Department of Anatomy, Histology and Embryology, K K Leung Brain Research Centre, The Fourth Military Medical University, Xi'an, PR China.
Neurochem Int. 2011 Jan;58(1):44-51. doi: 10.1016/j.neuint.2010.10.008. Epub 2010 Oct 23.
Enkephalinergic (ENKergic) neurons have been proposed to play crucial roles in pain modulation in the trigeminal subnucleus caudalis (Vc). To assist an advance in the research of ENKergic neurons, here we used preproenkephalin-green fluorescent protein (PPE-GFP) transgenic mice, in which all ENKergic neurons were fluorescent. We first performed fluorescent in situ hybridization combined with immunofluorescent histochemistry to confirm the specificity of this transgenic mouse and its advantages in showing ENKergic neurons in the Vc. Then based on this useful transgenic mouse, we examined the phenotypic diversity of PPE-GFP neurons by immunostaining for several markers that characterize ENKergic neuron subtypes. About 25.9±1.9% of GFP-positive neurons were regarded as immunoreactive for glutamic acid decarboxylase (GAD)(67) mRNA and 14.7±1.4% of GFP-positive neurons were positive for γ-aminobutyric acid. The proportions of calbindin-, calretinin-positive cells among the ENKergic neurons were 8.4±1.2% and 7.3±1.7%, respectively. Only 1.1±0.1% of GFP-positive neurons colocalized with parvalbumin and no GFP-positive neurons were found to co-express neuronal nitric oxide synthase. We then injected retrograde tracer into the thalamic regions and observed that a small number of ENKergic neurons in the Vc were retrogradely labeled with the tracer. The present results provide a detailed morphological evidence of the neurochemical features of ENKergic neurons. These results have broad implications for understanding the functional roles of ENKergic neurotransmission in the Vc.
啡肽能（ENKergic）神经元被认为在三叉神经尾核（Vc）的疼痛调制中发挥关键作用。为了协助 ENKergic 神经元研究的进展，我们使用了前啡肽原-绿色荧光蛋白（PPE-GFP）转基因小鼠，其中所有的 ENKergic 神经元都是荧光的。我们首先进行了荧光原位杂交结合免疫荧光组织化学，以确认这种转基因小鼠的特异性及其在显示 Vc 中的 ENKergic 神经元方面的优势。然后，基于这种有用的转基因小鼠，我们通过免疫染色几种标记物来检查 PPE-GFP 神经元的表型多样性，这些标记物可以表征 ENKergic 神经元亚型。约 25.9±1.9%的 GFP 阳性神经元被认为对谷氨酸脱羧酶（GAD）（67）mRNA 具有免疫反应性，而 14.7±1.4%的 GFP 阳性神经元对γ-氨基丁酸呈阳性。钙结合蛋白、钙调蛋白阳性细胞在 ENKergic 神经元中的比例分别为 8.4±1.2%和 7.3±1.7%。只有 1.1±0.1%的 GFP 阳性神经元与副甲状腺蛋白共表达，没有 GFP 阳性神经元被发现共表达神经元型一氧化氮合酶。然后，我们将逆行示踪剂注入丘脑区域，并观察到 Vc 中的少量 ENKergic 神经元被示踪剂逆行标记。这些结果提供了 ENKergic 神经元神经化学特征的详细形态学证据。这些结果对于理解 Vc 中 ENKergic 神经传递的功能作用具有广泛的意义。
