Primary Culture in Vitro and Identification of Murine Spiral Ganglion Neurons_West China Medical Journal

Authors：

LIU Tao ¹ , CHEN anfang ² , LIU Yi ³

Department of Otolaryngology,West China Hospital,Sichuan University,Chengdu 610041,China;

Corresponding?author：

Keywords：

螺旋神經元; 原代培養; 神經微絲蛋白; 免疫熒光細胞學

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目的：機械分離、培養小鼠耳蝸螺旋神經元，并進行免疫熒光細胞學鑒定，為后期進一步的實驗研究提供實驗材料。方法：采用初出生1～5天以內的昆明小鼠進行解剖、機械分離以獲得螺旋神經節組織，進行原代培養后，應用神經微絲蛋白（Neurofilament protein,NFP－H）單克隆抗體進行免疫熒光細胞學鑒定。結果：機械分離后獲得的螺旋神經節組織中的螺旋神經元，在體外培養條件下可以存活并進行正常分化。典型的螺旋神經元，其細胞形態呈橢圓形,胞體透明光滑、接近生理形態。熒光染色標記后，胞體和神經突起均顯色好，Schwann細胞和成纖維細胞未著色。結論：應用機械分離的方法獲得小鼠耳蝸螺旋神經節組織并進行培養，耳蝸螺旋神經元在體外可以穩定地存活生長。培養獲得的細胞形態和生存狀態接近生理狀態，滿足電生理、免疫細胞化學、藥理學等研究。應用特異性的神經微絲蛋白對培養獲得的螺旋神經元進行免疫熒光細胞學鑒定，特異性好，熒光顯色好。

Citation： LIU Tao,CHEN anfang,LIU Yi,et al.. Primary Culture in Vitro and Identification of Murine Spiral Ganglion Neurons. West China Medical Journal, 2008, 23(1): 104-105. doi: Copy

1.	Bianchi LN,Cohan CS.Developmental regulation of neurite-promoting factorinfliuencing statoacoustic neurons[Ｊ].Devel Brain Res,1991,64:167－74..
2.	李剛，唐癑秌，程薇波．應用LsAB技術顯示五羥色胺受體亞型1A在小鼠螺旋神經元上的分布[Ｊ]．聽力學及言語疾病雜志，2007，15（3）：235-237．.
3.	Harrison RG.Observation on the living developing nerve fiber[Ｊ].Ant Rec，1907,1:116．.
4.	Nakai J.Dissociated dorsal root ganglia in tissueculure[Ｊ].Am J Anat,1956,99(1):81．.
5.	黃世楷，等，譯．神經元形態與功能[Ｍ]．上海科技出版社，1986:6．.
6.	Kimura K,Mat sumoto N,Kitada M,et al.Neurite out growth from hippocampal neurons is promoted by choroid plexus ependymal cells in vit ro[Ｊ].J Neurocytol,2004,33:465-476..
7.	Deumens R,Koopmans G C,Den Bakker C G,et al.Alignment of glial cells stimulates directional neurite growth of CNS neurons in vitro[Ｊ].Neuroscience,2004,125:591-604..
8.	Gallo F,Morale M C,Spina2Purrello V,et al.Basic fibroblast growth factor(bFGF)act s on both neurons and glia to mediate the neurot rophic effect s of ast rocytes on L HRH neurons in culture[Ｊ].Synapse,2000,36:233-253..
9.	Chakrabort ty S,Kitada M,Mat sumoto N,et al.Choroid plexus ependymal cells enhance neurite outgrowth f rom dorsal root ganglion neurons in vitro[Ｊ].J Neurocy2tol,2000,29:707-717..
10.	Zhiping Zhou,Qing Liu,and Robin L.Davis.Complex regulationof spiral ganglion neuron firing patterns by neurotrophin-3[Ｊ].The Journal of Neurosicence,August 17,2005-25(33):7558-7566..
11.	徐定遠，劉世喜，劉進．內皮素1在豚鼠耳蝸螺旋神經節的分布[Ｊ]．臨床耳鼻咽喉頭頸外科雜志，2007，21（4）：176-177．.
12.	Hoorens A,Pipeleers D.Nicotinamide protents human beta cells against chemically induced necrosis,but not against cytokine induced apoptosis[Ｊ].Diabetologia,1999,42:55-59．.
13.	孫建軍，劉鋌．螺旋神經元原代培養及其免疫細胞化學鑒定[Ｊ]．中華耳鼻咽喉科雜志，1998，33（4）：196－198．.
14.	Han DT,Yamashita T,Harada N,et al.Calcium mobilization in isolated cochlear sprail ganglion cells of the guinea pigs[Ｊ].Acta Otol,1993,Suppl 506:26-29..
15.	Ross MD.Multipolar neurons in the spiral ganglion of rat[Ｊ].Acta otolaryngol,1973,76:381-394..
16.	Ard,MD,Morest DK,Hauger SH.Trophic interactions between the cochleovestibular ganglion of the chick embryo and its synaptic targets in cultures[Ｊ].Neurosicence,1985,16:151-170..
17.	Sobkowicz HM,Loftus JM,slapnick,et al.Tissue culture of the organ of corti[Ｊ].Acta Otolaryngol,1993,502:3-36..
18.	唐癑秌，Davis Robin.應用膜片鉗技術對螺旋神經元放電類型的初步研究[Ｊ]．聽力學及言語疾病雜志，2002，10（4）：250-252．.

