Effects of ethanol and other drugs on excitatory and inhibitory neurotransmission in the crayfish.

1. Crayfish exposed to 434 mM ethanol (EtOH) showed signs of hyperactivity within 0.5-2 h, at which times crayfish hemolymph EtOH concentration had reached 60-90 mM. 2. A 10-min exposure to 60-90 mM EtOH reduced presynaptic inhibition of excitatory postsynaptic currents (EPSCs) at the crayfish opener neuromuscular junction (NMJ) in vitro but did not significantly alter excitatory neurotransmission. The same concentrations of EtOH did not alter other potentials or currents associated with inhibition at this synapse, such as presynaptic inhibitory potentials (PIPs), inhibitory postsynaptic potentials (IPSPs), and inhibitory postsynaptic currents (IPSCs). 3. Intermediate EtOH concentrations (120-180 mM) applied for 10 min in vitro reduced the amplitude of excitatory postsynaptic potentials (EPSPs) by decreasing the membrane resistance of opener muscle fibers and by reducing the amplitude of EPSCs. 4. High EtOH concentrations (434 mM) applied for 10 min in vitro had yet greater depressive effects on measures of postsynaptic properties described above. The time course of EPSCs was also significantly reduced. In addition, presynaptic properties such as action-potential (AP) amplitude and frequency of spontaneous release of neurotransmitter were reduced by 434 mM EtOH. 5. Presynaptic inhibition, gamma-aminobutyric acid (GABA; 250-500 microM), muscimol (50 microM), and baclofen (75 microM) all reduced the depolarizing afterpotential of APs in the excitor axon and reduced EPSPs in opener muscle fibers. GABA (500 microM) and baclofen (75 microM) significantly reduced presynaptic AP amplitudes, whereas presynaptic inhibition, GABA (250 microM), and muscimol (50 microM) had no effect on AP amplitude. Bicuculline (250-500 microM), a GABAA antagonist, did not entirely eliminate presynaptic inhibition, whereas picrotoxin (50 microM), another GABAA antagonist, completely removed presynaptic inhibition. Thus presynaptic inhibitory mechanisms may involve both GABAA and GABAB receptors on the opener excitor axon. 6. Our data suggest that the behavioral hyperactivity seen at hemolymph EtOH concentrations of 60-90 mM is not accompanied by a change in excitatory synaptic transmission observed at the opener NMJ. Rather, crayfish hyperactivity may be due to depressive effects of EtOH on inhibitory synapses in the CNS similar to the disinhibition evoked by EtOH at the opener NMJ.     

New types of synaptic connections in crayfish stretch receptor organs: an electron microscopic study

Summary The synaptic input to crayfish (Orconectes limosus) stretch receptor neurons, and the synaptic interactions between the inhibitory and excitatory efferents were analysed by electron microscopy of serial sections. Several novel types of synaptic connections have been observed: (i) inhibitory synaptic input on the axon hillock and initial axon segment; (ii) serial synaptic terminals on the sensory cell body; (iii) simultaneous synaptic contacts of the same inhibitory terminal with sensory dendrites and muscle fibres; (iv) reciprocal synapses between the two types of inhibitory efferents; and (v) inhibitory synapses on the primary inhibitory axon. The possible functional significance of these synapses is discussed in the light of earlier electrophysiological and pharmacological findings.Fellow of the Alexander von Humboldt-Stiftung.     

Synaptic strength and horseradish peroxidase uptake in crayfish nerve terminals

Summary Uptake of horseradish peroxidase was studied by examining percentages of labelled synaptic vesicles in nerve endings of the excitatory axon innervating the opener muscle of the walking leg in the crayfish (Procambarus clarkii). Terminals on fibres with large excitatory postsynaptic potentials (EPSP) had higher percentage of labelled vesicles than terminals of fibres with small EJPs. The extent of labelling in the synaptic vesicle pool was greater for terminals with higher transmitter output. Evidence for three possible routes of synaptic vesicle formation was found. Movement of vesicles within the terminal as a whole appeared to be constrained, but rapid movement of vesicles within local populations probably occurs.     

Excitatory potentials induced by stimulation of the inhibitory axon at the crustacean neuromuscular junction.

