Presynaptic spike broadening reduces junctional potential amplitude

Presynaptic modulation of action potential duration may regulate synaptic transmission in both vertebrates and invertebrates. Such synaptic plasticity is brought about by modifications to membrane currents at presynaptic release sites, which, in turn, lead to changes in the concentration of cytosolic calcium available for mediating transmitter release. The 'primitive' neuromuscular junction of the jellyfish Polyorchis penicillatus is a useful model of presynaptic modulation. In this study, we show that the durations of action potentials in the motor neurons of this jellyfish are negatively correlated with the amplitude of excitatory junctional potentials. We present data from in vitro voltage-clamp experiments showing that short duration voltage spikes, which elicit large excitatory junctional potentials in vivo, produce larger and briefer calcium currents than do long duration action potentials, which elicit small excitatory junctional potentials.     

Calcitonin gene-related peptide regulates muscle acetylcholine receptor synthesis

Innervation of muscle by motoneurones induces the development of a characteristic, high density cluster of acetylcholine receptors (AChRs) at the neuromuscular junction. Studies in vitro show that the accumulation of AChRs at nerve-muscle contacts results from both increased insertion of new AChRs into the muscle plasma membrane beneath nerve terminals and redistribution of preexisting AChRs; these two modes of AChR accumulation may be separately controlled since factors have been identified that influence AChR redistribution but not synthesis. Although many aspects of muscle development are regulated by nerve-dependent muscle activity, junctional AChR clusters still develop when neuromuscular transmission is blocked by either curare or alpha-bungarotoxin, suggesting that their formation is mediated by nerve-derived trophic factors other than activity. A molecule immunologically related to calcitonin gene-related peptide (CGRP-I) has been found in motoneurones in a variety of mammals including man. Here we provide indirect evidence that CGRP-I may be a motoneurone-derived trophic factor that increases AChR synthesis at vertebrate neuromuscular junctions.     

Agonist-induced potentiation of acetylcholine sensitivity in denervated skeletal muscle

The entire surface membrane of denervated skeletal muscle is sensitive to the neuromuscular transmitter, acetylcholine (ACh), whereas in innervated muscle only the junctional area is sensitive. It has been proposed that this difference is due to a 'trophic' effect exerted by ACh in innervated muscle to keep the extrajunctional regions of the surface membrane insensitive to its depolarising action. Several studies have demonstrated an agonist-induced potentiation of ACh sensitivity, followed by desensitisation, at the endplate region of normal muscles. The potentiation has been attributed to a cooperative action of ACh on the receptors. Desensitisation of the extrajunctional regions of denervated muscles by ACh has also been described. We now provide evidence that the transmitter itself potentiates the ACh contracture and depolarisation responses of the denervated muscles of the rat in vitro and that it produces this effect by increasing the number of available ACh receptors on the surface membrane.     

Onset of neural function in the lateral line.

The development of the nervous system includes the formation of specific neuronal connections. Some insight into the mechanisms by which these connections are made may be obtained by determining the order in which the elements of a developing system begin to function and examining any changes that may occur in the system shortly after it begins functioning. I have investigated the lateral line of the South African clawed frog, Xenopus laevis and I have classified the afferent system into three elements. First, the transduction mechanism, which includes all components that couple the water vibrations to the release of transmitter from the hair cell. Second, the afferent synapse, at which there is transmitter release and subsequent postsynaptic conductance changes. And third, the afferent axon. I report here experiments suggesting that these elements become functional in the following order: first, the axon can conduct action potentials; second, transmission is established; third, the transduction mechanism can modulate transmitter release. This same sequence has been reported in a system related to the lateral line, the auditory pathway of mammals.     

