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Effect of membrane potential on acetylcholine-induced inward current in guinea-pig ileum
1Department of Physiology, University of Cologne, FRG.
Insights
Acetylcholine (ACh) triggers inward cation current (Ins, ACh) in guinea-pig ileum smooth muscle. This current
Area of Science:
- Electrophysiology
- Pharmacology
- Smooth Muscle Physiology
Background:
- Acetylcholine (ACh) is a key neurotransmitter in the gastrointestinal tract.
- Understanding ion channel function is crucial for smooth muscle regulation.
- The precise gating mechanisms of ACh-induced currents remain incompletely understood.
Purpose of the Study:
- To investigate the voltage-dependent gating of acetylcholine (ACh)-induced inward current (Ins, ACh).
- To characterize the biophysical properties of Ins, ACh in guinea-pig ileum longitudinal muscle cells.
Main Methods:
- Whole-cell patch clamp technique on isolated guinea-pig ileum longitudinal muscle cells.
- Voltage clamp and current clamp experiments.
- Ionic substitution experiments to determine channel selectivity.
Main Results:
- ACh (300 microM) induced a slow, sustained inward current (Ins, ACh) through non-selective cation channels.
- Ins, ACh amplitude increased instantaneously upon hyperpolarization and then relaxed, with relaxation time constants dependent on voltage.
- The steady-state activation of Ins, ACh followed Boltzmann kinetics, with half-maximal activation at -50 mV, indicating voltage-dependent gating.
Conclusions:
- ACh-induced depolarization in guinea-pig ileum smooth muscle is controlled by membrane potential.
- The voltage-dependent gating of Ins, ACh channels explains the observed membrane potential control.
- These findings elucidate a key mechanism in cholinergic smooth muscle regulation.
Abstract:
1. The whole-cell patch clamp technique with caesium aspartate internal solution was used with single isolated cells from the longitudinal muscle layer of guinea-pig ileum, to investigate the voltage-dependent gating of ACh-induced inward current. 2. In voltage clamp experiments, at holding potentials ranging from -80 to -30 mV, ACh (300 microM) produced a slow sustained inward current in physiological salt bath solution (PSS). The measurements of the reversal potentials on substituting Na+ by other monovalent and divalent cations showed that this current is through non-selective cation channels (Ins, ACh). 3. During hyperpolarizations, Ins, ACh instantaneously increased in amplitude and then relaxed to a new steady-state level. The I-V relationship of the instantaneous peak was linear with a reversal potential of 0 mV, while that of the steady state was bell-shaped. The time course of relaxation appeared to be monoexponential and its time constants were reduced by stronger hyperpolarizations. 4. These results were not affected by the organic Ca2+ antagonists D600 or nitrendipine (10 microM). Under this condition, maximal chord conductance of Ins, Ach which was observed at 0 mV was about 1.5 nS. The steady-state activation relationship was well fitted by Boltzmann's equation with a half-maximal activation (Vh) of -50 mV and a slope factor (k) of -15 mV at membrane potentials negative to 0 mV, but over 0 mV the degree of activation was again decreased. The time constants for relaxation also appeared to follow a sigmoid curve. 5. In current clamp experiments, superfusion of ACh (300 microM) depolarized the membrane up to -10 to 0 mV. Inward current injection resulting in the moderate hyperpolarization of the membrane (-70 to -80 mV) attenuated ACh-induced depolarization and stronger hyperpolarization (less than -80 mV) abolished it. 6. These results show that ACh-induced depolarization is controlled by the membrane potential, which is explained by the voltage-dependent gating of Ins, Ach.