Substance P depolarizes nerve terminals in an autonomic ganglion

The effect of substance P on presynaptic nerve terminals was examined by intracellular impalements of calyciform terminals within the chick ciliary ganglion. Substance P produced a slow depolarization of the nerve terminals which was associated with an increase in input resistance. The postsynaptic ciliary neurons were unaffected by exposure to substance P, indicating that at this synapse the physiological effects of substance P may be largely presynaptic in nature.     

Calcium currents in a vertebrate presynaptic nerve terminal: the chick ciliary ganglion calyx

Ca currents (ICa) were recorded from presynaptic nerve terminals in the chick ciliary ganglion. Ciliary neurons are innervated by a single nerve terminal that extends over a wide area of the neuron surface to form a 'calyx'. The neurons were dissociated enzymatically with the calyx intact and the patch clamp technique was used in the whole cell mode to record ion currents. A small inward ICa (peak current 20-80 pA) was recorded that was blocked by external Cd. Only one component of ICa was detected. This was recruited at positive membrane potentials, exhibited no evidence of inactivation during a 25-ms depolarizing pulse, and deactivated rapidly. Thus, the ICa recorded in this vertebrate presynaptic nerve terminal was similar to the high-voltage activated, fast deactivating, current reported in other neurons.     

Inward rectifier produced by Xenopus oocytes injected with mRNA extracted from carp olfactory epithelium

Ionic channels encoded by mRNA extracted from carp olfactory epithelium were investigated by injection into Xenopus laevis oocytes. The oocytes expressed an inward rectifier K+ -channel, as detected under two-electrode voltage clamp conditions. The results were as follows. An inactivating inward current appeared on hyperpolarization and increased with increasing extracellular K+ concentrations. The 0 current potentials plotted as a function of log [K+]0 in the range between 2 to 20 mMK+ fell on a straight line, with a slope of 58 mV per tenfold change in K+ concentration, indicating that the current carrier is K+. Chord conductances reached saturation levels on extreme hyperpolarization. The chord conductances at the saturation levels were 35.7, 22.5, and 13.4 muSec in 20, 10, and 5 mM extracellular K+, respectively. Extracellular application of 0.1 mM Cs+ or 0.1 mM Ba2+ blocked the inward current in 2 mM K+, whereas 1 microM TTX or 0.3 mM Cd2+ did not affect the inward current. Inactivation of the inward currents, which became clear on extreme hyperpolarization, was suppressed with decreasing extracellular Na+ concentration. The present results suggest that carp olfactory epithelium is rich in the inward rectifier and is an excellent source of mRNA for cloning cDNA coding the inward rectifier.     

Repolarization of the presynaptic action potential and short-term synaptic plasticity in the chick ciliary ganglion

Stimulation-induced increases in synaptic efficacy have been described as being composed of multiple independent processes that arise from the activation of distinct mechanisms at the presynaptic terminal. In the chick ciliary ganglion, four components of short-term synaptic plasticity have been described: F1 and F2 components of facilitation, augmentation, and potentiation. In the present study, intracellular recording from the presynaptic calyciform nerve terminal of the chick ciliary ganglion revealed that the late repolarization and afterhypolarization (AHP) phases of the presynaptic action potential are affected by repetitive stimulation and that the time course of these effects parallel that of facilitation. The effects of these changes in the presynaptic action potential time course on calcium influx were tested by using the recorded action potential waveforms as voltage command stimuli during whole-cell patch-clamp recordings from acutely isolated chick ciliary ganglion neurons. The "facilitated" action potential waveform (slowed repolarization, decreased AHP amplitude) evoked calcium current with slightly but significantly greater total calcium influx. Taken together, these results are consistent with the hypothesis that activity-dependent changes in the presynaptic action potential are one of several mechanisms contributing to the facilitation phase of stimulation-induced increases in transmitter release in this preparation.     

