Calcium currents in GH3 cultured pituitary cells under whole-cell voltage-clamp: inhibition by voltage-dependent potassium currents.

To isolate inward Ca2+ currents in GH3 rat pituitary cells, an inward Na+ current as well as two outward K+ currents, a transient voltage-dependent current (IKV) and a slowly rising Ca2+-activated current (IKCa), must be suppressed. Blockage of these outward currents, usually achieved by replacement of intracellular K+ with Cs+, reveals sustained inward currents. Selective blockage of either K+ current can be accomplished in the presence of intracellular K+ by use of quaternary ammonium ions. When IKCa and Na+ currents are blocked, the net current elicited by stepping the membrane potential (Vm) from -60 to 0 mV is inward first, becomes outward and peaks in 10-30 msec, and finally becomes inward again. Under this condition, in which both IKV and Ca2+ currents should be present throughout the duration of the voltage step, the Ca2+ current was not detected at the time of peak outward current. That is, plots of peak outward current vs. Vm are monotonic and are not modified by nisoldipine or low external Ca2+ as would be expected if Ca2+ currents were present. However, similar plots at times other than at peak current are not monotonic and are altered by nisoldipine or low Ca2+ (i.e., inward currents decrease and plots become monotonic). When K+ channels are first inactivated by holding Vm at -30 mV, a sustained Ca2+ current is always observed upon stepping Vm to 0 mV. Furthermore, substitution of Ba2+ for Ca2+ causes blockage of IKV and inhibition of this current results in inward Ba2+ currents with square wave kinetics. These data indicate that the Ca2+ current is completely inhibited at peak outward IKV and that Ca2+ conductance is progressively disinhibited as the transient K+ current declines due to channel inactivation. This suggests that in GH3 cells Ca2+ channels are regulated by IKV.     

Effects of quinidine on plateau currents of guinea-pig ventricular myocytes

Effects of quinidine on membrane currents forming the plateau of action potentials were studied using an isolated single ventricular cell from guinea-pig hearts. Quinidine (5 mg/l) produced a fall and shortening of the early part of the plateau, and delayed its later part and final repolarization, without changes in resting membrane potential. Application of quinidine caused a reversible depression of the peak Ca2+ current by about 30% of the control. Delayed outward K+ current, iK, also decreased to less than 20% of the control. Thus, an outward tail current upon repolarization to -40 mV from depolarizing voltage steps of the plateau ranges became inward. Current values at the end of 200 ms pulses in response to voltage steps to -60-0 mV were always positive and were not changed by the drug. The inward current elicited at potentials negative to resting potential level, also, decreased by 13% to 23% of the control in the presence of the drug, but the effect was not reversible upon wash-out of the drug. These results suggest that quinidine causes a non-specific depression of inward rectifier K+ current, iK1, with minor degree but has little effect on the window sodium current. Therefore, changes in the action potential repolarization produced by quinidine can be explained by its effects on both calcium current and delayed outward K+ current.     

Voltage-dependent calcium channel in the squid axon.

A voltage-dependent inward calcium current insensitive to tetrodotoxin has been measured in internally perfused or dialyzed squid giant axons. Sodium conductance was blocked by tetrodotoxin, and potassium conductance was irreversibly destroyed by the lack of potassium in the external and internal medium. Chloride conductance was eliminated by replacement of chloride with methylsulfonate. The calcium current was activated at about -40 mV and it peaked at about 0 mV. The peak inward current was 3 microA/cm2 when external Ca was 80 mM and 0.9 microA/cm2 in 8 mM Ca. In 80 mM Ca, the calcium current is turned on in less than 10 msec, and it does not decay appreciably for pulses up to 70 msec in duration. Barium can replace calcium, and cadmium blocks the calcium current.     

Dual modulation of K channels by thyrotropin-releasing hormone in clonal pituitary cells.

