CLARITY with neuronal tracing and immunofluorescence to study the somatosensory system in rats,Journal of Neuroscience Methods

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CLARITY with neuronal tracing and immunofluorescence to study the somatosensory system in rats
Journal of Neuroscience Methods ( IF 2.7 ) Pub Date : 2020-12-24 , DOI: 10.1016/j.jneumeth.2020.109048
Guadalupe Martínez-Lorenzana ₁ , Mohammed Gamal-Eltrabily ₁ , Irma Alejandra Tello-García ₁ , Ataulfo Martínez-Torres ₁ , Marymar Becerra-González ₁ , Abimael González-Hernández ₁ , Miguel Condés-Lara ₁

Affiliation

Background

The CLARITY technique enables researchers to visualize different neuronal connections along the nervous system including the somatosensory system.

New method

The present work describes the antero-lateral and dorsal column pathways until the thalamic and cortical stations, as well as descending oxytocinergic and vasopressinergic innervations by means of combined CLARITY, neuronal tracing, and immunofluorescence techniques. We used male Sprague-Dawley rats of 13, 30, and 60 days.

Results

The main results are as follows: A) CLARITY is a reliable technique that can be combined with fluorescent neuronal tracers and immunofluorescence techniques without major procedure modifications; B) at spinal level, some primary afferent fibers were labeled by CGRP, as well as the presence of neuronal populations that simultaneously project to the gracile and ventral posterolateral thalamic nuclei; C) corticothalamic connections were visible when retrograde tracers were injected at thalamic level; D) oxytocin receptors were expressed in the spinal dorsal horn by GABAergic-positive neurons, reinforcing previous outcomes about the possible mechanism for oxytocin blocking the primary afferent sensory input.

Comparison with existing methods and Conclusions

The CLARITY technique lets us observe in a transparent way the entire processed tissue compared with classical histological methods. CLARITY is a potentially useful tool to describe neuroanatomical structures and their neurochemical stratus.

中文翻译：

清晰的神经元示踪和免疫荧光研究大鼠的体感系统

背景

CLARITY技术使研究人员可以观察到沿神经系统（包括体感系统）的不同神经元连接。

新方法

目前的工作描述了直到丘脑和皮层站的前外侧和背柱通路，以及通过结合的CLARITY，神经元示踪和免疫荧光技术使催产素能和血管加压素的神经支配下降。我们使用了13、30和60天的雄性Sprague-Dawley大鼠。

结果

主要结果如下：A）清晰度是一种可靠的技术，可以与荧光神经元示踪剂和免疫荧光技术相结合，而无需对程序进行重大修改；B）在脊柱水平上，一些初级传入纤维被CGRP标记，同时存在神经元群体，这些神经元群体同时投射到脆弱和腹侧后外侧丘脑核；C）在丘脑水平注射逆行示踪剂时可见皮质丘脑连接；D）催产素受体由GABA能性阳性神经元在脊髓背角中表达，从而加强了先前有关催产素阻断主要传入感觉输入的可能机制的结果。