Hayashida K-I, Bynum T, Vincler M, Eisenach J C
Department of Anesthesiology and Center for the Pharmacologic Plasticity in the Presence of Pain, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157, USA.
Neuroscience. 2006 Jun 19;140(1):259-68. doi: 10.1016/j.neuroscience.2006.02.013. Epub 2006 Mar 31.
Acetylcholine reduces nociceptive input in part by activating inhibitory M2 muscarinic receptors on primary sensory neurons, and acetylcholinesterase inhibitors and muscarinic agonists produce analgesia in humans and animals. M2 muscarinic receptors are upregulated in animals with diabetic neuropathy, but their level of expression and function after peripheral nerve injury has not been previously examined. This study tested, using intracellular Ca(2+) response to membrane depolarization, the effect of the M2 muscarinic receptor agonist bethanechol on individual dorsal root ganglion cells from normal and L5-6 spinal nerve-ligated rats, followed by M2 muscarinic receptor immunostaining. We also examined functional transient receptor potential for vanilloids-1 activity by determining intracellular Ca(2+) response evoked by capsaicin in M2 muscarinic receptor immunoreactive cells. In normal dorsal root ganglion cells, bethanechol inhibited the Ca(2+) response in a concentration-related fashion, and this inhibition was blocked by the M2 muscarinic receptor antagonist gallamine. Cells expressing M2 muscarinic receptors by immunostaining were significantly inhibited by bethanechol, whereas those lacking positive staining were not. The proportion of studied dorsal root ganglion neurons with positive M2 muscarinic receptor staining increased significantly in the injured ipsilateral L5-6 and the uninjured ipsilateral L4 ganglia, but not in the contralateral dorsal root ganglion neurons compared with normals. In contrast, the proportion of neurons responding to capsaicin significantly decreased in the injured ipsilateral L5-6 dorsal root ganglion cells. These results suggest that inhibitory M2 muscarinic receptors are upregulated in small- and medium-sized axotomized dorsal root ganglion neurons and their uninjured neighbors following nerve injury, and may represent an appropriate target for analgesia in this setting.
乙酰胆碱部分通过激活初级感觉神经元上的抑制性M2毒蕈碱受体来减少伤害性传入，并且乙酰胆碱酯酶抑制剂和毒蕈碱激动剂在人和动物中可产生镇痛作用。在患有糖尿病性神经病变的动物中，M2毒蕈碱受体上调，但其在周围神经损伤后的表达水平和功能此前尚未得到研究。本研究使用对膜去极化的细胞内Ca(2+)反应，测试了M2毒蕈碱受体激动剂氨甲酰甲胆碱对正常大鼠和L5-6脊神经结扎大鼠单个背根神经节细胞的作用，随后进行M2毒蕈碱受体免疫染色。我们还通过测定辣椒素在M2毒蕈碱受体免疫反应性细胞中诱发的细胞内Ca(2+)反应，来检测功能性香草酸类受体-1的活性。在正常背根神经节细胞中，氨甲酰甲胆碱以浓度相关的方式抑制Ca(2+)反应，并且这种抑制被M2毒蕈碱受体拮抗剂加拉明阻断。通过免疫染色表达M2毒蕈碱受体的细胞被氨甲酰甲胆碱显著抑制，而那些缺乏阳性染色的细胞则未被抑制。与正常相比，在受伤的同侧L5-6和未受伤的同侧L4神经节中，M2毒蕈碱受体染色阳性的被研究背根神经节神经元的比例显著增加，但在对侧背根神经节神经元中未增加。相反，在受伤的同侧L5-6背根神经节细胞中，对辣椒素产生反应的神经元比例显著降低。这些结果表明，抑制性M2毒蕈碱受体在神经损伤后中小型轴突切断的背根神经节神经元及其未受伤的相邻神经元中上调，并且在这种情况下可能是镇痛的合适靶点。
