可视脑片膜片钳技术前庭核团脑片的制备

The Preparation of Vestibular Nuclei Slices for Visual Patch Clamp Techniques

夏交;孔维佳;朱云;张宇;周彦;郭长凯;

1:华中科技大学同济医学院附属协和医院耳鼻咽喉科

摘要

目的改良脑片制备方法,探讨影响脑片活性的实验因素。方法在国内外报道常用方法的基础上进行改良,制备含有前庭内侧核的脑干脑片,从肉眼观、镜下观和放电特性三个方面判断脑片活性,观察鼠龄、生理溶液、温度等因素对脑片活性的影响。结果建立了稳定的脑片制备方法,通过红外微分干涉相差技术可直接观察脑片表层下50100μm神经元的立体影像,并可稳定地进行电生理记录;鼠龄越小神经元活性越易恢复,在成年鼠制备脑片时采取心脏灌注、使用高蔗糖溶液、较高温度预孵育等措施可提高脑片活性;孵育和记录温度直接影响神经元的兴奋性,一定范围内提高温度可提高神经元兴奋性,但会缩短脑片存活时间。结论改良的脑片制备方法简单可行,可为电生理研究提供大量具有良好活性的神经元,脑片制备方法的选择和实验环境的控制应根据所选用动物的年龄而调整。

关键词

脑片;红外微分干涉相差术;全细胞记录;前庭核

基金项目(Foundation):

国家杰出青年科学基金(39925035);; “十一五”国家科技支撑计划(2007BAI18B13);; 国家自然科学基金(30371525)资助项目

作者

夏交;孔维佳;朱云;张宇;周彦;郭长凯;

参考文献

1 Murchison D,Griffith WH.High-voltage-activated calciumcurrents in basal forebrain neurons during aging[J].J Neuro-physiol,1996,76:158.

2 Murchison D,Griffith WH.Low-voltage activated calciumcurrents increasein basal forebrain neurons fromaged rats[J].J Neurophysiol,1995,74:876.

3 Li CL,Mcil wain H.Maintenance of resting membrane poten-tials in slices of mammalian oerebral cortex and other tissuesinvitro[J].J Physiol,1957,139:178.

4 Dodt HU,Zieglgansberger W.Visualizing unstained neuronsin living brain slices by infrared dic-videomicroscopy[J].Brain research,1990,537:333.

5 Stuart GJ,Dodt HU,Sakmann B.Patch-clamp recordingsfromthe soma and dendrites of neuronsin brain slices usingin-frared video microscopy[J].Pflugers Arch,1993,423:511.

6刘振伟,李立君,刘传缋.海马脑片盲法膜片钳全细胞记录技术[J].生理学报,2001,53:405.

7涂娅莉,刘应兵,张莉,等.大鼠视皮层神经元电生理和形态学特性在发育中的变化[J].生理学报,2003,55:206.

8雷革胜,万业宏,王玉英,等.应用红外可视脑片膜片钳技术研究大鼠海马ca1神经元的电生理特性[J].第四军医大学学报,2004,25:1 153.

9 Moyer JRJ,Brown TH.Methods for whole-cell recordingfromvisually preselected neurons of perirhinal cortex in brainslices fromyoung and aging rats[J].Journal of NeuroscienceMethods,1998,86:35.

10 Paxinos G,Watson C.The rat brain in stereotaxic coordi-nates[M].5thed.San Diego:Elsevier Academic Press,2005.59~65.

11 Aitken PG,Breese GR,Dudek FF,et al.Preparative meth-ods for brain slices:Adiscussion[J].Journal of NeuroscienceMethods,1995,59:139.

12 Colbert CM.Preparation of cortical brain slices for electro-physiological recording[J].Methods in Molecular Biology,2006,337:117.

13 Sekirnjak C,du Lac S.Physiological and anatomical proper-ties of mouse medial vestibular nucleus neurosn projecting tothe oculomotor nucleus[J].Journal of Neurophysiology,2006,95:3 012.

14 Feig S,Lipton P.N-methyl-d-aspartate receptor activa-tion and Ca2+account for poor pyramidal cell structure inhippocampal slices[J].Journal of Neurochemistry,1990,55:473.

15 Choi DW.Excitotoxic cell death[J].Journal of Neurobiolo-gy,1992,23:1 261.

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脑片红外微分干涉相差术全细胞记录前庭核

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夏交孔维佳朱云张宇周彦郭长凯