A new cellular mechanism for coupling inputs arriving at different cortical layers

Pyramidal neurons in layer 5 of the neocortex of the brain extend their axons and dendrites into all layers. They are also unusual in having both an axonal and a dendritic zone for the initiation of action potentials. Distal dendritic inputs, which normally appear greatly attenuated at the axon, must cross a high threshold at the dendritic initiation zone to evoke calcium action potentials but can then generate bursts of axonal action potentials. Here we show that a single back-propagating sodium action potential generated in the axon facilitates the initiation of these calcium action potentials when it coincides with distal dendritic input within a time window of several milliseconds. Inhibitory dendritic input can selectively block the initiation of dendritic calcium action potentials, preventing bursts of axonal action potentials. Thus, excitatory and inhibitory postsynaptic potentials arising in the distal dendrites can exert significantly greater control over action potential initiation in the axon than would be expected from their electrotonically isolated locations. The coincidence of a single back-propagating action potential with a subthreshold distal excitatory postsynaptic potential to evoke a burst of axonal action potentials represents a new mechanism by which the main cortical output neurons can associate inputs arriving at different cortical layers.     

Modulation of intracortical synaptic potentials by presynaptic somatic membrane potential

Traditionally, neuronal operations in the cerebral cortex have been viewed as occurring through the interaction of synaptic potentials in the dendrite and soma, followed by the initiation of an action potential, typically in the axon. Propagation of this action potential to the synaptic terminals is widely believed to be the only form of rapid communication of information between the soma and axonal synapses, and hence to postsynaptic neurons. Here we show that the voltage fluctuations associated with dendrosomatic synaptic activity propagate significant distances along the axon, and that modest changes in the somatic membrane potential of the presynaptic neuron modulate the amplitude and duration of axonal action potentials and, through a Ca2+-dependent mechanism, the average amplitude of the postsynaptic potential evoked by these spikes. These results indicate that synaptic activity in the dendrite and soma controls not only the pattern of action potentials generated, but also the amplitude of the synaptic potentials that these action potentials initiate in local cortical circuits, resulting in synaptic transmission that is a mixture of triggered and graded (analogue) signals.     

Dendritic spikes as a mechanism for cooperative long-term potentiation

Strengthening of synaptic connections following coincident pre- and postsynaptic activity was proposed by Hebb as a cellular mechanism for learning. Contemporary models assume that multiple synapses must act cooperatively to induce the postsynaptic activity required for hebbian synaptic plasticity. One mechanism for the implementation of this cooperation is action potential firing, which begins in the axon, but which can influence synaptic potentiation following active backpropagation into dendrites. Backpropagation is limited, however, and action potentials often fail to invade the most distal dendrites. Here we show that long-term potentiation of synapses on the distal dendrites of hippocampal CA1 pyramidal neurons does require cooperative synaptic inputs, but does not require axonal action potential firing and backpropagation. Rather, locally generated and spatially restricted regenerative potentials (dendritic spikes) contribute to the postsynaptic depolarization and calcium entry necessary to trigger potentiation of distal synapses. We find that this mechanism can also function at proximal synapses, suggesting that dendritic spikes participate generally in a form of synaptic potentiation that does not require postsynaptic action potential firing in the axon.     

Membrane potential has no direct role in evoking neurotransmitter release

Neurons communicate by secreting a transmitter that excites or inhibits other neurons at synapses. The role of presynaptic membrane potential in triggering transmitter release is still controversial. In one view, presynaptic action potentials trigger the release by the entry of calcium ions into presynaptic terminals through voltage-dependent calcium channels. Calcium acts at high local concentrations at release sites near channel mouths to cause neurosecretion. An opposing view is that, in addition to elevating presynaptic calcium, presynaptic potential stimulates transmitter release by a distinct direct action. The relative importance of depolarization and calcium entry in neurosecretion cannot be determined because the two events are tightly linked. To delineate the roles of presynaptic potential and calcium entry in transmitter release, we have used nitr-5, a photolabile calcium chelator, and a voltage-clamp technique to control intracellular calcium and membrane potential independently at a synapse formed between cell bodies of cultured neurons of the fresh water snail Helisoma trivolvis. We found transmitter release occurred when presynaptic calcium levels were elevated to concentrations of a few micromolar, and that presynaptic voltage had no direct effect on neurosecretion.     

