Electrical properties and membrane currents in the ciliateDidinium

Abstract

1. The electrical properties of the ciliateDidinium nasutum, aParamecium predator, have been investigated using techniques of constant current injection, voltage-clamp, exchange of solutions, and non-lethal deciliation. The electrical observations have been related to structural characteristics (Fig. 1). 2. The resting potential was primarily K-dependent. K concentrations beyond 1 mmol/1 depolarized the cell and reduced the input resistance. The resting conductance for Ca was low. Partial substitution of Ca by Mg changed the membrane potential very little and had no effect on the input resistance. Chloride did not influence the membrane properties. 3. The late V/I-relationship was characterized by pronounced outward and minimal inward rectification. It was very little modified by Ca concentrations ranging between 0.125 mmol/1 and 8 mmol/1. Raising the K concentration between 0.063 and 16 mmol/1 did not affect membrane rectification. 4. Membrane depolarization triggered a stimulus-graded action potential, which was Ca-dependent. The action potential rose with rates of up to 6 V/s; its overall duration was 100 to 500 ms. Repolarization occurred along a fast and a slow (shoulder) component and was followed by pronounced after-hyperpolarization. 5. Complete deciliation ofDidinium eliminated the regenerative depolarization of the membrane. The resting potential, input resistance and late current/voltage relationship were not modified by the removal of ciliary membranes. Under voltage-clamp deciliation removed the early inward current and the hump component of the late outward current. 6. Depolarizing voltage-clamp pulses of up to + 20 mV revealed a residual inward current following the early transient. The transient included two decay time constants (3.3 ms; 31.8 ms). An 0.8 nA persistent inward current was isolated using the difference current of ciliated and deciliated cells. A late outward current rose with depolarizations beyond 20 mV including an upward inflection (hump) about 50 ms after step onset. 7. A decreasing Ca/Ba mole-fraction increased the frequency and duration of spontaneous action potentials, and depressed the amplitudes of both, early inward and late outward current under voltage-clamp. In Ca-free Ba solution the unclamped membrane fired repetitively; the depolarizations included shoulders of 10 s or more. Inactivation of inward Ba currents was slow under depolarizing voltage-clamp; the late outward current was strongly depressed. In deciliated cells an early inward current was missing in Ca-free Ba solution, and the late I/V relation resembled that of deciliated cells in Ba-free Ca solution. 8. Tail currents following the late outward current of ciliated cells in Ca-solution and of deciliated cells in Ca- or Ba-solutions decayed with similar time constants. 9. In conclusion, the voltage-activated Ca conductance of the ciliary membrane ofDidinium compares well with data from ciliates so far studied. Unique is (i) a persistent inward current isolated using minor step depolarization or the difference current of ciliated and deciliated cells, (ii) a slow activating voltage-dependent K conductance, and (iii) a late, potentially Ca-dependent, outward current, which may be related to spatial separation of the cilia from the majority of the soma membrane. Ba interferes with the Ca channel which favours a previous two-binding site model. Ba can inhibit the K conductance from inside the cell after passing the ciliary Ca channel.