小鼠螺旋神经节细胞电压依赖性钙通道亚单位分析

Subunit of voltage-dependent calcium channels in the murine spiral ganglion cells

目的 分析小鼠螺旋神经节细胞电压依赖性钙通道亚单位的类型。方法 手术分离新生小鼠的耳蜗螺旋神经节细胞进行培养,24 h后提取RNA。逆转录后获得螺旋神经节细胞cDNA,根据电压依赖性钙通道7种亚单位序列设计的引物进行聚合酶链反应扩增,通过聚合酶链反应结果分析和DNA测序确定螺旋神经节细胞表达的亚单位类型。结果 逆转录聚合酶链反应扩增出α1D、α1E、α2/δ、β1和β3亚单位的片段,测序进一步证明小鼠螺旋神经节细胞中有这几种亚单位的存在。结论 小鼠螺旋神经节细胞中有α1D、α1E、α2/δ、β1和β3亚单位的表达。α1D和α1E亚单位的共表达证明在哺乳动物的螺旋神经节细胞中有L型和R型电压依赖性钙通道。

Objective To investigate subunit type of voltage-dependent calcium channels in the spiral ganglion cells of the mouse. Methods The spiral ganglion cells were dissected from cochleae of neonatal mice and cultured for 24 h. Total RNA was extracted from cultured spiral ganglion cells. After reverse transcription, resulting cDNA was amplified by polymerase chain reaction (PCR) with primers targeted to nucleotide sequences corresponding to 7 different calcium channel subunits. The types of calcium channel subunits were identified by PCR analysis and nucleotide sequencing. Results Reverse transcription (RT)-PCR products representing subunit gene expression were strongly and consistently amplified for α1D, α1E, α2/δ, β1 and β3 Nucleotide sequencing confirmed the identity of mouse cochlear subunit cDNAs. Conclusions α1D, α1E, α2/8δ, β1 and β3 subunits are expressed in spiral ganglion cells. And the coexpression of α1 D and α1 E demonstrate the presence of L-type and R-type calcium channels in mammalian spiral ganglion cells.