Synaptic events in a chloride-deficient condition were studied to elucidate functional aspects of presynaptic inhibitory synapses. The extracellular junctional potentials and nerve terminal potentials were concurrently recorded from a synaptic region. Inhibitory stimulation produced repetitive spikes on the inhibitory nerve terminal and then the excitatory nerve terminal, which resulted in the extracellular excitatory junctional potentials. Excitatory stimulation did not produce repetitive spikes on the inhibitory nerve terminal, indicating one-way signal transmission in this axo-axonal synapse from inhibitory to excitatory axon. The interval required for an inhibitory stimulation to produce the first response in the postsynaptic muscle membrane ranged widely from 10 to 800 msec. When gamma-aminobutyric acid (GABA, 1 times 10-minus 4 M) was added in these experimental conditions, the muscle membrane was transiently depolarized by about 10 mV. The action of GABA mimics that of the neurotransmitter at presynaptic inhibitory synapses. The experimental observations may be well explained by the concept of synapses on synapses, i.e., presynaptic inhibition, where the neurotransmitter may be GABA and chloride ions may be playing essential roles as in the case of postsynaptic inhibition.     

Ultrastructure of identified fast excitatory, slow excitatory and inhibitory neuromuscular junctions in the locust

Summary The specialized jumping muscle of the locust, the metathoracic extensor tibiae (ETi), is innervated by four physiologically different motoneurons, including FETi, a phasic excitor, SETi, a tonic excitor, and CI, a tonic common inhibitor. FETi neuromuscular junctions were examined in three phasic ETi bundles innervated by FETi. FETi terminals were characterized by patchy contacts on to granular sarcoplasm. The ETi accessory extensor, innvervated by both SETi and CI, contains two morphologically different types of axon ending. When this muscle was soaked in horseradish peroxidase, stimulation of SETi led to selective uptake in vesicles in terminals similar to those of FETi axons but containing smaller vesicles, while stimulation by CI caused increased uptake into terminals with more extensive contact directly on to fibrillar sarcoplasm. As has been observed in excitatory and inhibitory synapses in some crustacean and vertebrate nervous systems, the synaptic vesicles in the locust excitatory endings are round and electron-lucent while those in the inhibitory endings are more irregular in shape. The tonic neuromuscular junctions, SETi and CI, are more densely packed with vesicles, larger in cross-sectional area and appear to be of more complex shape than the smaller, vesicle-sparse, phasic FETi terminals. Following long duration stimulation at 10 Hz, the tonic neuromuscular junctions showed little morphological change. FETi endings, which fatigue within minutes at the same stimulation frequency, showed a 20% decrease in synaptic vesicle density and an increase in irregularly shaped membrane inclusions.     

Correlation of presynaptic and postsynaptic events during establishment of long-term facilitation at crayfish neuromuscular junction

Repetitive stimulation (10-20 Hz) of the motor axon supplying the opener muscle in the crayfish leg produces long-lasting enhancement of excitatory postsynaptic potentials. This long-term facilitation (LTF) was investigated by recording simultaneously from the presynaptic nerve terminal and from the innervated muscle fiber with intracellular microelectrodes. On cessation of stimulation, the facilitated postsynaptic potential declines in amplitude when monitored with low-frequency test stimuli. A rapid decline (phase I) occurs over the first 30 s and is succeeded by a more gradual decline lasting several minutes (phase II). Finally, a residual potentiation with a very slow decay (phase III) persists for several hours. Simultaneous pre- and postsynaptic recordings were made during induction of LTF with stimuli delivered at 20 Hz for 10 min. During the tetanus, excitatory postsynaptic potentials were enhanced 20-fold, while action potentials in the presynaptic terminal declined in amplitude from 108.6 to 97.2 mV, and the presynaptic membrane became hyperpolarized by 6.4 mV. The Na+ pump inhibitor ouabain (0.5-1.0 mM) abolished the hyperpolarization, indicating that the latter resulted from activation of an electrogenic Na+ pump. The reduction in amplitude of the presynaptic action potential was consistent with a reduced transmembrane concentration gradient for Na+. Thus, it is suggested that a significant accumulation of Na+ occurs during repetitive stimulation of crayfish motor axons. Decay of phase II of LTF, but not of phases I or III, had approximately the same time course as the decay of Na+ accumulation in the terminals, monitored by changes in the presynaptic action potential. Thus it is probable that in crayfish this phase of LTF is linked to an increased intraterminal Na+ concentration. Injection of Na+ from a microelectrode into the presynaptic terminal produced enhancement of the excitatory postsynaptic potential lasting for many minutes, as well as changes in presynaptic membrane potential and action potential similar to those seen during repetitive stimulation. The results provide the first direct measurements of electrical and ionic changes in axonal terminals during prolonged periods of activity leading to LTF, and support the hypothesis that accumulation of intraterminal Na+ is associated with one phase of LTF.     

