Cohen R I, Molina-Holgado E, Almazan G
Department of Pharmacology and Therapeutics, McGill University, Montreal, Que., Canada.
Brain Res Mol Brain Res. 1996 Dec 31;43(1-2):193-201. doi: 10.1016/s0169-328x(96)00176-3.
To determine if muscarinic receptor-activation plays a role in oligodendrocyte development, the effect of carbachol a stable acetylcholine analog, on gene expression and proliferation was investigated. Using Northern blot analysis we showed that carbachol caused a time and concentration-dependent increase in c-fos mRNA. This effect was blocked by atropine, a non-selective muscarinic antagonist. In addition, the muscarinic-stimulated c-fos increase was inhibited by 1-(5-isoquinoline-sulfonyl)-2-methylpiperazine (H-7), a potent inhibitor of protein kinase C (PKC), but not by N-2-(p-bromocinnamylamino)-ethyl-5-isoquinoline-sulfonamide (H-89), a potent inhibitor of protein kinase A, suggesting the involvement of PKC in mediating the response. Down-regulation of PKC by overnight pre-treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) blocked only the phorbol ester-stimulated c-fos accumulation while no effect was observed in the carbachol-induced response. These results suggested that carbachol stimulated an H-7 sensitive PKC pathway which may be different than that activated by TPA. Further evidence for two separate mechanisms of proto-oncogene induction was provided by the additive effect of carbachol and TPA. Induction of c-fos mRNA by carbachol was dependent on both influx of extracellular Ca2+ and release from intracellular stores, as both EDTA and BAPTA blocked the response. Since activation of muscarinic receptors can affect cell division in other cellular systems, the effect of carbachol on [3H]thymidine and bromodeoxyuridine incorporation into oligodendrocyte DNA was measured. Carbachol stimulated DNA synthesis in oligodendrocyte progenitors. This effect was mediated by muscarinic receptors as [3H]thymidine incorporation was prevented or significantly reduced by the addition of atropine. In conclusion, the present findings suggest that, the neurotransmitter, acetylcholine may act as a trophic factor in developing oligodendrocytes, regulating their growth and development in the central nervous system.
为了确定毒蕈碱受体激活在少突胶质细胞发育中是否起作用，研究了稳定的乙酰胆碱类似物卡巴胆碱对基因表达和增殖的影响。通过Northern印迹分析，我们发现卡巴胆碱导致c-fos mRNA呈时间和浓度依赖性增加。这种效应被非选择性毒蕈碱拮抗剂阿托品阻断。此外，毒蕈碱刺激的c-fos增加被蛋白激酶C（PKC）的强效抑制剂1-(5-异喹啉磺酰基)-2-甲基哌嗪（H-7）抑制，但不被蛋白激酶A的强效抑制剂N-2-(对溴肉桂酰胺基) -乙基-5-异喹啉磺酰胺（H-89）抑制，这表明PKC参与介导该反应。用12-O-十四酰佛波醇13-乙酸酯（TPA）过夜预处理下调PKC，仅阻断佛波酯刺激的c-fos积累，而在卡巴胆碱诱导的反应中未观察到影响。这些结果表明，卡巴胆碱刺激了一条对H-7敏感的PKC途径，该途径可能与TPA激活的途径不同.卡巴胆碱和TPA的相加效应为原癌基因诱导的两种独立机制提供了进一步的证据。卡巴胆碱诱导c-fos mRNA依赖于细胞外Ca2+内流和细胞内储存释放，因为EDTA和BAPTA均阻断了该反应.由于毒蕈碱受体的激活可以影响其他细胞系统中的细胞分裂，因此测量了卡巴胆碱对少突胶质细胞DNA中[3H]胸苷和溴脱氧尿苷掺入的影响。卡巴胆碱刺激少突胶质细胞祖细胞中的DNA合成。这种效应由毒蕈碱受体介导，因为加入阿托品可阻止或显著减少[3H]胸苷掺入。总之，目前的研究结果表明，神经递质乙酰胆碱可能作为发育中的少突胶质细胞中的一种营养因子,调节它们在中枢神经系统中的生长和发育。
