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M M Poo

Publications and source records attributed to M M Poo.

At least 37 records · Page 2Linked to original sources
PubMed1996

Essential role of filopodia in chemotropic turning of nerve growth cone induced by a glutamate gradient.

Pathfinding of growing neurites depends on turning of the growth cone in response to extracellular cues. Motile filopodia of the growth cone are known to be critical for mediating contact-dependent guidance of the growth cone. However, whether filopodia also play an essential role in growth cone turning response induced by a diffusible chemotropic substance is unclear. Growth cones of cultured Xenopus spinal neurons exhibited chemotropic turning responses in a gradient of glutamate within a limited range of concentrations. This turning response depends on the activation of the NMDA subtype of glutamate receptors and requires the presence of extracellular Ca2+. Time-lapse differential interference contrast microscopy with quantitative analysis of filopodia dynamics showed a close correlation between an increased number of filopodia on the side of the growth cone facing the glutamate source and the turning. Such filopodia asymmetry was observed within minutes after the onset of the glutamate gradient, before any detectable turning of the growth cone. In Ca(2+)-free medium, no filopodia asymmetry was induced by the glutamate gradient, and no growth cone turning was observed. Furthermore, elimination of filopodia with a low concentration of cytochalasin B completely abolished the turning response without substantially affecting neurite extension. Thus, filopodia may be required for chemotropic guidance of the growth cone, and an asymmetry in filopodia distribution may be an early cellular event responsible for determining the direction the growth cone advances.

Actins↗
PubMed1996

Expression of synapsin I correlates with maturation of the neuromuscular synapse.

Synapsins are a family of neuron-specific phosphoproteins that are localized within the presynaptic terminals in adult brain. Previous work has demonstrated that introduction of exogenous synapsins I(a + b) or IIa into Xenopus spinal neurons promoted maturation of the neuromuscular synapse in a nerve-muscle co-culture system. We have now studied the expression of endogenous Xenopus synapsin I during synaptic maturation in vivo and in culture, using a polyclonal antibody raised against Xenopus synapsin I. Immunoprecipitation experiments indicated that synapsin I was not detectable during the early phase of synaptogenesis in vivo, and exhibited a marked increase during the period of synaptic maturation. In contrast, the expression of synaptophysin, another synaptic vesicle protein, was detected at the start of nervous system formation, and remained at a high level thereafter. Similar expression profiles for the two proteins were also observed in immunocytochemical studies of Xenopus spinal neurons in culture: intense staining of synaptophysin was found on the first day, while synapsin I was not detected until after three days in culture. The expression of synapsin I correlated very well with the appearance of a bell-shaped amplitude distribution of spontaneous synaptic currents, a physiological parameter which reflects functional maturation of the neuromuscular synapse. In one-day-old cultures grown in the absence of laminin, an extracellular matrix protein known to be present at the neuromuscular junction, the amplitude distribution of virtually all synapses was skewed towards smaller values. In contrast, when laminin was used as a culture substrate, many synapses exhibited a bell-shaped amplitude distribution. Laminin treatment also induced synapsin I expression in one-day-old cultures. These results suggest that the expression of endogenous synapsin I may regulate maturation at neuromuscular synapses.

Animals↗
PubMed1996

Postsynaptic elevation of calcium induces persistent depression of developing neuromuscular synapses.

Synaptic activity is known to modulate neuronal connectivity in the nervous system. At developing Xenopus neuromuscular synapses in culture, repetitive postsynaptic application of ACh near the synapse leads to immediate and persistent synaptic depression, which was shown to be caused by reduction of presynaptic evoked transmitter release. However, little depression was found when ACh was applied to the muscle 20 microns or further from the synapse. Fluorescence imaging of cytosolic Ca2+ ([Ca2+]i) showed that each ACh pulse induced a transient elevation of myocyte [Ca2+]i that spread approximately 20 microns. Local photoactivated release of Ca2+ from the caged Ca2+ chelators nitr-5 or nitrophen in the postsynaptic cell was sufficient to induce persistent synaptic depression. These results support a model in which localized Ca2+ influx into the postsynaptic myocyte initiates transsynaptic retrograde modulation of presynaptic secretion mechanisms.

