WGA-Alexa transsynaptic labeling in the phrenic motor system of adult rats: Intrapleural injection versus intradiaphragmatic injection
WGA-Alexa transsynaptic labeling in the phrenic motor system of adult rats: Intrapleural injection versus intradiaphragmatic injection
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DOI：
10.1016/j.jneumeth.2014.12.013
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发表时间：
2015-02-15
影响因子：
3
通讯作者：
Goshgarian, Harry G.
中科院分区：
文献类型：
--
作者：
Buttry, Janelle L.;Goshgarian, Harry G.
背景：胸腔内注射CTB-Alexa 488，一种逆行示踪剂，提供了一种替代手术侵入性剖腹手术所需的标记技术。然而，CTB-Alexa 488不能穿过突触，将示踪剂限制在脊髓的膈核和肋间运动核。新方法：胸膜内注射经突触示踪剂WGA-Alexa 488，提供了一种标记脊髓和髓质的呼吸运动通路的方法。采用阴道内注射WGA-Alexa 594和迷走神经注射True blue。确认膈核和区分髓质的rVRG和NA。结果：胸膜内注射后，WGA-Alexa 488逆行转运至膈核和肋间运动核。随后，WGA-Alexa 488经突触从膈运动神经元传递到rVRG的前运动神经元，后者在激励期间向膈神经元提供下行驱动。此外，通过WGA-Alexa 488和True blue的双重标记，在NA的选定细胞中鉴定出WGA-Alexa 488。与现有方法比较：WGA-Alexa 488胸膜内注射后显示逆行跨突触标记，而之前注射cbb - alexa 488的方法仅显示逆行标记。结论：胸膜内注射WGA-Alexa氟偶联物是一种有效的经突触标记膈运动系统的方法，为侵入性剖腹手术提供了另一种选择。此外，该研究提供了rVRG和NA细胞之间直接突触关系的第一个解剖学证据。(C) 2014 Elsevier B.V.版权所有
Background: Intrapleural injection of CTB-Alexa 488, a retrograde tracer, provides an alternative labeling technique to the surgically invasive laparotomy required for intradiaphragmatic injection. However, CTB-Alexa 488 is incapable of crossing synapses restricting the tracer to the phrenic nuclei and the intercostal motor nuclei in the spinal cord.New method: Intrapleural injection of WGA-Alexa 488, a transsynaptic tracer, provides a method to label the respiratory motor pathway in both the spinal cord and medulla. Intradiaphragmatic injection of WGA-Alexa 594 and vagal nerve injections of True blue were used to. confirm the phrenic nuclei and to differentiate between the rVRG and the NA in the medulla.Results: Following intrapleural injection, WGA-Alexa 488 was retrogradely transported to the phrenic nuclei and to the intercostal motor nuclei. Subsequently WGA-Alexa 488 was transsynaptically transported from the phrenic motoneurons to the pre-motor neurons in the rVRG that provide the descending drive to the phrenic neurons during inspiration. In addition WGA-Alexa 488 was identified in select cells of the NA confirmed by a dual label of both WGA-Alexa 488 and True blue.Comparison with existing method: WGA-Alexa 488 demonstrates retrograde transsynaptic labeling following intrapleural injection whereas the previous method of injecting CTB-Alexa 488 only demonstrates retrograde labeling.Conclusions: Intrapleural injection of WGA-Alexa fluor conjugates is an effective method to transsynaptically label the phrenic motor system providing an alternative for the invasive laparotomy required for intradiaphragmatic injections. Furthermore, the study provides the first anatomical evidence of a direct synaptic relationship between rVRG and select NA cells. (C) 2014 Elsevier B.V. All rights reserved.