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本文介绍了一种能在同一神经元内同时显示ＨＲＰ逆行标记以及Ｆｏｓ和某类神经递质或相关酶（例如ＴＨ）的三重标记技术。首先将ＨＲＰ注入脑内某一核团（如中央杏仁核），动物存活４８ｈ后，再向胃内导入１％福尔马林２ｍｌ，以造成对胃的伤害性刺激，２ｈ后将动物处死。脑的切片（如本文的延髓切片）先用四甲基联苯胺－钨酸钠（ＴＭＢ－ＳＴ）法进行ＨＲＰ呈色反应，后用二氨基联苯胺（ＤＡＢ）和氯化钴作加强；然后按ＡＢＣ法进行Ｆｏｓ免疫组织化学反应，继之按ＰＡＰ法进行ＴＨ免疫组织化学反应。光镜下在延髓切片观察到７种标记神经元，即ＴＨ－、ＨＲＰ－或Ｆｏｓ－单标记神经元，ＴＨ和ＨＲＰ－、ＴＨ和Ｆｏｓ－或ＨＲＰ和Ｆｏｓ的双标记神经元以及ＴＨ和ＨＲＰ和Ｆｏｓ的三标记神经元。该方法为研究同时显示中枢神经元的功能活动状态、传出投射及所含神经递质提供了一个强有力的工具。
This article presents a triple-tagging technique that displays both HRP retrograde labeling and Fos and certain neurotransmitters or related enzymes (eg, TH) within the same neuron. Firstly, HRP was injected into a certain nucleus in the brain (such as the central amygdala). After the animals survived for 48 hours, 1 ml of formalin 2 ml was infused into the stomach to cause noxious stimulation to the stomach, and the animals were sacrificed after 2 hours. Brain sections (eg, medulla oblongata) were first HRP-colored with tetramethylbenzidine-sodium tungstate (TMB-ST) followed by diaminobenzidine (DAB) and cobalt chloride According to ABC method, Fos immunohistochemical reaction was carried out, followed by TH immunohistochemical reaction by PAP method. Seven labeled neurons, TH-, HRP- or Fos-mono-labeled neurons, double labeled neurons of TH and HRP-, TH and Fos- or HRP and Fos, as well as TH and HRP And Fos’s triple-labeled neurons. This method provides a powerful tool for studying both the functional activity of central neurons and the transmission and projection of neurotransmitters.