大鼠腳橋被蓋核至A7核區正腎上腺素神經元 之膽鹼性及麩胺酸性投射機制與特徵

Abstract

腳橋被蓋核區(pedunculopontine tegmental nucleus, PPTg)是一包含膽鹼性(cholinergic)、麩胺酸性(glutamatergic)等其他種類之神經元投射至許多區域的核區。在過去的動物行為實驗研究指出，在腳橋被蓋核給予電刺激或化學性刺激之後可以引發出鎮痛反應。而在前人研究結果也指出A7正腎上腺素神經元(NAergic A7 neurons)上擁有膽鹼性受器，A7正腎上腺素神經元已有文獻證實會投射軸突至脊髓背角(dorsal horn)，分泌正腎上腺素來調控痛覺傳遞。根據以上結果，我們假設A7正腎上腺素神經元接受來自腳橋被蓋核區之膽鹼性神經投射來調控下游之痛覺傳遞，並由型態方面及電生理方面進行實驗來探討兩者之間連結是否存在及其特性。經由腦定位儀手術將BDA正向神經追蹤劑注射至腳橋被蓋核區，經由免疫染色標定後觀察其投射之神經纖維和A7細胞群。在型態方面的實驗結果可以看到來自腳橋被蓋核被BDA標定之軸突纖維往A7細胞群延伸並形成似突觸之結構。而電生理實驗方面，在腳橋被蓋核予以單一電刺激可在A7細胞群引發一內向電流，同時施給木防已苦毒素(picrotoxin)和士的寧(strychnine)可以部份抑制此內向電流，顯示在此神經投射當中具有抑制性(inhibitory)神經纖維。高頻率電刺激則可在A7細胞群記錄到一可被蕈毒膽鹼性受器(mAChRs)拮抗劑-阿托品(atropine)部分抑制之突觸後興奮性電流，顯示在此神經投射中也具有膽鹼性的神經纖維。在此高頻刺激之後，也在A7細胞群此處記錄到麩胺酸性的非同步釋放(asynchronous release)的現象。在此神經投射也發現了自主回饋抑制(auto-inhibition)的調控現象。這些實驗結果證明從腳橋被蓋核區往A7正腎上腺素神經元的投射的確存在，也顯示出在此神經投射當中包含有抑制性，膽鹼性，以及麩胺酸性的神經纖維。此實驗結果也對於脊髓以上蕈毒膽鹼性系統與正腎上腺素之下行性痛覺調控路徑的交互作用提供了有利之證據。

Pedunculopontine tegmental nucleus (PPTg) is a nucleus containing cholinergic neurons, glutamatergic neurons, and other kinds of neurons projecting toward many regions. Many behavioral studies have shown that electrical or chemical stimulation at PPTg can induce antinociception response. Also previous studies have shown that there are cholinergic receptors on NAergic A7 cell group, which projects NAergic fibers to dorsal horn to modulate nociceptive signaling. From the above results, we hypothesized that NAergic A7 neurons receive cholinergic projection from PPTg to modulate descending nociceptive signaling. The projection from PPTg to NAergic A7 neurons was investigated by morphological and electrophysiological methods. BDA anterograde tracer was injected in PPTg by stereotaxic surgery, and the A7 neurons and the projection fibers from PPTg were labeled by immunohistochemistry. In the morphological results, labeled axon fibers spread toward to NAergic A7 neurons forming synaptic terminal contact-like structures. In electrophysiological experiments, single pulse at PPTg was used to evoke an inward current which could be partial blocked by co-application of picrotoxin (antagonist of GABAA receptors) and strychnine (antagonist of glycine receptors), suggesting there are inhibitory components in this projection. High frequency stimulation at PPTg evoked an inward current partially blocked by muscarinic antagonist, atropine, suggesting that cholinergic fibers are also involved in this projection. Moreover, calcium-dependent asynchronous glutamate release events were also found on A7 neurons in this experiment. There was an auto-inhibition in the cholinergic system between A7 and PPTg transmission. These results provide an evidence for the existence of the projection from PPTg toward NAergic A7 neurons, and there are least inhibitory, glutamatergic, and cholinergic fibers involved in this projection. The results in this study also provide an evidence for supraspinal interaction between muscarinic cholinergic system and NAergic descending pain pathway.