Acetylcholine↗
PubMed1996

Calcium and chemotropic turning of nerve growth cones.

Cultured Xenopus spinal neurons exhibit chemotropic turning toward the source of neurotransmitters acetylcholine and glutamate. Here we review the experimental evidence that transmitter-induced turning of the growth cone is mediated by an influx of Ca2+, that a gradient of intracellular Ca2+ within the growth cone is responsible for the directional growth cone response, and that asymmetric filopodia formation precedes and is essential for the turning response.

Animals↗
PubMed1995

Potentiation of transmitter release by ciliary neurotrophic factor requires somatic signaling.

Neurotrophic factors participate in the development and maintenance of the nervous system. Application of ciliary neurotrophic factor (CNTF), a protein that promotes survival of motor neurons, resulted in an immediate potentiation of spontaneous and impulse-evoked transmitter release at developing neuromuscular synapses in Xenopus cell cultures. When CNTF was applied at the synapse, the onset of the potentiation was slower than that produced by application at the cell body of the presynaptic neuron. The potentiation effect was abolished when the neurite shaft was severed from the cell body. Thus, transmitter secretion from the nerve terminals is under immediate somatic control and can be regulated by CNTF.

Acetylcholine↗
PubMed1995

Accelerated structural maturation induced by synapsin I at developing neuromuscular synapses of Xenopus laevis.

The role of synapsin I, a synaptic vesicle-associated phosphoprotein, in the maturation of nerve-muscle synapses was investigated in nerve-muscle co-cultures prepared from Xenopus embryos loaded with the protein by the early blastomere injection method. The stage of maturation of the synapses was analysed by electron microscopy as well as by whole-cell patch-clamp recording. The acceleration in the functional maturation of neuromuscular synapses induced by synapsin I was accompanied by a profound rearrangement in the ultrastructure of the nerve terminal. Nerve terminals formed by synapsin I-loaded neurons were characterized by a higher number of small synaptic vesicles organized in clusters and predominantly localized close to the nerve terminal plasma membrane, a smaller number of large dense-core vesicles and no significant change in the number of coated vesicles. Precocious development of active zone-like structures as well as deposition of basal lamina into the synaptic cleft were also observed at these synapses. These results support a role for synapsin I in the architectural changes which occur during synaptogenesis and lead to the maturation of quantal neurotransmitter release mechanisms.

Animals↗
PubMed1995

Calcium-dependent transmitter secretion from fibroblasts: modulation by synaptotagmin I.

Following endocytic uptake of acetylcholine (ACh), CHO fibroblasts exhibit Ca(2+)-dependent spontaneous quantal ACh release and depolarization-evoked ACh release, as detected by a whole-cell voltage-clamped myocyte in contact with the fibroblast. CHO fibroblasts transfected with synaptotagmin I, an integral membrane protein of synaptic vesicles, showed a reduced spontaneous quantal ACh release and an enhanced Ca(2+)-evoked ACh release, as compared with control cells. Biochemical and ultrastructural studies of endocytic activity using horseradish peroxidase as a marker further confirmed the inhibitory action of synaptotagmin I on spontaneous vesicular exocytosis and on elevated exocytosis induced by Ca2+. Through inhibition of exocytosis at the resting intracellular concentration of Ca2+ and removal of the inhibition upon depolarization-induced Ca2+ entry, synaptotagmin I could enhance the efficiency of excitation-secretion coupling.

Acetylcholine↗
PubMed1995

Plasticity of developing neuromuscular synapses.

Developing neuromuscular synapses are susceptible to modulation by the presence of synaptic activity and a number of chemical factors originated from either pre- or postsynaptic cells. In vitro studies of functional modulation of synaptic strength by electrical stimulation of pre- and postsynaptic cells suggested an essential role of retrograde interactions at developing synapses.