Transmembrane electrical activity in pituitary tumor cells can be altered by substances that either stimulate or inhibit their secretory activity. Using patch recording techniques, we have measured the resting membrane potentials, action potentials, transmembrane macroscopic ionic currents, and single Ca2+-activated K channel currents of GH3 and GH4/C1 rat pituitary tumor cells in response to thyrotropin-releasing hormone (TRH). TRH, which stimulates prolactin secretion, causes a transient hyperpolarization of the membrane potential followed by a period of elevated action potential frequency. In single cells voltage clamped and internally dialyzed with solutions containing K+, TRH application results in a transient increase in Ca2+-activated K currents and a more protracted decrease in voltage-dependent K currents. However, in cells internally dialyzed with K+-free solutions, TRH produces no changes in inward Ca2+ or Ba2+ currents through voltage-dependent Ca channels. The time courses of the effects on Ca2+-activated and voltage-dependent K currents correlate with the phases of hyperpolarization and hyperexcitability, respectively. During application of TRH to whole cells, single Ca2+-activated K channel activity increases in cell-attached patches not directly exposed to TRH. In contrast, TRH applied directly to excised membrane patches produces no change in single Ca2+-activated K channel behavior. We conclude that TRH (i) triggers intracellular Ca2+ release, which opens Ca2+-activated K channels, (ii) depresses voltage-dependent K channels during the hyperexcitable phase, which further elevated intracellular Ca2+, and (iii) does not directly modulate Ca channel activity.     

Effects of quinidine on action potentials and ionic currents in isolated canine ventricular myocytes

We examined the effects of quinidine (5-20 microM) on transmembrane action potentials and ionic currents of isolated canine ventricular myocytes. Collagenase treatment of canine ventricular tissue produced a yield of 40-60% healthy cells. Myocytes had normal resting and action potentials as measured using conventional microelectrodes. Quinidine decreased Vmax, amplitude, overshoot, and the duration of action potentials stimulated by passage of brief current pulses through the recording pipette. Recovery was complete after washout except that action potential duration was prolonged compared with control. A discontinuous single microelectrode voltage ("switch") clamp was used to measure ionic currents. Quinidine irreversibly reduced steady-state outward current as measured with three different voltage clamp protocols. Quinidine reversibly decreased peak calcium current as well as the slowly inactivating and/or steady-state inward currents in the plateau voltage range, presumably both "late" sodium (tetrodotoxin-sensitive) and calcium (tetrodotoxin-insensitive) currents. The effect on calcium current showed both tonic and use-dependent block. Thus, quinidine has a multitude of actions on both inward and outward currents, which combine to produce the net effect of quinidine on action potential configuration.     

Blocking and isolation of a calcium channel from neurons in mammals and cephalopods utilizing a toxin fraction (FTX) from funnel-web spider poison

A Ca2+-channel blocker derived from funnel-web spider toxin (FTX) has made it possible to define and study the ionic channels responsible for the Ca2+ conductance in mammalian Purkinje cell neurons and the preterminal in squid giant synapse. In cerebellar slices, FTX blocked Ca2+-dependent spikes in Purkinje cells, reduced the spike afterpotential hyperpolarization, and increased the Na+-dependent plateau potential. In the squid giant synapse, FTX blocked synaptic transmission without affecting the presynaptic action potential. Presynaptic voltage-clamp results show blockage of the inward Ca2+ current and of transmitter release. FTX was used to isolate channels from cerebellum and squid optic lobe. The isolated product was incorporated into black lipid membranes and was analyzed by using patch-clamp techniques. The channel from cerebellum exhibited a 10- to 12-pS conductance in 80 mM Ba2+ and 5-8 pS in 100 mM Ca2+ with voltage-dependent open probabilities and kinetics. High Ba2+ concentrations at the cytoplasmic side of the channel increased the average open time from 1 to 3 msec to more than 1 sec. A similar channel was also isolated from squid optic lobe. However, its conductance was higher in Ba2+, and the maximum opening probability was about half of that derived from cerebellar tissue and also was sensitive to high cytoplasmic Ba2+. Both channels were blocked by FTX, Cd2+, and Co2+ but were not blocked by omega-conotoxin or dihydropyridines. These results suggest that one of the main Ca2+ conductances in mammalian neurons and in the squid preterminal represents the activation of a previously undefined class of Ca2+ channel. We propose that it be termed the "P" channel, as it was first described in Purkinje cells.     