TEA-sensitive potassium channels and inward rectification in regenerated rat sciatic nerve

Sucrose gap and intra-axonal recording techniques were used to identify the types of ion channels and inward rectification that are present in regenerated axons of adult (greater than 8 weeks) rat sciatic nerve after crush injury. In sucrose gap recordings, 4-aminopyridine (4-AP) led to slight broadening of the compound action potential (CAP) in normal nerve, and a greater broadening in regenerated nerves. By 12 days after sciatic nerve crush, regenerated nerves manifested an afterhyperpolarization (AHP) lasting up to 250 ms that was sensitive to tetraethylammonium (TEA). A similar TEA-sensitive AHP could be elicited with repetitive stimulation. Hyperpolarizing constant current steps (0.1 to 0.5 mA; 600-900 ms duration) applied across the sucrose gap through regenerated axons evoked membrane hyperpolarizations with a depolarizing, Cs(+)-sensitive relaxation in the response to hyperpolarization, which is characteristic of inward rectification, occurring after about 70 ms. The relaxation was present as early as 21 days after nerve crush. Intra-axonal recordings showed burst firing in 4-AP that was terminated by an AHP that temporally correlated with the TEA-sensitive AHP, and a relaxation in the response to hyperpolarizing current, similar to that of whole nerve recordings. The results demonstrate that in addition to voltage-sensitive sodium channels and 4-AP-sensitive potassium channels, there are TEA-sensitive and inwardly rectifying channels on mammalian regenerated peripheral nerve axons.     

Novel inward rectifier blocked by Cd2+ in crayfish muscle.

The characteristics of a voltage- and time-dependent inward rectifying current were examined with voltage clamp techniques in crayfish muscle. The inward current, carried by K+, was activated by hyperpolarization. Although this inward current increased with the extracellular K+ concentration [( K+]o), the voltage-dependence of the underlying conductance was independent of [K+]o. The current was unaffected by Cs+ and Ba2+, but was blocked by low concentrations of Cd2+. Therefore, this inward rectifier is different than previously described ones.     

Effect of nifedipine on neurons of guinea pig celiac ganglion.

The effect of nifedipine on electrophysiological membrane properties and nicotinic neurotransmission of guinea pig celiac ganglion neurons was studied using intracellular recordings in vitro. Nifedipine in concentrations of 0.1-10 microM did not affect membrane potential, membrane input resistance or the amplitude and duration of action potentials induced by intracellular current injection. Higher doses of nifedipine (0.1-1 mM) significantly reduced the amplitude and extended the duration of action potentials induced by intracellular current injection. Superfusion of the ganglia with nifedipine in concentrations of 0.1-10 microM significantly inhibited nicotinic fast excitatory postsynaptic potentials (f-EPSPs) and orthodromic action potentials evoked by nerve stimulation. This depressant effect of nifedipine on synaptic transmission was eliminated with high Ca2+ (12.5 mM). Nifedipine (10 microM) did not affect the postsynaptic effect of exogenous acetylcholine (ACh), but significantly reduced the quantal content but not the quantal size of evoked f-EPSPs in a low Ca2+ (0.5 mM), high Mg2+ (5.5 mM) Krebs solution. Nifedipine in concentration of 10 microM did not affect afterspike hyperpolarization (AH) and post-tetanic hyperpolarization (PTH), which have been recognized to be generated mainly by an increase of calcium-dependent potassium conductance. Higher doses of nifedipine (0.1-1 mM) significantly depressed AH and PTH. These experimental results suggest that nifedipine in concentrations of 0.1-10 microM exerts an inhibitory effect on nicotinic neurotransmission without affecting the membrane properties of the guinea pig celiac ganglion neurons. This inhibitory effect of nifedipine on synaptic transmission may result from blocking L-type calcium channels and reducing the quantal release of ACh from the presynaptic nerve terminals.     

Acetylcholine responses in snail neurons: Increase and decrease in potassium conductance succeeding inward currents

In the identified neurons B1 and B3 of the buccal ganglion of Helix pomatia, the initial acetylcholine (ACh) inward current was succeeded by two types of secondary responses. The secondary responses consisted either in an outward current or in a long-lasting inward current or in a combination of both. The secondary outward current was decreased with membrane hyperpolarization, associated with a decrease of membrane resistance and abolished in Ca2+-free Co2+ solution. It is assumed to be a K+ current activated by an influx of Ca2+. The secondary inward current also decreased with membrane hyperpolarization, but was associated with an increase of the membrane resistance and could be mimicked by an injection of Na+ into the cells. It is suggested to be due to a block of K+ channels by intracellular Na+. When the secondary responses appeared combined, the outward current preceded the inward current.     