Presynaptic glycine receptors enhance transmitter release at a mammalian central synapse

Glycine and GABAA (gamma-aminobutyric acid A) receptors are inhibitory neurotransmitter-gated Cl- channels localized in postsynaptic membranes. In some cases, GABAA receptors are also found presynaptically, but they retain their inhibitory effect as their activation reduces excitatory transmitter release. Here we report evidence for presynaptic ionotropic glycine receptors, using pre- and postsynaptic recordings of a calyceal synapse in the medial nucleus of the trapezoid body (MNTB). Unlike the classical action of glycine, presynaptic glycine receptors triggered a weakly depolarizing Cl- current in the nerve terminal. The depolarization enhanced transmitter release by activating Ca2+ channels and increasing resting intraterminal Ca2+ concentrations. Repetitive activation of glycinergic synapses on MNTB neurons also enhanced glutamatergic synaptic currents, indicating that presynaptic glycine receptors are activated by glycine spillover. These results reveal a novel site of action of the transmitter glycine, and indicate that under certain conditions presynaptic Cl- channels may increase transmitter release.     

Long-term potentiation is associated with increases in quantal content and quantal amplitude.

Long-term potentiation (LTP) of synaptic transmission in CA1 neurons of the hippocampus, elicited by the conjunction of presynaptic firing and postsynaptic depolarization, is an important model of plasticity, which may underlie memory storage. Although induction of LTP takes place in the postsynaptic cell, it is not clear whether it is expressed through an enhancement of transmitter release or through an increased postsynaptic response to the same amount of transmitter. Analysis of the trial-to-trial amplitude fluctuations of synaptic signals, that is quantal analysis, gives an important insight into the probabilistic mechanisms of transmission, although attempts to apply it to the mode of expression of LTP have so far yielded inconsistent results, at least in part because they have relied on models of transmitter release that have not been confirmed experimentally. Here we report clear evidence for quantal fluctuation in a subset of cells. Induction of LTP in these cells causes abrupt increases in either quantal content or quantal amplitude, or both. This shows that two different mechanisms can underlie the maintenance of LTP.     

Hyperpolarization following activation of K+ channels by excitatory postsynaptic potentials

We have postulated that an excitatory postsynaptic potential (e.p.s.p.) may open voltage-sensitive K+ ('M') channels, in an appropriate depolarizing range, and that this could alter the e.p.s.p. waveform. Consequently, the fast e.p.s.p. in neurones of sympathetic ganglia, elicited by a nicotinic action of acetylcholine (ACh), could be followed by a hyperpolarization, produced by the opening of M channels during the depolarizing e.p.s.p. and their subsequent slow closure (time constant-150 mg). This introduces the concept that transmitter-induced p.s.ps may trigger voltage-sensitive conductances other than those initiating action potentials, and that in the present case this could produce a true post-e.p.s.p. hyperpolarization. (Some hyperpolarizations other than inhibitory postsynaptic potentials (i.p.s.ps) have been reported to follow e.p.s.ps.) We show here that this is so.     

A physiological role for GABAB receptors in the central nervous system

The role of GABA in synaptic transmission in the mammalian central nervous system is more firmly established than for any other neurotransmitter. With virtually every neuron studied, the synaptic action of GABA is mediated by bicuculline-sensitive GABAA receptors which selectively increase chloride conductance. However, it has been shown that GABA has a presynaptic inhibitory action on transmitter release that is insensiive to bicuculline and is selectively mimicked by baclofen. The receptors involved in this action are referred to as GABAB receptors, to distinguish them from the classic bicuculline-sensitive GABAA receptors. In hippocampal pyramidal cells an additional postsynaptic action of GABA and baclofen has been reported that is also insensitive to GABAA antagonists, and may be mediated by GABAB receptors on the postsynaptic neuron. This action of GABA and baclofen involves an increase in potassium conductance. Synaptic activation of pathways converging on hippocampal pyramidal cells results in a slow inhibitory postsynaptic potential which involves an increase in potassium conductance, and it has been suggested that GABAB receptors might be responsible for this synaptic potential. However, to establish convincingly that GABAB receptors are physiologically important in the central nervous system, a selective GABAB antagonist is required. Here we provide this missing evidence. Using the hippocampal slice preparation, we now report that the phosphonic acid derivative of baclofen, phaclofen, is a remarkably selective antagonist of both the postsynaptic action of baclofen and the bicuculline-resistant action of GABA, and that it selectively abolishes the slow inhibitory postsynaptic potential in pyramidal cells.     