Animals↗
PubMed1995

Spontaneous quantal transmitter secretion from myocytes and fibroblasts: comparison with neuronal secretion.

When exogenous ACh is loaded into the cytoplasm of cultured amphibian myocytes and fibroblasts, the cells undergo spontaneous quantal ACh secretion, as detected by the appearance of pulsatile membrane currents in Xenopus myocytes which are manipulated into contact with the cells. These currents resemble in many ways the miniature endplate currents (MEPCs) observed at developing neuromuscular synapses formed on these Xenopus myocytes. Analyses of the frequency, amplitude, and time course of these currents suggests similarity in the cellular mechanisms involved in the packaging and secretion of ACh quanta in fibroblasts, myocytes, and developing neurons. The size of the ACh packets released by the non-neuronal cells were found to be very similar to the size of the neuronal ACh quanta, which are thought to result from the exocytotic release of synaptic vesicles. Moreover, the kinetics with which the ACh packets are discharged from all three cell types are comparable, although the speed of secretion in non-neuronal cells is somewhat slower and more irregular. The spontaneous quantal ACh secretion from neurons and myocytes was decreased by reducing cytosolic Ca2+ level and enhanced by activation of protein kinase C with phorbol ester, but secretion from fibroblasts was unaffected by both treatments. The spontaneous secretion from fibroblasts did show some sensitivity to a rise in cytosolic Ca2+ after treatment with a Ca2+ ionophore. These observations support the hypothesis that the basic machinery for transmitter secretion operating in neurons derive from a more ubiquitous mechanism used for constitutive secretion and membrane trafficking in non-neuronal cells, and neuronal differentiation involves expression of additional unique components for the regulation of the spontaneous quantal secretion.

Acetylcholine↗
PubMed1995

Overexpression of synaptophysin enhances neurotransmitter secretion at Xenopus neuromuscular synapses.

Previous studies have suggested the importance of synaptophysin (p38), a major integral membrane protein of the synaptic vesicle, in transmitter secretion, but few have directly addressed its functional role at intact synapses. In the present study, injection of synthetic mRNA for synaptophysin into one of the early blastomeres of a Xenopus embryo resulted in elevated synaptophysin expression in 1 and 2 d embryos and in cultured spinal neurons derived from the injected blastomere, as shown by immunocytochemistry. At neuromuscular synapses made by neurons overexpressing synaptophysin [p38(+)] in 1 d cell cultures, the spontaneous synaptic currents (SSCs) showed a markedly higher frequency, as compared to control synapses. This increase in frequency was not accompanied by a change in the mean amplitude or the amplitude distribution of the SSCs, suggesting that synaptophysin is not involved in determining the size of transmitter quanta. The impulse-evoked synaptic currents (ESCs) of synapses made by p38(+) neurons showed increased amplitude as well as reduced fluctuation and delay of onset of ESCs. Under high-frequency tetanic stimulation at 5 Hz, the rate of tetanus-induced depression was faster for p38(+) neurons. Taken together, these results suggest a role for synaptophysin in the late steps of transmitter secretion, affecting the probability of vesicular exocytosis and/or the number of synaptic vesicles initially docked at the active zone.

Animals↗
PubMed1994

Study on the induction of spontaneous transmitter release at early nerve-muscle contacts in Xenopus cultures.

During synaptogenesis, spontaneous acetylcholine release is rapidly induced by the contact of the growing neurite with the myocyte. Using Xenopus nerve-muscle co-cultures, we investigated the role of Ca2+ in the presynaptic mechanisms mediating this induction process. We found that Ca2+ influx is not necessary to induce spontaneous release and that an increase in cytosolic Ca2+ is not sufficient to trigger this release in the absence of the target cell.

Acetylcholine↗
PubMed1994

Synapsin IIa accelerates functional development of neuromuscular synapses.

