大鼠脑塑料切片乙酰胆碱酯酶组织化学研究的最佳参数。

L R Williams, M R Donald, K S Jodelis
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引用次数: 2

摘要

描述了一种改进的方法,以提高保存和光学分辨率的乙酰胆碱酯酶(AChE)在成年大鼠脑结构。根据Hedreen等人的研究,最佳组织制备方法为:在0.1M Sorensen磷酸缓冲液中,用4%的多聚甲醛、0.1%的glutaraldehyde和7%的蔗糖固定,pH为7.4,用50 mM的NH4Cl和2%的蔗糖在缓冲液中冲洗,丙酮脱水,用LKB组织树脂真空浸润,在4℃下聚合,在37℃下将10微米的片段在AChE组织化学反应混合物中过夜,并进行银强化。从大鼠基底前脑到同侧海马的胆碱能投射中乙酰胆碱酯酶的活性证明了该技术的有效性。该方法提供了ache阳性轴突过程的良好演示,并使胆碱能神经元的药物组织化学可视化成为可能。该程序为胆碱能神经元的分析提供了一种方便的方法,避免了其他乙酰胆碱酯酶组织化学程序中固有的潜在伪影。
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Optimal parameters for the histochemical demonstration of acetylcholinesterase in plastic sections of rat brain.

A modified method for improved preservation and optical resolution of acetylcholinesterase (AChE)-containing structures in adult rat brain is described. Optimal tissue preparation included fixation in paraformaldehyde 4%, glutaraldehyde 0.1%, and sucrose 7% in 0.1M Sorensen's phosphate buffer, pH 7.4, rinsing in buffer 50 mM with respect to NH4Cl and 2% with respect to sucrose, acetone dehydration, vacuum infiltration with LKB Historesin, and polymerization at 4 C, overnight incubation of 10 microns sections at 37 C in the AChE histochemical reaction mixture and silver intensification according to Hedreen et al. Demonstration of AChE enzyme activity in the cholinergic projection from the rat basal forebrain to the ipsilateral hippocampus exemplifies the usefulness of the technique. The method provides an excellent demonstration of AChE-positive axonal processes and enables the pharmacohistochemical visualization of cholinergic neurons. This procedure offers a convenient method for analysis of cholinergic neurons that avoids potential artifacts inherent in other AChE histochemical procedures.

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