Presynaptic inhibition in Aplysia involves a decrease in the Ca2+ current of the presynaptic neuron.

By voltage clamping presynaptic cell L10 and using pharmacologic separation techniques, we have analyzed the specific ionic currents in the presynaptic cell that correlate with presynaptic inhibition while assaying transmitter release with intracellular recordings from postsynaptic cells. We have found that presynaptic inhibition can be elicited in conditions in which the Na+ and the various K+ channels are pharmacologically blocked and depolarizing current pulses produce only an inward Ca2+ current. Both inward currents and tail currents at and above the K+ reversal potential were always less inward during presynaptic inhibition. The changes in conductance associated with presynaptic inhibition were voltage sensitive and paralleled the voltage sensitivity of the Ca2+ channel. We therefore conclude that presynaptic inhibition is caused by a direct transmitter-mediated decreased of presynaptic Ca2+-channel conductance.     

Early afterdepolarizations: mechanism of induction and block. A role for L-type Ca2+ current

Early afterdepolarizations (EADs) are a type of triggered activity found in heart muscle. We used voltage-clamped sheep cardiac Purkinje fibers to examine the mechanism underlying EADs induced near action potential plateau voltages with the Ca2+ current agonist Bay K 8644 and the effect of several interventions known to suppress or enhance these EADs. Bay K 8644 produced an inward shift of the steady-state current-voltage relation near plateau voltages. Tetrodotoxin, lidocaine, verapamil, nitrendipine, and raising [K]o abolish EADs and shift the steady-state current-voltage relations outwardly. Using a two-pulse voltage-clamp protocol, an inward current transient was present at voltages where EADs were induced. The voltage-dependence of availability of the inward current transient and of EAD induction were similar. The time-dependence of recovery from inactivation of the inward current transient and of EAD amplitude were nearly identical. Without recovery of the inward current transient, EADs could not be elicited. The inward current transient was enhanced with Bay K 8644 and blocked by nitrendipine, but was not abolished by tetrodotoxin or replacement of [Na]o with an impermeant cation. These results support a hypothesis that the induction of EADs near action potential plateau voltages requires 1) a conditioning phase controlled by the sum of membrane currents present near the action potential plateau and characterized by lengthening and flattening of the plateau within a voltage range where, 2) recovery from inactivation and reactivation of L-type Ca2+ channels to carry the depolarizing charge can occur. Our results suggest an essential role for the L-type Ca2+ "window" current and provide a framework for understanding the role of several membrane currents in the induction and block of EADs.     

Trimebutine maleate has inhibitory effects on the voltage-dependent Ca2+ inward current and other membrane currents in intestinal smooth muscle cells

We examined effects of trimebutine maleate on the membrane currents of the intestinal smooth muscle cells by using the tight-seal whole cell clamp technique. Trimebutine suppressed the Ba2+ inward current through voltage-dependent Ca2+ channels in a dose-dependent manner. The inhibitory effect of trimebutine on the Ba2+ inward current was not use-dependent. It shifted the steady-state inactivation curve to the left along the voltage axis. Trimebutine also had inhibitory effects on the other membrane currents of the cells, such as the voltage-dependent K+ current, the Ca2(+)-activated oscillating K+ current and the acetylcholine-induced inward current. These relatively non-specific inhibitory effects of trimebutine on the membrane currents may explain, at least in part, the dual actions of the drug on the intestinal smooth muscle contractility, i.e. inhibitory as well as excitatory.     

Noninactivating, tetrodotoxin-sensitive Na+ conductance in rat optic nerve axons.