We have investigated the possible involvement of the synaptic vesicle protein synapsin IIa in synapse development. Synapsin IIa was introduced into Xenopus embryonic spinal neurons by early blastomere injection, and nerve-muscle cultures were prepared. Synaptic currents were measured by comparing synapses in which the presynaptic neuron either contained [syn IIa (+)] or lacked (control) exogenous synapsin IIa. Syn IIa (+) synapses had a 3.6-fold increase in the frequency and a 2.1-fold increase in the amplitude of spontaneous synaptic currents, compared to controls, after 2 days in culture. Synapsin IIa also increased the amplitude of evoked synaptic currents by 2.3-fold in 2-day cultures. The evoked synaptic current amplitudes of syn IIa (+) synapses had a lower coefficient of variation indicating a more stable evoked response. These enhanced synaptic activities were independent of the presence or absence of the protein in the postsynaptic muscle cell. The findings indicate a role for synapsin IIa in synapse maturation.

Acetylcholine↗
PubMed1994

Turning of nerve growth cones induced by neurotransmitters.

Pathfinding by growing nerve processes in the developing nervous system depends on the turning response of the growing tip, the growth cone, to extracellular guidance cues. There is evidence in vivo and in cell culture that some growth cones exhibit chemotropic behaviour, but the identity of endogenous chemoattractants remains elusive. Neurotransmitters appear early in the developing embryo and may have morphogenic roles in development. In cell culture a number of neurotransmitters were found to induce growth inhibition or retraction of neurites. Here we report positive turning responses of the nerve growth cone in a defined extracellular gradient of the neurotransmitter acetylcholine (ACh). The growth cone response depends on the activation of neuronal nicotinic ACh receptors, requires the presence of extracellular Ca2+, and appears to be mediated by Ca(2+)-calmodulin-dependent protein kinase II. Fluorescence imaging of cytosolic Ca2+ concentration ([Ca2+]i) at the growth cone showed a small but significant evaluation of [Ca2+]i within minutes of the onset of ACh application and before the turning of the growth cone. These findings suggest that neurotransmitters may serve as specific chemoattractants for growth cone guidance and that cytosolic Ca2+ may act as a second messenger in the cytoplasm of the growth cone to initiate the turning response.

Acetylcholine↗
PubMed1994

Calcium-dependent postsynaptic exocytosis: a possible mechanism for activity-dependent synaptic modulation.

Elevation of cytosolic Ca2+ level in the postsynaptic cell is critical for the induction of many forms of activity-dependent synaptic modulation. Based on our recent evidence that in muscle cells and fibroblasts constitutive exocytosis is increased by elevating cytosolic Ca2+ levels, we hypothesize that Ca(2+)-dependent exocytosis at the postsynaptic site may provide a mechanism for a localized, activity-dependent synaptic modulation.

Acetylcholine↗
PubMed1994

Evoked neuronal secretion of false transmitters.

The ability of differentiated neurons to secrete false transmitters in response to depolarization was examined by loading exogenous transmitters into the neuronal cytoplasm with a whole-cell recording pipette. We found that within minutes following loading of exogenous glutamate into the cytoplasm of cholinergic Xenopus spinal neurons, depolarization-evoked glutamate secretion could be detected by an acutely dissociated hippocampal pyramidal neuron manipulated into contact with the spinal neuron. Similarly, when ACh was loaded into a glutamatergic hippocampal neuron, evoked ACh secretion could be detected by a myocyte. The evoked secretion of the false transmitter was Ca2+ dependent and appeared to be vesicular in nature. These results suggest that differentiated neurons are capable of packaging and secreting multiple nonpeptide transmitters, provided that sufficient concentrations of the transmitters are available in the cytoplasm.

Acetylcholine↗
PubMed1994

Retrograde interactions during formation and elimination of neuromuscular synapses.

Maturation of neuromuscular synapses depends on dynamic interactions between presynaptic motor neurons and postsynaptic muscle cells. Recent studies have addressed the cellular mechanisms underlying these interactions in cell cultures and in developing animals. Retrograde signals from the postsynaptic muscle cells appear to play critical roles in all stages of synapse development, from the initial synaptogenesis to the stabilization or elimination of the synapse.

Animals↗