Long-term facilitation alters transmitter releasing properties at the crayfish neuromuscular junction

Synaptic transmission at the neuromuscular junction of the excitatory axon supplying the crayfish opener muscle was examined before and after induction of long-term facilitation (LTF) by a 10-min period of stimulation at 20 Hz. Induction of LTF led to a period of enhanced synaptic transmission, which often persisted for many hours. The enhancement was entirely presynaptic in origin, since quantal unit size and time course were not altered, and quantal content of transmission (m) was increased. LTF was not associated with any persistent changes in action potential or presynaptic membrane potential recorded in the terminal region of the excitatory axon. The small muscle fibers of the walking-leg opener muscle were almost isopotential, and all quantal events could be recorded with an intracellular microelectrode. In addition, at low frequencies of stimulation, m was small. Thus it was possible to apply a binomial model of transmitter release to events recorded from individual muscle fibers and to calculate values for n (number of responding units involved in transmission) and p (probability of transmission for the population of responding units) before and after LTF. In the majority of preparations analyzed (6/10), amplitude histograms of evoked synaptic potentials could be described by a binomial distribution with a small n and moderately high p. LTF produced a significant increase in n, while p was slightly reduced. The results can be explained by a model in which the binomial parameter n represents the number of active synapses and parameter p the mean probability of release at a synapse. Provided that a pool of initially inactive synapses exists, one can postulate that LTF involves recruitment of synapses to the active state.     

Presynaptic membrane potential and transmitter release at the crayfish neuromuscular junction

Release of transmitter was evoked at neuromuscular junctions of the crayfish opener muscle by passage of current through an intracellular electrode impaling a branch of the motor axon close to a muscle fiber. Membrane-potential changes in the presynaptic axon branch were monitored, together with postsynaptic potentials. Depolarization of impaled secondary axonal branches by more than 10 mV led to an increase in asynchronous transmitter release. The release was facilitated by prolonged (50-500 ms) depolarizations and it decayed rapidly when depolarization was terminated. Ca2+ was essential for facilitated release; however, no indication of a Ca spike was found at the recording site. Input-output curves for the synapse were obtained by applying depolarizing pulses of varying amplitude to the axon branch. Transmitter output was strongly influenced by both amplitude and duration of the applied depolarization. During normal synaptic transmission, propagated Na+-dependent action potentials were recorded in the secondary axonal branches but there was no evidence for a calcium-dependent component for these action potentials. Evoked release was dependent on Ca2+ and was steeply dependent on the amplitude of the action potential, which could be made variable in size by application of tetrodotoxin (TTX). Prolonged depolarization of axonal branches resulted in enhancement of transmitter release evoked by an action potential. The enhancement occurred in spite of a simultaneous reduction of the amplitude of the action potential. Morphological features of the terminals were investigated after injection of lucifer yellow into the axon. An electrical model incorporating the morphological features suggests that membrane-potential changes set up in the main axon reach the nearest terminals with 30-40% attenuation, while events originating in the terminals would be severely attenuated in the main axon. Comparison of the crayfish synapse with other frequently studied synapses shows both similarities and differences, suggesting that it is not possible to apply findings made in one synapse to all others.     