The ionic current underlying the upstroke of axonal action potentials is carried by rapidly activating, voltage-dependent Na+ channels. Termination of the action potential is mediated in part by the rapid inactivation of these Na+ channels. We previously demonstrated that an influx of Na+ plays a critical role in the cascade leading to irreversible anoxic injury in central nervous system white matter. We speculated that a noninactivating Na+ conductance mediates this pathological Na+ influx and persists at depolarized membrane potentials as seen in anoxic axons. In the present study we measured the resting compound membrane potential of rat optic nerves using a modified "grease-gap" technique. Application of tetrodotoxin (2 microM) to resting nerves ([K+]o = 3 mM) or to nerves depolarized by 15 or 40 mM K+ resulted in hyperpolarizing shifts of membrane potential. We interpret these shifts as evidence for a persistent, noninactivating Na+ conductance. This conductance is present at rest and persists in nerves depolarized sufficiently to abolish classical transient Na+ currents. PK/PNa ratios were estimated at 35.5, 23.2, and 88 in 3 mM, 15 mM, and 40 mM K+, respectively. We suggest that this noninactivating Na+ conductance may provide an inward pathway for Na+ ions, necessary for the operation of Na+, K(+)-ATPase. Under pathological conditions, such as anoxia, this conductance is the likely route of Na+ influx, which causes damaging Ca2+ entry through reverse operation of the Na(+)-Ca2+ exchanger. The presence of this conductance in white matter axons may provide a therapeutic opportunity for diseases such as stroke and spinal cord injury.     

Facilitation of Ca2+-channel currents in bovine adrenal chromaffin cells.

Ca2+-channel currents in primary cultures of bovine adrenal chromaffin cells were studied using the whole-cell patch-clamp method. Parameters of a double-pulse protocol were systematically varied to characterize facilitation by a prepulse (P1) of Ca2+-channel current during a test pulse (P2). The pulses were usually separated by 30 msec, an interval sufficient for decay of any measurable P1 tail currents. The Ca2+-channel current amplitude during P2 increased when P1 voltage was more positive than 0 mV. The effect became progressively greater with more positive P1 voltage. With a 60-msec P1 to +80 mV, the current amplitude typically increased by 25%-35% during a 60-msec P2. Comparison of facilitated and control inward Ca2+-channel current I(V) curves showed that facilitation was also strongly dependent on P2 test voltage. Facilitation of Ca2+-channel currents is a voltage-dependent phenomenon and is not dependent on Ca2+ entry. When short repetitive voltage-clamp pulses were applied, the Ca2+-channel current amplitude increased with each pulse. This suggests that Ca2+-channel facilitation could enhance release of catecholamines from chromaffin cells during a train of action potentials.     

Receptors of the serotonin 1C subtype expressed from cloned DNA mediate the closing of K+ membrane channels encoded by brain mRNA.

The modulation of K+ channels by serotonin (5-HT) receptors was studied by coinjecting Xenopus oocytes with mRNA transcribed in vitro from a cloned 5-HT 1C subtype (5-HT1C) receptor gene, together with size-fractionated mRNA isolated from rat cerebral cortex that expresses K+ channels. After intracellular loading with EGTA to block Ca2(+)-dependent chloride currents, these oocytes responded to 5-HT with an inward current associated with a decrease in membrane conductance. Membrane current responses were small or absent in oocytes injected with either mRNA alone. We conclude that 5-HT1C receptors are able to cause the closing of a class of K+ channels expressed by cortex mRNA in a Ca2(+)-independent manner. The coupling between the receptors and channels appears to be mediated by the inositol phospholipid second messenger pathway, since activation of this pathway by application of serum evoked a similar closing current.     

