Synaptic structures of the nociceptive primary afferent central terminals in the substantia gelatinosa of spinal trigeminal nucleus candalis

本实验旨在研究小鼠脊髓背角内CGRP、SP免疫反应阳性（CGRP、SP-IR）初级传入终末与GABA免疫反应阳性（GABA-IR）中间神经元的关系。采用ABC法对游离切片进行包埋前GABA免疫染色。超薄切片收集在Formvar涂层的镍网格上。免疫胶体金法检测CGRP和SP。GABA阳性细胞体分布于背角浅层。CGRP或SP阳性终末与胶状质中间神经元的CABA阳性树突形成突触前连接。这些结果表明，初级传入纤维传递的信息可以直接传递给GABA能神经元，伤害性信息激活抑制回路，为了直接显示CI终末与GABA和Met-脑啡肽能中间神经元之间的突触，采用辣椒素处理和免疫标记方法。用免疫金标记法对小鼠腰髓后角和三叉神经脊束核浅层进行电镜观察。GABA（15 nm）和Met-Enk（5 nm）用不同大小的IgG金进行双标记。肾小球CI-终端突触前接触周围的树突表现出GABA和Met-Enk免疫反应（IR）。非肾小球CI-终末也与GABA-和Met-Enk-IR索马体形成突触前接触。在单个树突和胞体中，GABA和Met-Enk可能同时存在于某些中间神经元中。本研究结果表明，伤害性初级传入神经元通过抑制性GABA能和Met-脑啡肽能中间神经元调节背角浅层和三叉神经脊束核尾侧的疼痛传递。

The present study was designed to elucidate the relationship of CGRP-, SP-immunoreactive (CGRP-, SP-IR) primary afferent central terminals with GABA-immunoreactive (GABA-IR) interneurons in the mouse dorsal horn. Pre-embedding immunostaining for GABA was done on free-floating sections with ABC method. Ultrathin sections were collected on Formvar-coated nickel grids. The grids were immunostained for CGRP or SP with postembedding immunogold technique. The GABA-IR cell bodies were distributed throughout the superficial dorsal horn. The CGRP- or SP-IR terminals were presynaptic to the CABA-IR dendrite of interneurons in the substantia gelatinosa. These findings suggest that the information transmitted by primary afferent fibers can be relayd directly to GABAergic neurons, and inhibitory circuits are activated by nociceptive information.In order to directly demonstrate the synapses between CI-terminals and GABA- and Met-Enkephalinergic interneurons, capsaicin treatment and immunolabeling methods were utilized. Superficial layr of the lumbar spinal cord dorsal horn and spinal trigeminal nucleus candalis of the mouse were examined by electron microscopy after post-embedding immunogold labeling procedures. Double-labeling with different size of IgG gold was used for GABA (15nm) and Met-Enk (5nm). Glomerular CI-terminals made presynaptic contacts with surrounding dendrites exhibiting GABA and Met-Enk immunoreactivities (IR). Nonglomerular CI-terminals also made presynaptic contacts with GABA- and Met-Enk-IR soma. Frequently a single dendrite and somata showed double labeling, i.e.GABA and Met-Enk might coexist in some of the interneurons. The present findings strongly suggest that nociceptive primary afferent neurons modulate the pain transmission by themselves via inhibitory GABAergic and Met-Enkephalinergic interneurons in the superficial layr of dorsal horn and spinal trigeminal nucleus caudalis.