Novel mechanism for presynaptic inhibition: GABA(A) receptors affect the release machinery

Presynaptic inhibition is produced by increasing Cl(-) conductance, resulting in an action potential of a smaller amplitude at the excitatory axon terminals. This, in turn, reduces Ca(2+) entry to produce a smaller release. For this mechanism to operate, the "inhibitory" effect of shunting should last during the arrival of the "excitatory" action potential to its terminals, and to achieve that, the inhibitory action potential should precede the excitatory action potential. Using the crayfish neuromuscular preparation which is innervated by one excitatory axon and one inhibitory axon, we found, at 12 degrees C, prominent presynaptic inhibition when the inhibitory action potential followed the excitatory action potential by 1, and even 2, ms. The presynaptic excitatory action potential and the excitatory nerve terminal current (ENTC) were not altered, and Ca(2+) imaging at single release boutons showed that this "late" presynaptic inhibition did not result from a reduction in Ca(2+) entry. Since 50 microM picrotoxin blocked this late component of presynaptic inhibition, we suggest that gamma-aminobutyric acid-A (GABA(A)) receptors reduce transmitter release also by a mechanism other than affecting Ca(2+) entry.     

Intramembranous organization of lobster excitatory neuromuscular synapses

Summary The fine structure of identified neuromuscular synapses of the single excitatory axon to the distal accessory flexor muscle in lobster limbs was examined with freeze-fracture and serial thin-section electron microscopy. The latter technique reveals presynaptic dense bars with synaptic vesicles aligned on either side of these bars and often fused to the membrane, suggesting exocytosis and confirming our previous contention that these bars are active zones of transmitter release. The intramembranous organization of these active zones, as revealed in freeze-etched tissue, is a ridge-like elevation of the P-face of the axolemma with a matching trough on the complementary E-face. The ridge on the P-face has rows of large scattered intramembranous particles along the apex and is often bordered by a series of small, circular depressions which are presumed to represent exocytotic vesicles attached to the presynaptic membrane. Complementing these depressions are a few volcano-like protuberances seen occasionally on the E-face membrane. Because such evidence for transmitter release occurred in both stimulated and non-stimulated preparations, it demonstrates that chemical fixatives employing aldehydes induce transmitter release. The postsynaptic receptor sites of these excitatory synapses are characterized by oval-shaped patches of densely packed particles on the E-face, arranged in a random pattern on the sarcolemma. The complementary P-face view exhibits a regular square array of particle imprints or pits.     

GABA-immunoreactivity in processes presynaptic to the terminals of afferents from a locust leg proprioceptor

Summary Individually labelled sensory neurons from the femoral chordotonal organ, a proprioceptor at the femoro-tibial joint of a locust hindleg, were analysed by intracellular recording, and by electron microscopical immunocytochemistry to reveal the arrangement of their input and output synapses and to determine whether the input synapses were GABAergic. Intracellular recordings from these sensory neurons show spikes superimposed on a barrage of synaptic potentials during movements of the femoro-tibial joint. These synaptic inputs can be mimicked by GABA. Input synapses are made onto the vesicle-containing terminals of afferents and are often closely associated with the output synapses. By contrast, the axons of the afferents in the neuropil have no vesicles and neither make nor receive synapses. The input synapses to the afferent terminals are made from processes typically a few microns in diameter, whereas the output synapses are made onto much smaller processes of only 0.1–0.2 m. Input synapses at which an afferent terminal is the only postsynaptic element are common. Where the synapse is dyadic the second postsynaptic element does not usually appear to be a chordotonal afferent. The output synapses from the afferent terminals are usually dyadic. At 78% of the input synapses, the presynaptic neurite showed immunoreactivity to a GABA antibody, supporting the physiological evidence that the presynaptic effects can be mediated by the release of GABA. The remaining (22%) immunonegative synapses are intermingled with those showing GABA immunoreactivity, but their putative transmitter is unknown. These morphological observations suggest that the presynaptic control of the chordotonal afferents is largely mediated by GABAergic neurons, but because other types of neuron also appear to be involved, presynaptic modulation may be more complex than has yet been revealed by the physiology.     

