EFFECTS OF 8-BROMO-CYCLIC GMP ON MEMBRANE-POTENTIAL OF SINGLE SWINE TRACHEAL SMOOTH-MUSCLE CELLS

Cyclic GMP relaxes swine tracheal smooth muscle. Relaxation occurs bec ause of decreases in intracellular calcium concentration ([Ca++](i)) t hat are thought to occur through hyperpolarization which inhibits calc ium influx. Activation of K+ channels has been suggested as the underl ying mechanism for the hyperpolarization. In the present study, the ef fects of 8-bromo-guanosine 3',5'-cyclic monophosphate (8-Br-cGMP, a me mbrane-permeable analog of cyclic GMP) on acetylcholine (ACh)-induced increases in [Ca++](i) were examined by laser scanning confocal micros copy in flue 3-loaded single cells. Membrane potential and currents we re measured by the perforated-configuration of patch-clamp method. 8-B romo-cGMP (1 mu M-0.1 mM) inhibited 0.1 mu M ACh-induced oscillations in [Ca++](i) in a concentration-dependent manner. Spontaneous changes in membrane potential were observed by the patch-clamp method. Acetylc holine (0.03 mu M) did not affect the time-averaged mean potential. Th e spontaneous changes in membrane potential were reduced and the cells were depolarized by 0.1 mu M ACh and to a greater degree by 1 mu M AC h. 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 potentia l and did not induce changes in membrane potential in cells treated wi th 0.1 mu M ACh. In voltage-clamped cells, ACh (0.1 mu M) induced osci llations in calcium-activated K+ currents. 8-Bromo-cGMP (0.1 mM) inhib ited these ACh-induced oscillations in currents, but had no significan t effects on spontaneous changes in membrane current in unstimulated c ells. These data indicate that 8-Br-cGMP inhibits ACh-induced increase s in [Ca++](i) by mechanisms other than regulation of membrane potenti al.