Cell Biotechnology and Molecular Biology of the Isolated Spiral Ganglion Cells in the Ear with Profound Hearing Loss

严重听力损失耳中分离螺旋神经节细胞的细胞生物技术和分子生物学

• 批准号: 09671731
• 负责人: KAWASE Tetsuaki
• 金额: $ 1.92万
• 依托单位: Tohoku University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09671731/
• 关键词: spiral ganglion; Ca^<2+>; hearing loss; LIVE and DEAD Viability; Cytotoxity Assay; Fluo-3 and Fura-Red; LASER-scanning microscope

Intracellular Ca^<2+> plays an important role in the cellular functions of various tissues. In the spiral ganglion cells, neurotransmitter release is known to be dependent on cytosolic Ca^<2+> responses through voltage-dependent Ca^<2+> channels. We examined cytosolic Ca^<2+> responses induced by depolarization in the isolated spiral ganglion cells of the kanamycin-intoxicated guinea-pig to evaluate the activity of theCa^<2+> channel in pathological conditions.Albino guinea-pigs (weight 250-250g) received intramuscular injections of 400 mg/kgkanamycin 5 times per week for 3 weeks. Guinea pigs without treatment were used as controls. Control guinea-pigs showed normal ABR responses. On the other hand, kanamycin-intoxicatedguinea pigs showed no detectable responses below 65 dB SPL evoked by click. Spiral ganglion cells of the guinea-pig cochlea were mechanically isolated. Dynamic changes in intracellular Ca^<2+>concentration were estimated with Fluo-3/Fura-Red ratiometric method using a LASER-scanning microscope. Activation of the Ca^<2+> channel was obtained by high K+-induced depolarization.High K+-induced deporalization elevated the cytosolic Ca^<2+> concentration of the ganglion cells. Some of the cells showed oscillatory increases in the cytosolic Ca^<2+> level. Furthermore, the cytosolicCa^<2+> response induced by depolarization was decreased in damaged spiral ganglion cells compared with that in control cells.

细胞内Ca ^2+在各种组织的细胞功能中起重要作用。在螺旋神经节细胞中，已知神经递质的释放依赖于通过电压依赖性Ca ^2+通道的胞质Ca ^2+反应。我们检测了卡那霉素中毒豚鼠螺旋神经节细胞去极化引起的胞浆Ca ^<2 +>反应，以评估病理状态下Ca ^<2 +>通道的活性。白化病豚鼠（体重250 - 250 g）每周5次肌肉注射400 mg/kg卡那霉素，共3周。未处理的豚鼠用作对照。对照组豚鼠ABR反应正常。而卡那霉素中毒豚鼠在65 dB SPL以下的短声诱发下无明显反应。机械分离豚鼠耳蜗螺旋神经节细胞。用Fluo-3/Fura-Red比值法在激光扫描显微镜下测定细胞内Ca ^<2 +>浓度的动态变化。高K+诱导的去极化可激活Ca ^<2 +>通道，高K+诱导的去孔作用可提高神经节细胞胞浆Ca ^<2 +>浓度。有些细胞胞浆Ca ^2+水平呈振荡性升高。此外，与对照组相比，受损螺旋神经节细胞由去极化引起的胞浆Ca ^2+反应降低。

项目成果

Obara Y,Kawase T et.al.: "Cytosolic Ca2+ response induced by depolarization in the isolated spiral ganglion cells of the kanamycin-intoxicated guinea-pig" VIIth Korea-Japan joint meeting of Otoarhinolaryngology-head and neck surgery. 60- (1998)

Obara Y、Kawase T 等人：“卡那霉素中毒豚鼠的分离螺旋神经节细胞去极化诱导的胞质 Ca2 反应”第七届韩日耳鼻咽喉头颈外科联席会议。

Kawase T,et al.: "The acoustic reflex for filtered broadband stimuli : A lesser contribution of the lower frequency neurons." Tohoku J.Exp.Med. 185. 131-137 (1998)

Kawase T 等人：“过滤宽带刺激的声反射：低频神经元的贡献较小。”

Obara Y,Kawase T et.al.: "Cytosolic Ca2+ response induced by depolarization in the isolated spiral ganglion cells of the kanamycin-intoxicated guinea-pig" VIIth Korea-Japan joint meeting of Otoarhinolaryngology-head and neck surgery. 60 (1998)

Obara Y、Kawase T 等人：“卡那霉素中毒豚鼠的分离螺旋神经节细胞去极化诱导的胞质 Ca2 反应”第七届韩日耳鼻咽喉头颈外科联席会议。