Impulse activity of a crayfish motoneuron regulated its neuromuscular synaptic properties

1. Previous studies have demonstrated that initial transmitter release, fatigability, and the morphology of identified crayfish neuromuscular synapses adapt to long-term changes in motoneuron impulse activity. 2. Experiments were performed to determine whether these long-term, adaptive alterations in neuromuscular synaptic physiology are triggered by changes in neuromuscular synaptic activity, muscle activity, or neuronal impulse activity. The fast closer excitor of the crayfish claw, a phasic motoneuron, was studied. Either the central or the peripheral region of the motoneuron was selectively stimulated in vivo by blocking impulse activity midway along the motor axon with localized application of tetrodotoxin and stimulating either central or distal to the blocked region. 3. Neither muscle activity nor transmitter release from the neuromuscular synapses was required to trigger the changes in synaptic physiology. Stimulation central to the block induced changes in neuromuscular transmission that included a long-lasting decrease in initial transmitter release and increased fatique resistance. 4. Because peripheral stimulation also produced decreased initial transmitter release, it appears that increased impulse activity in either region of the motoneuron can produce the synaptic changes. These results along with earlier findings suggest that neuronal depolarization induces adaptive, long-term changes in synapses. 5. These results are discussed in relation to findings at vertebrate and invertebrate synapses.     

Different kinds of axon terminals forming symmetric synapses with the cell bodies and initial axon segments of layer II/III pyramidal cells. II. Synaptic junctions

Summary Four different types of axon terminals form symmetric synapses with the cell bodies and initial axon segments of pyramidal cells in layer II/III of rat visual cortex. One type belongs to chandelier cells, and the other three kinds of terminals have origins that have not been established yet. These latter are referred to as large, medium-sized and dense terminals. The purpose of the present study was to examine the synaptic junctions formed by all four types of terminal. The synapses formed by the chandelier cell terminals are readily recognized in thin sections because of the characteristic features of both the terminals and the initial axon segments, which are the neuronal elements postsynaptic to them. In en face views of these axo-axonal synapses the junctions can be seen to have presynaptic dense projections that form a grid in which they are triagonally spaced, and have an average centre-to-centre spacing of 84 nm. As an ensemble the projections form the presynaptic grid, which usually has an oval or round outline, but may be notched on one side where projections are absent. The synaptic junctions of the large, medium-sized and dense terminals were examined by making reconstructions of the terminals from serial thin sections. It was found that at the interfaces between the axon terminals and the cell bodies of pyramidal cells, several separate synaptic junctions may be present, in addition to a number of puncta adhaerentia. Thus, there may be as many as five separate synaptic junctions and as few as one. It was also found that while the proportion of the area of the synaptic interface occupied by synaptic junctions was between 12% and 26% for dense terminals, for medium sized terminals it was 10–15%, and for the one large terminal reconstructed it was only 8%. Thus, there can be multiple synaptic junctions between each of these types of axon terminals and a pyramidal cell, and because many of the terminals forming symmetric junctions are boutons en passant, a number of vesicle release sites exist between the presynaptic axon and its postsynaptic partner. The axon terminals forming symmetric synapses in the cerebral cortex are assumed to be inhibitory, and consequently it is suggested that this arrangement of multiple release sites is designed to ensure that stimulation of the presynaptic axon results in an effective level of hyperpolarization of the postsynaptic neuron.     

High rates of excitatory miniature currents in crayfish claw opener muscle evoked by high concentrations of gamma-aminobutyric acid (GABA) in normal and Ca2+-deficient superfusions

High concentrations (0.5 mol/l) of the neutral amino acid GABA were used to evoke release of transmitter quanta from excitatory terminals at voltage clamped crayfish muscle fibres in normal and Ca²⁺-deficient superfusions. An experiment in which the release of transmitter quanta proceeded at high rates in both normal and Ca²⁺-deficient superfusion was analyzed in detail indicating a Ca²⁺-independent mechanism of release. In the normal superfusion, on application of GABA, the release rates ñ increased within a few seconds up to about 6000 quanta/s and thereafter declined exponentially with a time constant $\text{τ}{}_{\mathrm{<mtext>q</mtext>}}\text{) = 18.5}\text{s}$, most likely due to depletion of a readily releasable store of transmitter in the excitatory nerve terminals comprising at least 110,000 quanta per muscle fibre. Assuming that about 1900 excitatory synapses exist per muscle fibre [9], it results that about 58 quanta can be associated with each synapse in agreement with morphological data [15] which show that between 47–117 vesicles exist in a single glutamatergic synapse of crayfish.     