Subcellular distribution of L-type Ca2+ channels responsible for plateau potentials in motoneurons from the lumbar spinal cord of the turtle

L-type calcium channels mediate the persistent inward current underlying plateau potentials in spinal motoneurons. Electrophysiological analysis shows that plateau potentials are generated by a persistent inward current mediated by low threshold L-type calcium channels located in the dendrites. As motoneurons express L-type calcium channels of the CaV1.2 and CaV1.3 subtypes, we have investigated the subcellular distribution of these channels using antibody labelling. The plateau generating a persistent inward current is modulated by the activation of metabotropic receptors. For this reason, we also examined the relationship between CaV1.2 and CaV1.3 subunits in motoneurons and presynaptic terminals labelled with antibodies against synapsin 1a. Motoneurons in the spinal cord of the adult turtle were identified as large neurons, immunopositive for choline acetyltransferase, located in the ventral horn. In these neurons, CaV1.2 subunits were present in the cell bodies and axons. Patches of CaV1.3 subunits were seen in association with the cell membrane of the somata and both the proximal and distal dendrites. Double labelling with an antibody against synapsin 1a showed that CaV1.3 subunits, but not CaV1.2 subunits, were always located at synaptic sites. The distribution of CaV1.2 and CaV1.3 strongly suggests that the persistent inward current underlying plateau potentials in spinal motoneurons is mediated by CaV1.3 and not by CaV1.2. Our findings also show that CaV1.3 may be located in the somatic and dendritic membrane adjacent to particular presynaptic terminals.     

Inward rectification in response to FMRFamide in Aplysia neuron L2: summation with transient K current

The response of Aplysia abdominal ganglion neuron L2 to the molluscan neuroactive peptide Phe-Met-Arg-Phe-NH2 (FMRFamide) was studied in voltage-clamp experiments. In all of the experiments, focal application of the peptide to the soma activated an inward rectifier current and reduced the apparent amplitude of the transient K current, IA. In a few cells, Na and K currents were activated in addition to these effects. Voltage-jump experiments were performed to study the ionic dependence, kinetics, and voltage dependence of the inward rectifier. Inward rectification increased exponentially during hyperpolarizing pulses and recovered exponentially on return to the resting potential. The reversal potential was variable, but was near -40 mV at the beginning of experiments. Inward rectification was insensitive to changes in external Na, Ca, or K concentration, but lowering the external Cl concentration had complicated effects on current amplitude. When KCl microelectrodes were used, perfusion with low-Cl external saline increased the amplitude of the peptide-dependent inward rectifier and shifted its reversal potential to a more positive voltage. With KAc microelectrodes, perfusion with low-Cl saline reduced the amplitude of the current. Inward rectification increased when a KAc microelectrode was withdrawn and replaced with a low-resistance KCl electrode, even when there was no measurable change in reversal potential. These results suggest that the FMRFamide-dependent inward rectifier is a Cl current that, like the current described by Chesnoy-Marchais (1982, 1983), is modulated by intracellular Cl. FMRFamide reduced the apparent amplitude of IA without affecting the voltage dependence of IA activation or inactivation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Localization of a FMRFamide-related peptide in efferent neurons and analysis of neuromuscular effects of DRNFLRFamide (DF2) in the crustacean Idotea emarginata

In the ventral nerve cord of the isopod Idotea emarginata, FMRFamide-immunoreactive efferent neurons are confined to pereion ganglion 5 where a single pair of these neurons was identified. Each neuron projects an axon into the ipsilateral ventral and dorsal lateral nerves, which run through the entire animal. The immunoreactive axons form numerous varicosities on the ventral flexor and dorsal extensor muscle fibres, and in the pericardial organs. To analyse the neuromuscular effects of a FMRFamide, we used the DRNFLRFamide (DF2). DF2 acted both pre- and postsynaptically. On the presynaptic side, DF2 increased transmitter release from neuromuscular endings. Postsynaptically, DF2 depolarized muscle fibres by approximately 10 mV. This effect was not observed in leg muscles of a crab. The depolarization required Ca2+, was blocked by substituting Ca2+ with Co2+, but not affected by nifedipine or amiloride. In Idotea, DF2 also potentiated evoked extensor muscle contractions. The amplitude of high K+ contractures was increased in a dose dependent manner with an EC50 value of 40 nm. In current-clamped fibres, DF2 strongly potentiated contractions evoked by current pulses exceeding excitation-contraction threshold. In voltage-clamped fibres, the inward current through l-type Ca2+ channels was increased by the peptide. The observed physiological effects together with the localization of FMRFamide-immunoreactive efferent neurons suggest a role for this type of peptidergic modulation for the neuromuscular performance in Idotea. The pre- and postsynaptic effects of DF2 act synergistically and, in vivo, all should increase the efficacy of motor input to muscles resulting in potentiation of contractions.     