Gonadotropin-releasing hormone induces oscillatory membrane currents in rat gonadotropes

Electrophysiological studies were performed to characterize membrane currents of rat gonadotropes under basal conditions and after exposure to secretagogues. Gonadotropes were identified in primary cultures of rat anterior pituitaries by a reverse hemolytic plaque assay. Giga-seal patch clamp recording with the cell-attached configuration was used to monitor membrane currents in these cells. Spontaneous spikes in basal current were seen. These were blocked by methoxyverapamil and probably reflect Ca2+-dependent action potentials. Brief GnRH stimulation induced slow oscillatory changes in membrane current that evolved into a series of large amplitude inward pulses after about 8 min. Treatment with TRH had no effect, and depolarization with K+ led to delayed inward currents without any oscillatory behavior. Under conditions of Ca2+ channel blockade, GnRH stimulation did not induce pulses of inward current, but did lead to oscillatory activation of a small conductance ion channel apparently selective for K+. Taken together these results suggest that GnRH induces oscillations in intracellular Ca2+ and that these oscillations are controlled by biochemical processes.     

Changes in left ventricular repolarization and ion channel currents following a transient rate increase superimposed on bradycardia in anesthetized dogs

INTRODUCTION: We previously demonstrated in dogs that a transient rate increase superimposed on bradycardia causes prolongation of ventricular refractoriness that persists for hours after resumption of bradycardia. In this study, we examined changes in membrane currents that are associated with this phenomenon. METHODS AND RESULTS: The whole cell, patch clamp technique was used to record transmembrane voltages and currents, respectively, in single mid-myocardial left ventricular myocytes from dogs with 1 week of complete AV block; dogs either underwent 1 hour of left ventricular pacing at 120 beats/min or did not undergo pacing. Pacing significantly heightened mean phase 1 and peak plateau amplitudes by approximately 6 and approximately 3 mV, respectively (P < 0.02), and prolonged action potential duration at 90% repolarization from 235+/-8 msec to 278+/-8 msec (1 Hz; P = 0.02). Rapid pacing-induced changes in transmembrane ionic currents included (1) a more pronounced cumulative inactivation of the 4-aminopyridine-sensitive transient outward K+ current, Ito, over the range of physiologic frequencies, resulting from a approximately 30% decrease in the population of quickly reactivating channels; (2) increases in peak density of L-type Ca2+ currents, I(Ca.L), by 15% to 35 % between +10 and +60 mV; and (3) increases in peak density of the Ca2+-activated chloride current, I(Cl.Ca), by 30% to 120% between +30 and +50 mV. CONCLUSION: Frequency-dependent reduction in Ito combined with enhanced I(Ca.L) causes an increase in net inward current that may be responsible for the observed changes in ventricular repolarization. This augmentation of net cation influx is partially antagonized by an increase in outward I(Ca.Cl).     

Activation thresholds of K(V), BK andCl(Ca) channels in smooth muscle cells in pial precapillary arterioles

We have previously shown expression of voltage-gated K+ channels (K(V)) in smooth muscle of cerebral arterioles and suggested the channels function to oppose voltage-dependent Ca2+ entry. However, other studies indicate that large conductance Ca2+-activated K+ (BK) channels serve this function and chloride (Cl-) channels may have the opposite effect. In this study we compared the activation thresholds and absolute current amplitudes for K(V) channels, BK channels and Cl- channels at physiological membrane potentials in intact precapillary arterioles from the rabbit cerebral circulation. Patch-clamp recordings were made to measure current and membrane potential, and a video scan line was used to detect external diameter. Two strategies to determine the basal current-voltage relationship of BK channels showed the channels contributed current only at voltages positive of -35 mV, even though voltage-dependent Ca2+-entry occurred. Ca2+-activated and niflumic acid-sensitive Cl- current was detected but, between -50 and -10 mV, both BK and Cl- channel currents were much smaller and contributed less to the membrane potential compared with K(V) channel current. Furthermore, in the absence of an exogenous vasoconstrictor agent, block of K(V) channels but not BK or Cl- channels caused constriction, although in the presence of endothelin-1 block of BK or K(V) channels caused constriction. The data indicate K(V) channels are the first inhibitory mechanism to activate when there is depolarisation in precapillary arteriolar smooth muscle cells of the cerebral circulation.     

