血清铁、血清铁蛋白、转铁蛋白饱和度均升高,差异有统计学意义(P<0.05);而血红蛋白、总铁结合力均无明显改变。与对照组比较,糖尿病组小鼠血糖水平在喂养8周后明显升高,喂养16周时胰岛素抵抗指数(HOMA-IR)明显升高,差异均有统计学意义(P<0.05)。与对照组比较,糖尿病组小鼠血糖水平在葡萄糖耐量试验的60、120 min时较高,差异有统计学意义(P<0.05);而在15、30 min时仅略有升高。与对照组比较,胰岛素耐量实验各检测时间点糖尿病组小鼠血糖水平均较高,差异有统计学意义(P<0.05)。与对照组比较,糖尿病组小鼠在2、3 k Hz下的ABR阈值较高,差异均有统计学意义(P<0.05);而在1、4 k Hz下的ABR阈值仅略有升高。糖尿病可造成小鼠内毛细胞Ct BP2标记带状突触数量减少,但在耳蜗毛细胞中VGLUT3、MyosinⅦa免疫荧光阳性反应未受影响,另外,耳蜗Corti氏器、螺旋神经节细胞及神经丝未见损伤。结论膳食铁过量诱导的2型糖尿病小鼠可能出现内耳毛细胞带状突触数量减少,从而扰乱小鼠听力脑干反应,呈现高频段小鼠听力损伤。
文摘Background Previous studies have suggested that primary degeneration of hair cells causes secondary degeneration of spiral ganglion neurons （SGNs）,but the effect of SGN degeneration on hair cells has not been studied.In the adult mouse inner ear ouabain can selectively and permanently induce the degeneration of type 1 SGNs while leaving type 2 SGNs,efferent fibers,and sensory hair cells relatively intact.This study aimed to investigate the dynamic changes in hair cell ribbon synapse induced by loss of SGNs using ouabain application to the round window niche of adult mice.Methods In the analysis,24 CBA/CAJ mice aged 8-10 weeks,were used,of which 6 normal mice were used as the control group.After ouabain application in the round window niche 6 times in an hour,ABR threshold shifts at least 30 dB in the three experimental groups which had six mice for 1-week group,six for 1-month group,and six for 3-month group.All 24 animals underwent function test at 1 week and then immunostaining at 1 week,1 month,and 3 months.Results The loss of neurons was followed by degeneration of postsynaptic specializations at the afferent synapse with hair cells.One week after ouabain treatment,the nerve endings of type 1 SGNs and postsynaptic densities,as measured by Na/K ATPase and PSD-95,were affected but not entirely missing,but their partial loss had consequences for synaptic ribbons that form the presynaptic specialization at the synapse between hair cells and primary afferent neurons.Ribbon numbers in inner hair cells decreased （some of them broken and the ribbon number much decreased）,and the arrangement of the synaptic ribbons had undergone a dynamic reorganization：ribbons with or without associated postsynaptic densities moved from their normal location in the basal membrane of the cell to a more apical location and the neural endings alone were also found at more apical locations without associated ribbons.After 1 month,when the neural postsynaptic densities had completed their degeneration,most ribbons were lost and
文摘目的利用三维建模的方法对耳蜗内毛细胞带状突触进行计数分析,解决由于内毛细胞带状突触数量少、位置深在造成的计数困难,为内毛细胞带状突触的可塑性研究提供有效可靠的计数方法。方法取小鼠耳蜗基底膜,使用免疫荧光双标记的方法对突触前 RIBEYE 和突触后膜 GluR2&3 进行标记,激光共聚焦显微镜进行光学连续切片,每个荧光色对代表一个突触的存在。使用 3ds max 进行三维建模,对内毛细胞带状突触进行计数。结果耳蜗基底膜内毛细胞的带状突触显示清晰,每个内毛细胞的带状突触数量为(16.10±1.03)个。结论利用免疫荧光双标,激光共聚焦显微镜光学连续切片,3ds max 三维建模所得的内毛细胞带状突触数量准确,方法简单可行,是一种对内毛细胞带状突触进行计数的可靠方法。