Denervation-activated inward rectifier in frog slow skeletal muscle fibers

We tested whether the absence of an inward rectifier channel in slow skeletal muscle fibers of the frog is regulated by innervation. Normal and denervated slow fibers were identified according to their passive electrical properties. In current-clamp experiments, anomalous rectification was quantified as the ratio of effective resistances for hyperpolarizing and depolarizing pulses. In isotonic potassium solution, this ratio was 0.45 +/- 0.1 (n = 14) for twitch fibers, whereas slow fibers displayed linear behavior [ratio = 1.0 +/- 0.05 (n = 15)]. However, denervated slow fibers showed anomalous rectification (ratio, 0.48 +/- 0.07; n = 5). This finding was supported by voltage-clamp experiments in which denervated slow fibers displayed (1) an inward rectifier current during hyperpolarizing pulses, (2) an increase in this current when [K(+)](o) was increased, and (3) a current inhibition after application of Ba(2+). These results suggest that frog slow fibers, which normally do not possess inward rectifier channels, can express them after denervation.     

Unusual ultrastructural features in intrafascicular ganglion cells of the rat pelvic nerve

The incidental finding of four ectopic ganglion cells within the pelvic nerve of a normal rat prompted a thorough electron microscopic investigation of the ultrastructural features of these neurons. They were found to enwrap presynaptic terminals inside crater-like invaginations; the appositional surfaces were made more complex by the presence of slender dendritic appendages and sheet-like processes of glial cells. The presynaptic elements contained both clear and dense-cored vesicles, and appeared similar to those characterizing SIF (paraneuronal) cells. In addition, cilia were encountered in both the invaginated processes and most of the Schwann cells associated with the pre- and postsynaptic nerve cells and their processes. Overall, these features were deemed worth reporting because 1) of the unusual features of synaptic input from a SIF cell to a ganglion cell associated with the pelvic plexus, and 2) the ectopic ganglion cells possibly represent the sole example, other than ciliary neurones in the avian ciliary ganglion, of postsynaptic cells encasing presynaptic endings inside their perikarya.     

Hyperpolarization-activated (Ih) current in mouse vestibular primary neurons

The presence of a hyperpolarization-activated inward current (Ih) was investigated in mouse vestibular primary neurons using the whole-cell patch-clamp technique. In current-clamp configuration, injection of hyperpolarizing currents induced variations of membrane voltage with prominent time-dependent rectification increasing with current amplitudes. This effect was abolished by 2 mM Cs+ or 100 microM ZD7288. In voltage-clamp configuration, hyperpolarization pulses from -60 mV to -140 mV triggered a slow activating and non inactivating inward current that was sensitive to the two blockers, but insensitive to 5 mM Ba2+. Changing Na+ and K+ concentrations demonstrated that Ih current is carried by both these monovalent cations. This is the first demonstration of a Ih current in vestibular primary neurons.     

Neural control of the heart: developmental changes in ionic conductances in mammalian intrinsic cardiac neurons

The expression and properties of ionic channels were investigated in dissociated neurons from neonatal and adult rat intracardiac ganglia. Changes in the hyperpolarization-activated and ATP-sensitive K+ conductances during postnatal development and their role in neuronal excitability were examined. The hyperpolarization-activated nonselective cation current, Ih, was observed in all neurons studied and displayed slow time-dependent rectification. An inwardly rectifying K+ current, IK(IR), was present in a population of neurons from adult but not neonatal rats and was sensitive to block by extracellular Ba2+ Using the perforated-patch recording configuration, an ATP-sensitive K+ (KATP) conductance was identified in > or = 50% of intracardiac neurons from adult rats. Levcromakalim evoked membrane hyperpolarization, which was inhibited by the sulphonylurea drugs, glibenclamide and tolbutamide. Exposure to hypoxic conditions also activated a membrane current similar to that induced by levcromakalim and was inhibited by glibenclamide. Changes in the complement of ion channels during postnatal development may underlie observed differences in the function of intracardiac ganglion neurons during maturation. Furthermore, activation of hyperpolarization-activated and KATP channels in mammalian intracardiac neurons may play a role in neural regulation of the mature heart and cardiac function during ischaemia-reperfusion.     

Excitation of substantia nigra pars compacta neurones by 5-hydroxy-tryptamine in-vitro.