1. Bianchi LN,Cohan CS.Developmental regulation of neurite-promoting factorinfliuencing statoacoustic neurons[Ｊ].Devel Brain Res,1991,64:167－74..
2. 李剛，唐癑秌，程薇波．應用LsAB技術顯示五羥色胺受體亞型1A在小鼠螺旋神經元上的分布[Ｊ]．聽力學及言語疾病雜志，2007，15（3）：235-237．.
3. Harrison RG.Observation on the living developing nerve fiber[Ｊ].Ant Rec，1907,1:116．.
4. Nakai J.Dissociated dorsal root ganglia in tissueculure[Ｊ].Am J Anat,1956,99(1):81．.
5. 黃世楷，等，譯．神經元形態與功能[Ｍ]．上海科技出版社，1986:6．.
6. Kimura K,Mat sumoto N,Kitada M,et al.Neurite out growth from hippocampal neurons is promoted by choroid plexus ependymal cells in vit ro[Ｊ].J Neurocytol,2004,33:465-476..
7. Deumens R,Koopmans G C,Den Bakker C G,et al.Alignment of glial cells stimulates directional neurite growth of CNS neurons in vitro[Ｊ].Neuroscience,2004,125:591-604..
8. Gallo F,Morale M C,Spina2Purrello V,et al.Basic fibroblast growth factor(bFGF)act s on both neurons and glia to mediate the neurot rophic effect s of ast rocytes on L HRH neurons in culture[Ｊ].Synapse,2000,36:233-253..
9. Chakrabort ty S,Kitada M,Mat sumoto N,et al.Choroid plexus ependymal cells enhance neurite outgrowth f rom dorsal root ganglion neurons in vitro[Ｊ].J Neurocy2tol,2000,29:707-717..
10. Zhiping Zhou,Qing Liu,and Robin L.Davis.Complex regulationof spiral ganglion neuron firing patterns by neurotrophin-3[Ｊ].The Journal of Neurosicence,August 17,2005-25(33):7558-7566..
11. 徐定遠，劉世喜，劉進．內皮素1在豚鼠耳蝸螺旋神經節的分布[Ｊ]．臨床耳鼻咽喉頭頸外科雜志，2007，21（4）：176-177．.
12. Hoorens A,Pipeleers D.Nicotinamide protents human beta cells against chemically induced necrosis,but not against cytokine induced apoptosis[Ｊ].Diabetologia,1999,42:55-59．.
13. 孫建軍，劉鋌．螺旋神經元原代培養及其免疫細胞化學鑒定[Ｊ]．中華耳鼻咽喉科雜志，1998，33（4）：196－198．.
14. Han DT,Yamashita T,Harada N,et al.Calcium mobilization in isolated cochlear sprail ganglion cells of the guinea pigs[Ｊ].Acta Otol,1993,Suppl 506:26-29..
15. Ross MD.Multipolar neurons in the spiral ganglion of rat[Ｊ].Acta otolaryngol,1973,76:381-394..
16. Ard,MD,Morest DK,Hauger SH.Trophic interactions between the cochleovestibular ganglion of the chick embryo and its synaptic targets in cultures[Ｊ].Neurosicence,1985,16:151-170..
17. Sobkowicz HM,Loftus JM,slapnick,et al.Tissue culture of the organ of corti[Ｊ].Acta Otolaryngol,1993,502:3-36..
18. 唐癑秌，Davis Robin.應用膜片鉗技術對螺旋神經元放電類型的初步研究[Ｊ]．聽力學及言語疾病雜志，2002，10（4）：250-252．.

West China Medical Journal

Primary Culture in Vitro and Identification of Murine Spiral Ganglion Neurons

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