Curare can open and block ionic channels associated with cholinergic receptors

Curare has long been regarded as a typical competitive antagonist of acetylcholine (ACh) at the vertebrate neuromuscular junction. Recently, however, it has been shown that curare can also block the channels opened by ACh at the frog neuromuscular junction as well as on rat and Aplysia neurones; moreover, curare is able to depolarize rat myotubes and thus behaves as an agonist for the cholinergic receptor of this preparation (see ref. 6). Using the single channel recording technique, we have now found that, on rat myotubes, curare can both open and block in the same cell the channels controlled by the cholinergic receptor.     

Postsynaptic translation affects the efficacy and morphology of neuromuscular junctions

Long-term synaptic plasticity may be associated with structural rearrangements within the neuronal circuitry. Although the molecular mechanisms governing such activity-controlled morphological alterations are mostly elusive, polysomal accumulations at the base of developing dendritic spines and the activity-induced synthesis of synaptic components suggest that localized translation is involved during synaptic plasticity. Here we show that large aggregates of translational components as well as messenger RNA of the postsynaptic glutamate receptor subunit DGluR-IIA are localized within subsynaptic compartments of larval neuromuscular junctions of Drosophila melanogaster. Genetic models of junctional plasticity and genetic manipulations using the translation initiation factors eIF4E and poly(A)-binding protein showed an increased occurrence of subsynaptic translation aggregates. This was associated with a significant increase in the postsynaptic DGluR-IIA protein levels and a reduction in the junctional expression of the cell-adhesion molecule Fasciclin II. In addition, the efficacy of junctional neurotransmission and the size of larval neuromuscular junctions were significantly increased. Our results therefore provide evidence for a postsynaptic translational control of long-term junctional plasticity.     

用昆虫谷氨酸电位检测磷酰胺酯化合物的活性

采用电生理细胞内微电极记录果蝇３龄幼虫神经－肌肉接点的兴奋性接点电位（ｅｘｃｉｔａｔｏｒｙ ｊｕｎｃｔｉｏｎａｌ ｐｏｔｅｎ－ｔｉａｌｓ，ＥＪＰ），观察加药前后ＥＪＰ幅值的 变化以及电位是否被阻断及阻断时间，实验结果初步表明：１８种磷酰胺酯ＢＰＯＮ系列均可阻断ＥＪＰ，但它们的阻断时间相关很大，从７．２５ｍｉｎ到４３．００ｍｉｎ。这些化合物对谷氨酸电位的作用可分为三种：１．电位逐渐被阻断；２．电位先     

Action potential refractory period in ureter smooth muscle is set by Ca sparks and BK channels

In excitable tissues the refractory period is a critical control mechanism preventing hyperactivity and undesirable tetani, by preventing subsequent stimuli eliciting action potentials and Ca2+ entry. In ureteric smooth muscle, peristaltic waves that occur as invading pacemaker potentials produce long-lasting action potentials (300-800 ms) and extraordinarily long (more than 10 s) refractory periods, which prevent urine reflux and kidney damage. For smooth muscles neither the mechanisms underlying the refractory period nor the link between excitability and refractoriness are properly understood. Here we show that a negative feedback process, which depends on Ca2+ loading the sarcoplasmic reticulum (SR) during the action potential and on the subsequent activation of local releases of Ca2+ from the SR (sparks), stimulating plasmalemmal Ca2+-sensitive K+ (BK) channels, determines the refractory period of the action potential. As sparks gradually reduce the Ca2+ load in the SR, electrical inhibition is released, the refractory period is terminated and peristaltic contractions occur again. The refractory period can be manipulated, for example from 10 s to 100 s, by altering the Ca2+ content of the SR or release mechanism or by inhibiting BK channels. This insight into the control of excitability and hence function provides a focus for therapies directed at pathologies of smooth muscle.     

