Reeve H L, Vaughan P F, Peers C
Institute for Cardiovascular Studies, Leeds University, U.K.
Neuropharmacology. 1995 Mar;34(3):319-26. doi: 10.1016/0028-3908(94)00161-k.
The effects of muscarine on whole-cell Ca2+ channel currents in SH-SY5Y cells were studied using conventional and perforated-patch-clamp techniques, with 10 mM Ba2+ as charge carrier. Muscarine (10-300 microM) caused concentration-dependent inhibitions of Ca2+ channel currents which were only reversible when perforated-patch recordings were used. Inhibition of currents was associated with slowing of activation kinetics in approximately 50% of cells. In the presence of 5 microM nifedipine, muscarine was still able to inhibit currents, but after pre-exposure of cells to 1 microM omega-conotoxin GVIA the inhibitory effects of muscarine were almost completely lost. In the presence of 100 microM muscarine, Bay K 8644 (5 microM) was still able to enhance current amplitudes. Pre-treatment of cells with pertussis toxin (250 ng/ml for 16-24 hr) or inclusion of 1 mM GDP-beta-S in the patch-pipette prevented the inhibitory actions of muscarine. Hexahydrosiladifenidol (0.1-1 microM) antagonized the actions of muscarine (calculated pA2 7.1) but the presence of 10 microM pirenzipine or 0.1 microM methoctramine in the bath solution did not alter the degree of current inhibition caused by 100 microM muscarine. In summary, these results indicate that muscarine in SH-SY5Y cells causes inhibition of N-type Ca2+ channels via a M3 receptor coupled to a pertussis toxin-sensitive G-protein.
采用常规膜片钳技术和穿孔膜片钳技术，以10 mM Ba2+作为载流子，研究了毒蕈碱对SH-SY5Y细胞全细胞膜Ca2+通道电流的影响。毒蕈碱（10 - 300 μM）引起Ca2+通道电流的浓度依赖性抑制，只有在使用穿孔膜片记录时才是可逆的。电流抑制与约50%的细胞中激活动力学减慢有关。在存在5 μM硝苯地平的情况下，毒蕈碱仍能抑制电流，但在细胞预先暴露于1 μM ω-芋螺毒素GVIA后，毒蕈碱的抑制作用几乎完全丧失。在存在100 μM毒蕈碱的情况下，Bay K 8644（5 μM）仍能增强电流幅度。用百日咳毒素（250 ng/ml，作用16 - 24小时）预处理细胞或在膜片吸管中加入1 mM GDP-β-S可阻止毒蕈碱的抑制作用。六氢硅二苯基哌啶醇（0.1 - 1 μM）拮抗毒蕈碱的作用（计算得出的pA2为7.1），但在浴液中存在10 μM哌仑西平或0.1 μM美索曲明并不会改变100 μM毒蕈碱引起的电流抑制程度。总之，这些结果表明，SH-SY5Y细胞中的毒蕈碱通过与百日咳毒素敏感的G蛋白偶联的M3受体导致N型Ca2+通道的抑制。
