Choi J, Farley J M
Department of Pharmacology and Toxicology, University of Mississippi Medical Center, Jackson, USA.
J Pharmacol Exp Ther. 1998 May;285(2):588-94.
Cyclic GMP relaxes swine tracheal smooth muscle. Relaxation occurs because of decreases in intracellular calcium concentration ([Ca++]i) that are thought to occur through hyperpolarization which inhibits calcium influx. Activation of K+ channels has been suggested as the underlying mechanism for the hyperpolarization. In the present study, the effects of 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP, a membrane-permeable analog of cyclic GMP) on acetylcholine (ACh)-induced increases in [Ca++]i were examined by laser scanning confocal microscopy in fluo 3-loaded single cells. Membrane potential and currents were measured by the perforated-configuration of patch-clamp method, 8-Bromo-cGMP (1 microM-0.1 mM) inhibited 0.1 microM ACh-induced oscillations in [Ca++]i in a concentration-dependent manner. Spontaneous changes in membrane potential were observed by the patch-clamp method. Acetylcholine (0.03 microM) did not affect the time-averaged mean potential. The spontaneous changes in membrane potential were reduced and the cells were depolarized by 0.1 microM ACh and to a greater degree by 1 microM ACh. This result is consistent with previous observations of ACh-induced depolarization in intact tissue. The application of 0.1 mM 8-Br-cGMP had no significant effects on spontaneous changes in membrane potential and did not induce changes in membrane potential in cells treated with 0.1 microM ACh. In voltage-clamped cells, ACh (0.1 microM) induced oscillations in calcium-activated K+ currents. 8-Bromo-cGMP (0.1 mM) inhibited these ACh-induced oscillations in currents, but had no significant effects on spontaneous changes in membrane current in unstimulated cells. These data indicate that 8-Br-cGMP inhibits ACh-induced increases in [Ca++]i by mechanisms other than regulation of membrane potential.
环磷酸鸟苷可使猪气管平滑肌舒张。其舒张是由于细胞内钙浓度（[Ca++]i）降低所致，一般认为这是通过超极化抑制钙内流实现的。有研究表明钾离子通道的激活是超极化的潜在机制。在本研究中，运用激光扫描共聚焦显微镜，在负载了荧光素-3的单细胞中检测了8-溴鸟苷3',5'-环一磷酸（8-溴-cGMP，一种可透过细胞膜的环磷酸鸟苷类似物）对乙酰胆碱（ACh）诱导的[Ca++]i升高的影响。采用膜片钳技术的穿孔膜片配置来测量膜电位和电流。8-溴-cGMP（1微摩尔/升 - 0.1毫摩尔/升）以浓度依赖的方式抑制0.1微摩尔/升ACh诱导的[Ca++]i振荡。通过膜片钳技术观察到膜电位的自发变化。乙酰胆碱（0.03微摩尔/升）不影响时间平均的平均电位。膜电位的自发变化减少，细胞被0.1微摩尔/升ACh去极化，1微摩尔/升ACh使其去极化程度更大。这一结果与之前在完整组织中观察到的ACh诱导的去极化现象一致。施加0.1毫摩尔/升的8-溴-cGMP对膜电位的自发变化无显著影响，且未在经0.1微摩尔/升ACh处理的细胞中诱导膜电位变化。在电压钳制的细胞中，乙酰胆碱（0.1微摩尔/升）诱导钙激活钾电流振荡。8-溴-cGMP（0.1毫摩尔/升）抑制这些ACh诱导的电流振荡，但对未受刺激细胞的膜电流自发变化无显著影响。这些数据表明8-溴-cGMP通过调节膜电位以外的机制抑制ACh诱导的[Ca++]i升高。