Ultrastructure of the segmental giant neuron of crayfish

The ultrastructure of the crayfish segmental giant (SG) neuron is described, and compared to other identified and unidentified crayfish neurons. The SG was specifically stained by intracellular injection of horseradish peroxidase and is divided into four regions of interest. In the dorsal region, finger-like dendrites of the SG make contact with the through-conducting giant fibres (GF). These contacts are physiologically defined rectifying electrical synapses. They are characterized by the presence of 30-95 nm agranular vesicles in the presynaptic GFs, some postsynaptic density in the SG, and a narrowing of the intermembrane cleft to approximately 5 nm. There is little evidence for connecting cytoplasmic bridges. Unidentified neurons make chemical input with either round or elliptical vesicle types onto SG bottlenecks close to the electrical synapses. Ventral to the GFs, dendritic profiles of the SG make three sorts of contact with unidentified neurons. (a) Regions of close membrane apposition (approximately 5 nm) are presumed to be electrical output synapses, but there are no vesicles such as at the input synapses, and, again, little sign of connecting bridges. (b) Chemical input is received from unidentified presynaptic neurons containing either round or elliptical vesicles. These synapses are characterized by 30-75 nm presynaptic agranular vesicles, widened cleft (approximately 20 nm), granular cleft material and postsynaptic density. There is no sign of any presynaptic density. (c) Very occasional SG profiles containing vesicles and making output synapses to unidentified neurons occur. In the lateral neuropil at the edge of the ganglion the SG gives rise to a small tuft of very fine dendrites. These are nearly all laden with vesicles and ramify in a complex region of neuropil containing many small profiles which are also vesicle-laden. The SG axon diminishes in diameter as it progresses along its peripheral nerve root, and finally terminates at a blind ending near the base of the swimmerets. It is sheathed along its entire length, and there is no sign of vesicles within it. We conclude that the SG axon makes no peripheral output.     

家兔下橄榄背侧副核的超微结构特点——电镜研究

本实验对7只家兔的下橄揽背侧副核进行了电镜观察。除发现已报道过的轴树突触、轴体突触及以树突为中心的突触球外,还见此核内有轴轴突触、轴突与胞体棘形成的突触,以轴突为中心的突触球及核内微纤维束。轴树突触最常见,突触后成分为树突或棘。轴体突触较少见。有一轴突终末与一胞体棘形成突触,同时还与一树突形成突触。轴轴突触前、后成分中均含圆形囊泡,有时形成轴-轴-树突触串。家兔下橄榄背侧副核内见有两种突触球,一种以树突为中心,另一种以轴突为中心。还发现两个轴突终末同时与另一轴突终末形成轴→轴→轴突触。下橄榄背侧副核内的复杂突触形式,说明传入冲动在其中可能经过扩散、会聚、突触前抑制及整合等复杂的过程。     

Effect of calcium on excitatory neuromuscular transmission in the crayfish

1. The effects of varying the external Ca concentration from 1.8 to 30 mM/l. ((1/8)-2 times normal) have been studied at the in vitro crayfish excitatory neuromuscular junction. Electrophysiological techniques were used to record transmembrane junctional potentials from muscle fibres and extracellular junctional currents from the vicinity of nerve terminals.2. The excitatory junctional potential amplitude was proportional to [Ca](0) (n), where n varied between 0.68 and 0.94 (mean 0.82) when [Ca](0) was varied from 1.8 to 15 mM/l.3. The increase in junctional potential amplitude on raising [Ca](0) resulted primarily from an increase in the average number of quanta of excitatory transmitter released from the presynaptic nerve terminal by the nerve impulse.4. The size of the quanta, synaptic delay, presynaptic potential and electrical properties of the muscle membrane were little affected by changes in calcium concentration in the range studied.     

