Osborne N N, FitzGibbon F, Schwartz G
Nuffield Laboratory of Ophthalmology, University of Oxford, U.K.
Vision Res. 1991;31(7-8):1119-27. doi: 10.1016/0042-6989(91)90038-7.
Cultured retinal pigment epithelium cells prepared from post-mortem adult human eyes are shown to contain muscarinic receptors associated with phosphoinositide turnover. Carbachol at a concentration of 100 microM induced a four-fold increase in 3H-inositol phosphates (more than 76% is in the form of 3H-inositol-1-phosphate) accumulation within 45 min in cells prelabelled with 3H-myoinositol and exposed to 5 mM LiCl. The EC50 of carbachol was approx. 70 microM and the saturation concentration was about 1 mM. The carbachol-induced response was blocked by both atropine and pirenzepine, the former being most effective. Pre-exposure of cells to carbachol resulted in desensitization and a drastic reduction in the subsequent carbachol-induced stimulation of 3H-inositol phosphates. The carbachol response could be attenuated by the biologically active phorbol ester, 4 beta-phorbol 12-myristate 13-acetate, and this was nullified by the protein kinase C inhibitor, staurosporine. The biologically inactive phorbol ester, 4 alpha-phorbol 12,13 dideconoate, did not attenuate the carbachol-induced stimulation of 3H-inositol phosphates. Pertussis toxin failed to influence the carbachol receptor-mediated phosphoinositide turnover. These studies provide clear evidence for the occurrence of muscarinic receptors coupled to phosphoinositide hydrolysis on human retinal pigment epithelium cells.
从成年人类死后眼睛制备的培养视网膜色素上皮细胞显示含有与磷酸肌醇代谢相关的毒蕈碱受体。在预先用³H-肌醇标记并暴露于5 mM氯化锂的细胞中，100 μM浓度的卡巴胆碱在45分钟内使³H-肌醇磷酸酯（超过76%为³H-肌醇-1-磷酸形式）积累增加了四倍。卡巴胆碱的EC50约为70 μM，饱和浓度约为1 mM。卡巴胆碱诱导的反应被阿托品和哌仑西平阻断，前者最为有效。细胞预先暴露于卡巴胆碱会导致脱敏，并使随后卡巴胆碱诱导的³H-肌醇磷酸酯刺激大幅降低。卡巴胆碱反应可被生物活性佛波酯4β-佛波醇12-肉豆蔻酸酯13-乙酸酯减弱，而这被蛋白激酶C抑制剂星形孢菌素消除。生物无活性的佛波酯4α-佛波醇12,13-二癸酸酯不会减弱卡巴胆碱诱导的³H-肌醇磷酸酯刺激。百日咳毒素未能影响卡巴胆碱受体介导的磷酸肌醇代谢。这些研究为人类视网膜色素上皮细胞上存在与磷酸肌醇水解偶联的毒蕈碱受体提供了明确证据。