Incoming serotonergic fibres are known to make direct synaptic contact with dopamine-containing neurones in the substantia nigra pars compacta (SNc). However, the effects of 5-HT (5-hydroxytryptamine) on these cells have not been thoroughly investigated. In the present study we show that application of 10-50 microM 5-HT increases the firing frequency of SNc neurones in-vitro, and produces inward rectification in a voltage region negative to -50mV. This effect is sensitive to extracellular Cs+, but not to Ba2+, and has similar properties as the intrinsic inward rectifier current, Ih. Antagonists of the 5-HT1A and 5-HT2 receptors were inefficacious. It is concluded that 5-HT excites SNc neurones via an enhancement of the conductance underlying Ih.     

Sodium dependency of the inward potassium rectifier in horizontal cells isolated from the white bass retina

The ionic properties underlying the inwardly rectifying potassium current in cultured voltage-clamped white bass horizontal cells were studied. Anomalous rectification was apparent upon membrane hype rpolarization with a reversal potential depolarized from the predicted value of E_K In raised extracellular potassium, the current increased and the reversal potential shifted toward a more depolarized membrane potential. Solutions containing decreased sodium caused a rapid decrease in the inward rectifier current but only slightly affected the reversal potential. Extracellular cesium or barium caused a reversible voltage-dependent reduction of the inward current. We interpret these results to mean that the inward rectifying channel in white bass horizontal cells is mainly permeable to potassium ions, but is sodium dependent. It may shape the photoresponses of the horizontal cells and may contribute to a hyperpolarization activated conductance increase measured in situ.     

Proctolin activates an inward current whose voltage dependence is modified by extracellular Ca2+.

The pentapeptide proctolin modulates the activity of the rhythmic pattern generators in the crustacean stomatogastric nervous system. Proctolin strongly excites the lateral pyloric and the inferior cardiac neurons of the stomatogastric ganglion (STG), causing them to fire extended high-frequency bursts of action potentials (Hooper and Marder, 1987; Nusbaum and Marder, 1989a,b). We now report that proctolin depolarizes these cells maximally at membrane potentials close to the threshold for action potential generation. In voltage clamp, proctolin evokes an inward current, carried at least partially by Na+, that shows strong outward rectification. Removal of extracellular Ca2+ markedly increases the amplitude of the proctolin-evoked current and linearizes its current-voltage curve. The properties of the proctolin current make it ideally suited to contribute to the activity-dependent modulation of the pyloric network of the STG.     

Electrophysiological characterization of rat striatal neurons in vitro following a unilateral lesion of dopamine cells

The effects of a unilateral 6 to 19-week lesion of dopamine cells on the excitability of rat striatal neurons were investigated in vitro using the intracellularly recorded membrane properties of neurons obtained ipsilateral and contralateral to 6-hydroxydopamine (6-OHDA) injection sites. Neurons ipsilateral to the lesion site and in striatal tissue depleted of dopamine exhibited resting membrane potentials and membrane resistances similar to those recorded in contralateral striatal neurons. Denervation appeared to have no appreciable effect on the proportion of neurons exhibiting various patterns of neuronal spiking (repetitive, bursting, or single spike) evoked by depolarizing current pulses. Current-voltage determinations revealed nominal rectification in the majority of neurons and marked nonlinearty consistent with inward rectification at potentials hyperpolarized and depolarized to rest in a large proportion of the remaining neurons. Neurons ipsilateral to 6-OHDA lesion sites exhibited these relationships in the same proportion as contralateral control cells. However, ipsilateral neurons with nominal rectification exhibited an average rate constant for the early onset of small hyperpolarizing membrane transients which was significantly smaller than that of controls. This finding suggests that intrinsic membrane parameters regulating the excitability of certain striatal neurons may be under the influence of dopamine or other factors closely associated with nigrostriatal nerve terminals. Published 1993 Wiley-Liss, Inc.     

Presynaptic modulation of transmitter release by the early outward potassium current inAplysia

The mechanism involved in presynaptic modulation of transmitter release was studied in an identified synapse of Aplysia californica. Presynaptic hyperpolarization induces a decrease in he evoked postsynaptic potential amplitude. This is shown to be due to a reduction in the presynaptic spike amplitude during the hyperpolarization. The decreased presynaptic spike amplitude with hyperpolarization is explained s resulting from the superimposition of an early outward potassium current on the transient inward current. It is suggested that the presynaptic hyperpolarizing conditioning pulse decreases inactivation of the early outward current, which shunts the transient inward current. The superimposition of these two currents (transient inward current and the early outward current) induces a decrease in presynaptic spike amplitude, which in turn reduces the synaptic output from the terminal.