M2-receptor subtype does not mediate muscarine-induced increases in [Ca2+]i in nociceptive neurons of rat dorsal root ganglia
M2-receptor subtype does not mediate muscarine-induced increases in [Ca2+]i in nociceptive neurons of rat dorsal root ganglia
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DOI：
10.1152/jn.2000.84.4.1934
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发表时间：
2000-10-01
影响因子：
2.5
通讯作者：
Kress, M
中科院分区：
文献类型：
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作者：
Haberberger, R;Scholz, A;Kress, M
多种M受体亚型存在于感觉神经元上，可能参与伤害性感觉的调节。在本研究中，我们从功能([Ca~(2+)](I)测定)、转录(RT-PCR)和翻译(免疫组织化学)水平研究了成年大鼠腰背根神经节(DRG)M受体亚型M2和M3(M2R，M3R)的存在。分离的中型神经元(直径20-35 mm)培养1d后，76%的神经元[Ca~(2+)](I)升高。[Ca~(2+)](I)在去除细胞外钙后没有升高，在重复应用激动剂后也不脱敏。[Ca~(2+)]~(2+)的这种升高可能是通过M3R的表达来解释的，M3R可以通过三磷酸肌醇诱导内储存的钙释放。事实上，M受体拮抗剂阿托品和M3R拮抗剂4-二苯基乙酰氧基-N-(2-氯乙基)-哌啶盐酸盐(4-DAMP)都能拮抗这种作用。总RNA的RT-PCR和单细胞RT-PCR证实了M3R的药理作用，表明M3R的mRNA在腰背根节和单个感觉神经元中存在。此外，RT-PCR还发现M2R的表达，这似乎不是钙变化的原因，因为M2受体拮抗剂加拉胺不能阻止M2R的表达。免疫组织化学显示M2R和M3R在中等大小的腰背根节神经元中存在，这些神经元也共同表达凝集素I-B4的结合部位，凝集素I-B4主要是皮肤伤害性感受器的标志。在推测为伤害性感受的I-B4阳性神经元中出现M受体，提示这些乙酰胆碱受体参与了伤害性刺激的处理过程的调制。
Multiple muscarinic receptor subtypes are present on sensory neurons that may be involved in the modulation of nociception. In this study we focused on the presence of the muscarinic receptor subtypes, M2 and M3 (M2R, M3R), in adult rat lumbar dorsal root ganglia (DRG) at the functional ([Ca2+](i) measurement), transcriptional (RT- PCR), and translational level (immunohistochemistry). After 1 day in culture exposure of dissociated medium-sized neurons (20-35 mm diam) to muscarine was followed by rises in [Ca2+](i) in 76% of the neurons. The [Ca2+](i) increase was absent after removal of extracellular calcium and did not desensitize after repetitive application of the agonist. This rise in [Ca2+](i) may be explained by the expression of M3R, which can induce release of calcium from internal stores via inositoltrisphospate. Indeed the effect was antagonized by the muscarinic receptor antagonist atropine as well as by the M3R antagonist, 4-diphenylacetoxy-N-(2 chloroethyl)-piperidine hydrochloride (4-DAMP). The pharmacological identification of M3R was corroborated by RT- PCR of total RNA and single-cell RT- PCR, which revealed the presence of mRNA for M3R in lumbar DRG and in single sensory neurons. In addition, RT- PCR also revealed the expression of M2R, which did not seem to contribute to the calcium changes since it was not prevented by the M2 receptor antagonist, gallamine. Immunohistochemistry demonstrated the presence of M2R and M3R in medium-sized lumbar DRG neurons that also coexpressed binding sites for the lectin I-B4, a marker for mainly cutaneous nociceptors. The occurrence of muscarinic receptors in putative nociceptive I-B4-positive neurons suggests the involvement of these acetylcholine receptors in the modulation of processing of nociceptive stimuli.