Phosphatidylinositol system's role in serotonin-induced facilitation at the crayfish neuromuscular junction

1. In a crustacean neuromuscular preparation, the walking leg opener muscle of the freshwater crayfish Procambarus clarkii, application of serotonin (1 microM) produces presynaptic depolarization and long-lasting facilitation of excitatory postsynaptic potentials (EPSPs). The frequency of spontaneously released transmitter quanta also increases. Facilitation of evoked EPSPs declines after serotonin application in two phases. 2. Serotonin-induced facilitation was examined using simultaneous pre- and postsynaptic intracellular microelectrode recording. A presynaptic microelectrode recorded action potentials and membrane potential of a presynaptic axonal branch, and one or more postsynaptic microelectrodes recorded EPSPs in muscle fibers innervated by the excitatory motor axon. Components of the phosphatidylinositol second messenger system and pharmacologic agents affecting this system were injected through the presynaptic electrode, and changes in synaptic transmission were measured. 3. Presynaptic injection of inositol 1,4,5-triphosphate (IP3) causes presynaptic depolarization, increases the frequency of spontaneously released transmitter quanta, and promotes a relatively short-lasting facilitation of evoked EPSPs. These actions are consistent with elevation of intracellular Ca2+ and resemble the early phase of serotonin-induced facilitation. 4. Application of a phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), that activates protein kinase C (C-kinase), produces a long-lasting, low-level facilitation of evoked EPSPs. Application of another phorbol ester, phorbol-12-monoacetate (PTMA), which does not activate C-kinase has no effect. 5. Presynaptic injection of RA 233, a phospholipase C (PLP-C) inhibitor, blocks all aspects of serotonin-induced facilitation. This compound was found to have no general deleterious effects on synaptic transmission and does not block other forms of synaptic facilitation in this preparation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mechanisms of presynaptic inhibition studied using paired-pulse facilitation

An investigation was made of the effect of presynaptic inhibition on paired-pulse facilitation (PPF) of group Ia afferent excitatory postsynaptic potentials (EPSPs). The main finding from this study was that PPF was enhanced during presynaptic inhibition of compound Ia EPSPs. This increase in PPF is identical to that seen at other synapses when the probability of transmitter release is decreased by lowering the extracellular calcium or raising the extracellular magnesium concentration, providing unequivocal evidence that presynaptic inhibition is associated with a decrease in the probability of transmitter release. Further, by analogy with the effects of reduced calcium influx on PPF at other synapses, the results support the idea that presynaptic inhibition is associated with reduced calcium influx into nerve terminals.     

Axon hillocks and initial segments in spinal trigeminal nucleus with emphasis on synapses including axo-axo-axonic contacts

Summary As a part of a continuing study of the feline spinal trigeminal nucleus, the fine structure and synaptic arrangements on the axon hillock and axon initial segment of neurons in this region are described here. Transmission electron microscopy has been used to characterize qualitatively the axon hillock and initial segment and associated synapses in pars interpolaris. Axon hillocks and initial segments are easily identified in continuity with somata or as isolated profiles in the neuropil, and they receive synaptic contacts: these we regard as axo-axonic. The presynaptic terminals contain either mainly round or mainly flattened synaptic vesicles and have Type I (asymmetric) or Type II (symmetric) thickenings respectively at their contacts with the axon hillock or initial segment. I report here also the unusual arrangement of three separate axons in a serial synaptic complex. Some of the round vesicle Type I contacts onto the axon hillock-initial segment region also receive Type II contacts from one or more flattened vesicle terminals, thus formingan axo-axo-axonic complex. These flattened vesicle terminals lack the usual features of a presynaptic dendrite. It has been shown that in this nucleus some round vesicle terminals, especially those postsynaptic to flattened vesicle terminals, are primary afferents from the periphery. Therefore the round vesicle terminal presynaptic to the axon hillock-initial segment region, some of which are included in the axo-axo-axonic complex may also be a primary afferent directly contacting the spike generator area of the relay neuron and under presynaptic control of a flattened vesicle synapse. The latter may possibly be an intrinsic contact. This strategic situation of round vesicle terminals and the axo-axo-axonic complex at the axon hillock or initial segment has major implications relevant to the overall output of these neurons.