Effects of stimulating the acetylcholine receptor on the current-voltage relationships of the smooth muscle membrane studied by voltage clamp of potential recorded by micro-electrode.

1. A double sucrose-gap voltage-clamp technique is described for use on smooth muscle strips longer than about 2 mm. It involves intracellular recording by microelectrode of the membrane potential of a narrow region of the strip ("node") sandwiched between two streams of deionized sucrose solution. Current was passed into the node across one or both sucrose streams. 2. Preliminary experiments in which potential was recorded intracellularly at two points during polarization of a "short cable" preparation, formed by folding over a strip of smooth muscle, suggested that a node width of less than 0-15 mm was needed to achieve uniform potential during inward current flow. However, when node width between sucrose-gaps was reduced to 0-5 mm, spontaneous electrical activity was lost, and below 0-5 mm spike threshold was raised and the regenerative spike became graded. The currents flowing during the application of rectangular voltage-clamp command potentials were described. 3. Using taenia smooth muscle it was shown by recording with a second, independent micro-electrode that potential was not uniform for up to 200 ms or more following a step change in potential under voltage-clamp in nodes 0-4-0-5 mm wide where current was passed across both sucrose gaps. However, reasonably uniform nodal potentials were obtained using ramps with relatively slow rates of rise (25 mV/s). 4. Using such slow ramp commands under voltage clamp, the effects of carbachol on the current-voltage relationship of longitudinal muscle of ileum and taenia were studied in hypertonic solution. 5. In the presence of carbachol (10(-6) to 10(-5) g/ml.) additional inward current flowed across the membrane (in some experiments an equilibrium potential was observed at which this current reversed direction). The magnitude of this additional current was linearly related to potential at potentials negative to the resting potential. At potentials positive to the resting membrane potential, this additional current increased with depolarization over the range -40 to -10 mV; in ileum the effect of this additional inward current on the current-voltage relationship was to produce a region of net inward current where before, in the absence of carbachol, a net outward current existed. In taenia the additional inward current flowing in the presence of carbachol was too small to produce a region of net inward current; thus carbachol produced regenerative slow oscillations of potential (slow waves) in ileum but not in taenia. 6. These results support a previous suggestion that activation of the acetylcholine receptor of ileal smooth muscle produces an additional inward current in the membrane which increases with depolarization and is responsible for the regenerative slow waves seen when muscarinic stimulants are applied. A similar effect apparently operates in taenia but the additional inward current is too small to produce regenerative slow waves.