Patch-clamp study of the ionic currents underlying action potentials in cultured frog pituitary melanotrophs

The ionic conductance mechanisms underlying the action potential behaviour of frog melanotrophs in primary culture were studied by using the patch-clamp technique in whole-cell configuration. The action potentials spontaneously generated by these cells were predominantly sodium spikes with a calcium component. Voltage-dependent sodium, calcium, potassium and calcium-activated potassium currents were identified and analysed separately. The voltage-dependent sodium current was characterized by its fast kinetic, its low-threshold activation, its voltage-dependent inactivation and a tetrodotoxin sensitivity. Calcium currents were identified on the basis of their ionic selectivity to divalent cations (Ba2+, Ca2+, Co2+) and their time course. Only two of the three well-documented calcium currents could be detected in frog melanotrophs. A sustained calcium current (ICaS) and an inactivating calcium current (ICaN) were elicited by step depolarizations up to -20 mV. ICaN inactivated for membrane potentials more positive than -50 mV; its inactivation appeared to be both voltage- and calcium-dependent. Transient calcium current (ICaT) has never been observed. Two types of potassium currents were identified: voltage-dependent potassium (IKV) and calcium-activated potassium currents, (IK[Ca]). They were both suppressed by tetraethylammonium chloride, whereas only IK(Ca) was blocked by cobalt. These major ionic currents underlying spontaneous electrical activity are assumed to be involved in the process of alpha-melanocyte-stimulating hormone release. The present study provides the ground for future investigations regarding the relationships between the electrical and secretory activities in amphibian pars intermedia cells.     

Sustained inward current during pacemaker depolarization in mammalian sinoatrial node cells

Several time- and voltage-dependent ionic currents have been identified in cardiac pacemaker cells, including Na(+) current, L- and T-type Ca(2+) currents, hyperpolarization-activated cation current, and various types of delayed rectifier K(+) currents. Mathematical models have demonstrated that spontaneous action potentials can be reconstructed by incorporating these currents, but relative contributions of individual currents vary widely between different models. In 1995, the presence of a novel inward current that was activated by depolarization to the potential range of the slow diastolic depolarization in rabbit sinoatrial (SA) node cells was reported. Because the current showed little inactivation during depolarizing pulses, it was called the sustained inward current (I(st)). A similar current is also found in SA node cells of the guinea pig and rat and in subsidiary pacemaker atrioventricular node cells. Recently, single-channel analysis has revealed a nicardipine-sensitive, 13-pS Na(+) current, which is activated by depolarization to the diastolic potential range in guinea pig SA node cells. This channel differs from rapid voltage-gated Na(+) or L-type Ca(2+) channels both in unitary conductance and gating kinetics. Because I(st) was observed only in spontaneously beating SA node cells, ie, it was absent in quiescent cells dissociated from the same SA or atrioventricular node, an important role of I(st) for generation of intrinsic cardiac automaticity was suggested.     

Early Afterdepolarization Formation in Cardiac Myocyte:

Early Afterdepolarization Formation, introduction: Early afterdepolarizations (EADs) are among the mechanisms proposed to underlie ventricular arrhythmias. Sea anemone toxin, ATXII, known to delay Na inactivation and to induce plateau level voltage oscillations, was used to study the formation of EADs. Methods and Results: Action potential and membrane currents were studied in rat ventricular myocytes using whole cell current and voltage clamp techniques. Phase plane trajectories were generated by plotting membrane potential (V) versus the first time derivative of membrane potential (dV/dt). Under current clamp conditions, ATXII (40 nM) consistently prolonged the action potential and induced EADs. The EADs developed at a plateau voltage between -10 and -40 mV. Calcium channel blockers, verapamil 10 μM and cobalt 4 μM, and the sarcoplasmic reticulum modulator, ryanodine (1 μM) did not antagonize ATXII effects on the action potential and EADs. However, Na channel blockers, tetrodotoxin 0.3μM and lidocaine 40μM. and rapid stimulation consistently shortened the prolonged action potential and suppressed EADs. Under voltage clamp conditions in the presence of ATXII, a slowly decaying inward current followed the fast inward current during depolarizing pulses. Membrane currents flowing at or later than 100 msec after the test pulse were analyzed. The control isochronal current-voltage (I-V) curves showed no late inward currents. In the presence of ATXII, all the isochronal I-V curves showed an inward current that was more prominent between -40 and 0 mV. The ATXII-induced current at the 100-msec isochronc activated at a potential of approximately -60 mV, peaked at about -20 mV, and reversed at +40 mV consistent with the Na current I-V curve. The isochronal I-V curves obtained after lidocaine superfusion resembled those of the control. The phase plane trajectory of the action potential obtained with ATXII showed an oscillatory behavior corresponding to the t AD range of potential; within this voltage range, the isochronal I-V curves were shown to cross the abscissa three times instead of once. Conclusion: These results suggest that, in this experimental model, neither sarcolemmal L-type Ca current nor sarcoplasmic reticulum Ca release plays a significant role in the genesis of ATXII-induced EADs. EADs are generated by a voltage-dependent balance between a markedly prolonged Na inward current and K outward currents within the voltage plateau range of the action potential hut not by Ca current reactivation and inactivation.     