Are baclofen-sensitive GABAB receptors present on primary afferent terminals of the spinal cord?

The site of action of the antispastic drug baclofen has long been considered to reside in the spinal cord although supraspinal effects have also been reported. This beta-chlorophenyl derivative of the neurotransmitter gamma-aminobutyric acid (GABA) depresses both monosynaptic and polysynaptic transmission in the cord possibly through a decrease in transmitter release rather than by any antagonism at postsynaptic receptors. Recently, baclofen has been shown to be a selective ligand for a bicuculline-insensitive GABA receptor (GABAB) site that occurs widely in the mammalian central nervous system including the spinal cord. The apparent importance of the cord in the therapeutic effects of this drug prompted us to ask whether they involve GABAB site activation. As an initial step we have located these receptors by autoradiography, comparing them with classical GABAA sites. We report here that GABAB sites, unlike GABAA sites, are present in high concentrations in laminae I, II, III and IV of the dorsal horn and that after the neonatal administration of capsaicin this binding is reduced by 40-50%.     

Dual-component NMDA receptor currents at a single central synapse

Present thinking about the way that the NMDA (N-methyl-D-aspartate) class of glutamate receptor operates at central synapses relies mainly on information obtained from single-channel and whole-cell recordings from cultured neurons stimulated by exogenous NMDA receptor agonists. The mechanisms that operate in the postsynaptic membrane of a normal neuron following release of the natural transmitter are far less clear. An important problem is that most normal neurons receive many excitatory synapses (10(3)-10(5) per cell) and these synapses are located on slender dendritic elements far away from the somatic recording site, making the study of discrete synaptic events difficult. Typically, when populations of synapses are activated, NMDA receptor-mediated synaptic potentials appear as slowly rising, long-lasting waves superimposed on faster, non-NMDA-receptor potentials. Although believed to be critical for NMDA receptor function, this slow time-course would not be predicted from single-channel kinetics and its origin remains puzzling. We have now analysed the events occurring at the level of a single excitatory synapse using a simple, small, neuron--the cerebellar granule cell--which has an unusually simple glutamatergic input. By applying high-resolution whole-cell recording techniques to these cells in situ, we were able to study the nature of elementary NMDA receptor-mediated synaptic currents. Contrary to expectations, the prominent currents are fast but are followed by slow ones. Both types of current are strongly voltage-dependent but differ subtly in this respect. Furthermore, the currents are absent unless glycine is provided.     

Localization of dystrophin to postsynaptic regions of central nervous system cortical neurons

Moderate non-progressive cognitive impairment is a consistent feature of Duchenne muscular dystrophy (DMD), although no central nervous system (CNS) abnormality has been identified. Recent studies have elucidated the molecular defect in DMD, including the absence of the protein dystrophin in affected individuals. Normal brain tissue contains dystrophin messenger RNA and dystrophin is present in low abundance in the brain and seems to be regulated in this tissue, at least in part, by a promoter that differs from that in muscle. Until now, antibodies and immunocytochemical methods used to demonstrate dystrophin at the plasma membrane of mouse and human muscle have proven inadequate to localize precisely dystrophin in the mammalian CNS. We have now made an antibody (anti 6-10) which is much more sensitive than those previously available to immunolabel dystrophin in the CNS. Using this antibody, we found that in the mouse, dystrophin is particularly abundant in the neurons of the cerebral and cerebellar cortices, and that it is localized at postsynaptic membrane specializations. Dystrophin may have a different role in neurons than in muscle, and an alteration at the synaptic level may be the basis of the cognitive impairment in DMD.