GTP-gamma-S increases the duration of backward swimming behavior and the calcium action potential in marine Paramecium

Behavioral and electrophysiological experiments were made to examine the hypothesis that G-proteins modulate the voltage-dependent calcium channel in the marine ciliate Paramecium calkinsi. It was found that guanosine-5′-O-(3-thiotriphosphate) (GTP-gamma-S), an analogue of GTP that binds to and activates G-proteins, increased the duration of backward swimming behavior in reversibly permeabilized Paramecium in an irreversible and concentration-dependent manner. At 1 mumol l-1 GTP-gamma-S, the duration of backward swimming behavior was increased fivefold. Other nucleotides and related compounds did not have a significant effect on the backward swimming behavior. To evaluate whether the behavioral effects were due to ion channel modulation, the calcium action potential in intact Paramecium was monitored before and after guanine nucleotide injection. Within 5 min after the injection of GTP-gamma-S or GTP into the cell, the duration of the calcium action potential was prolonged at least threefold. Like the behavioral response, the GTP-gamma-S effect on the calcium action potential duration was irreversible, whereas the effect of GTP began to decay after 6 min. GDP-beta-S, which binds to and inactivates G-proteins, markedly reduced the calcium action potential within 5 min after injection. These results support the hypothesis that the voltage-dependent calcium channels present in Paramecium are modulated by GTP-binding proteins.

A TTX-resistant propagating calcium action potential

The cross-commissural (CC) cell in the supraesophageal ganglion of the giant barnacle, Balanus nubilus, was stimulated intrasomatically and antidromically in normal saline and 3 X 10(-7) M tetrodotoxin (TTX) saline. The action potential in normal saline contained both sodium and calcium components, each independently capable of propagation. Evidence that the action potential in TTX saline was calcium dependent included: the amplitude of the spike in TTX saline increased monotonically with increasing calcium; it was blocked by the calcium channel blockers La, Ni, Cd and Co; and equimolar substitution of Ba or Sr for Ca in TTX saline supported regenerative activity. Evidence that the calcium component could propagate alone included: the distance over which the calcium action potential traveled exceeded the space constant of the axon; the biphasic nature of the extracellularly recorded action potential, which propagated to the axon in the nerve root, indicated inward regenerative current was occurring on the axon; electrotonically spread potentials were clearly distinguishable from active regenerative potentials. In addition, optical experiments using the calcium indicator dye arsenazo III (28) showed that the relative magnitude of the calcium signal was not diminished along the axon in TTX saline compared with normal saline. The critical external calcium concentration necessary to support the propagating action potential in TTX saline was estimated to be between 1.25 and 5 mM. To our knowledge, this is the first direct observation of a neuron with sodium and calcium channels of apparently normal kinetics where the calcium component alone can propagate in the absence of an outward current blocker. Our results suggest that there is a greater density of calcium channels on the CC axon than on the axons of other neurons.

Sodium-Calcium Action Potential Associated with Contraction in the Heliozoan Actinocoryne Contractilis

The electrophysiology of the contractile protozoan Actinocoryne contractilis was studied with conventional intracellular recording techniques. Resting membrane potential (−78 mV, s.d. = 8, N = 18) was dependent upon external K+. Rapid action potentials (overshoot up to 50 mV) were evoked either by mechanical stimulation or by current injection. Graded membrane depolarizations induced by graded mechanical stimuli correspond to receptor potentials. The receptor potential was mainly Na+-dependent; the action potential was also mainly Na+-dependent, but involved a minor Ca2+-dependence. The two components of the action potential could be separated in Ca2+-free solution containing EGTA (1 mmol l−1), in low-Na+ solutions or by the addition of Co2+. The repolarizing phase of the action potential was sensitive to TEA ions and to 4-aminopyridine (4-AP). Action potentials were followed in 10–20 ms by a rapid all-or-none contraction of the axopods and stalk. Contraction was blocked in Ca2+-free solution containing EGTA and by Co2+, which suggests a requirement of external Ca2+ for this event. Contraction was also abolished by 4-AP.