Mechanism of calcium current modulation underlying presynaptic facilitation and behavioral sensitization in Aplysia.

Behavioral sensitization of the gill-withdrawal reflex of Aplysia is caused by presynaptic facilitation at the synapses of the mechanoreceptor sensory neurons of the reflex onto the motor neurons and interneurons. The presynaptic facilitation has been shown to be simulated by serotonin (the putative presynaptic facilitatory transmitter) and by cyclic AMP and to be accompanied by an increase in the Ca2+ current of sensory neuron cell bodies exposed to tetraethylammonium. This increase in the Ca2+ current could result from either a direct action on the Ca2+ channel or an action on an opposing K+ current. Here we report voltage clamp experiments which indicate that the increase in Ca2+ current associated with presynaptic facilitation results from a decrease in a K+ current. Stimulation of the connective (the pathway that mediates sensitization) or application of serotonin causes a decrease in a voltage-sensitive, steady-state outward current measured under voltage clamp as well as an increase in the transient net inward and a decrease in the transient outward currents elicited by brief depolarizing command steps. The reversal potential of the steady-state synaptic current is sensitive to extracellular K+ concentration, and both the steady-state synaptic current and the changes in the transient currents are blocked by K+ current blocking agents and by washout of K+. These results suggest that serotonin and the natural transmitter released by connective stimulation act to decrease a voltage-sensitive K+ current. The decrease in K+ current prolongs the action potential, and this in turn increases the duration of the inward Ca2+ current and thereby enhances transmitter release.     

Egg-laying hormone of Aplysia induces a voltage-dependent slow inward current carried by Na+ in an identified motoneuron.

This report presents studies on ionic currents in Aplysia motoneuron B16 that are modulated by the neuropeptide egg-laying hormone (ELH) of Aplysia. ELH induces an inward current that persists in the presence of the peptide and that decays slowly after ELH is removed from the bath. The effect is not due to a decrease in the delayed potassium current, the calcium-activated potassium current, or the transient potassium current. Current-voltage measurements indicate that ELH produces increased inward currents from -80 mV to approximately equal to 0 mV. The effect is particularly enhanced in the region from -40 mV to -25 mV where a negative slope conductance due to voltage-dependent slow inward current is observed. The slow inward current and the response to ELH persist in saline solutions in which Ca2+ is replaced with Co2+ but are eliminated when Na+ is replaced with equimolar concentrations of either Tris or N-methyl-D-glucamine. The response to ELH is unaffected by replacing chloride with equimolar acetate; by increasing the potassium concentration; or by adding tetraethylammonium chloride, CsCl, 4-amino-pyridine, or tetrodotoxin to the saline bath. In addition, the reversal potentials for the ELH response (range, -28 to +46 mV), obtained from difference current-voltage relationships, are consistent with an increase in the Na+-dependent slow inward current. We conclude that at least one of the effects of ELH on B16 is to increase a slow inward current carried by Na+.