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Jordi Molgó - One of the best experts on this subject based on the ideXlab platform.
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Vesicle size and transmitter release at the frog neuromuscular junction when quantal acetylcholine content is increased or decreased.
The Journal of Physiology, 2002Co-Authors: William Van Der Kloot, Jordi Molgó, Roger Cameron, Cesare ColasanteAbstract:We investigated whether the synaptic vesicles at the neuromuscular junction change size when their acetylcholine (ACh) content is altered. The size of the Miniature Endplate Potential (MEPP) increased 3- or 4-fold in preparations pre-treated in a hypertonic solution in which the anion was gluconate. We measured the dimensions of synaptic vesicles in such preparations and in controls. The size of the vesicles and size distribution were indistinguishable. Quanta contained about half of the usual amount of ACh in preparations stimulated in the presence of hemicholinium-3, an inhibitor of choline uptake, or in NH(4)(+), which diminishes the proton gradient for ACh uptake into the vesicles. Neither treatment changed the size of the synaptic vesicles. ACh content and vesicle size were both decreased in preparations stimulated in (-)-vesamicol, an inhibitor of ACh uptake in vesicles. Since the other inhibitors decreased ACh content by a similar amount without altering vesicle size, (-)-vesamicol may decrease vesicle size by acting on another target. We also found that a hypertonic solution in which the anion was aspartate increased quantal size similar to gluconate. Both anions have high hydration energy and a large volume. When these treatments increased quantal size the mean 20-80 % rise time of MEPPs recorded with an extracellular electrode was 170 micros. In the controls it was 97 micros. Perhaps some of the added ACh is bound within the vesicles, which slows the rise. Our major conclusion is that ACh content can change notably without any change in the size of the synaptic vesicles.
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Recycling and refilling of transmitter quanta at the frog neuromuscular junction.
The Journal of Physiology, 2000Co-Authors: William Van Der Kloot, Roger Cameron, Cesare Colasante, Jordi MolgóAbstract:1. Fluorescent dyes have been used at the frog neuromuscular junction to label synaptic vesicular membrane. Retrieved membrane is reformed into vesicles, which are released along with pre-existing vesicles. Consequently, if vesicular refilling with acetylcholine (ACh) is depressed by inhibitors, two sizes of quanta should be released: normal and smaller. As recycling continues the fraction of smaller size quanta should increase exponentially. 2. We enhanced the rate of quantal release by elevating the K+ concentration. The principal inhibitors were (-)-vesamicol (VES), hemicholinium-3 (HC3), and NH4+. Quantal size measurements were fitted to one and to two cumulative lognormal probability distribution functions. When two fitted better, the statistical significance assessment took into account the three additional parameters used in calculating the fit. 3. After recycling in the presence of inhibitor, many sets were fitted better by two lognormal functions. As recycling continued, the fraction of the Miniature Endplate Potential voltage-time integrals ( MEPPs) in the larger sub-population decreased exponentially. 4. The size of the releasable pool was estimated by counting the quanta released by carbonyl cyanide m-chlorophenylhydrazone (CCCP). This was compared to pool sizes calculated from the inhibitor experiments. The two estimates of pool size were indistinguishable, with mean values ranging from about 170,000 to 270,000. 5. With all of the treatments tested, the means of the sizes in the smaller sub-population of MEPPs were about 1/3 those of the larger sub-populations. 6. Recycling synaptic vesicles appear to be incorporated into the releasable pool from which they have roughly the same probability of release as the pre-existing vesicles.
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A new conotoxin isolated from Conus consors venom acting selectively on axons and motor nerve terminals through a Na + -dependent mechanism
European Journal of Neuroscience, 1999Co-Authors: Frédéric Le Gall, Evelyne Benoit, Philippe Favreau, César Mattei, Françoise Bouet, Jean-louis Menou, André Ménez, Yves Letourneux, Jordi MolgóAbstract:A novel conotoxin was isolated and characterized from the venom of the fish-hunting marine snail Conus consors. The peptide was identified by screening chromatography fractions of the crude venom that produced a marked contraction and extension of the caudal and dorsal fins in fish, and noticeable spontaneous contractions of isolated frog neuromuscular preparations. The peptide, named CcTX, had 30 amino acids and the following scaffold: X11CCX7CX2CXCX3C. At the frog neuromuscular junction, CcTx at nanomolar concentrations selectively increased nerve terminal excitability so that a single nerve stimulation triggered trains of repetitive or spontaneous synaptic Potentials and action Potentials. In contrast, CcTx had no noticeable effect on muscle excitability even at concentrations 100 x higher than those that affected motor nerve terminals, as revealed by direct muscle stimulation. In addition, CcTx increased Miniature Endplate Potential (MEPP) frequency in a Ca2+-free medium supplemented with ethylene glycol-bis-(beta-aminoethyl ether)-N,N,N', N'-tetraacetic acid (EGTA). Blockade of voltage-dependent sodium channels with tetrodotoxin (TTX) either prevented or suppressed the increase of MEPP frequency induced by the toxin. CcTx also produced a TTX-sensitive depolarization of the nodal membrane in single myelinated axons giving rise, in some cases, to repetitive and/or spontaneous action Potential discharges. In addition, CcTx increased the nodal volume of myelinated axons, as determined using confocal laser scanning microscopy. This increase was reversed by external hyperosmolar solutions and was prevented by pretreatment of axons with TTX. It is suggested that CcTx, by specifically activating neuronal voltage-gated sodium channels at the resting membrane Potential, produced Na+ entry into nerve terminals and axons without directly affecting skeletal muscle fibres. CcTx belongs to a novel family of conotoxins that targets neuronal voltage-gated sodium channels.
J. Vautrin - One of the best experts on this subject based on the ideXlab platform.
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Hypertonic treatment reversibly increases the ratio of giant skew-Miniature Endplate Potentials to bell-Miniature Endplate Potentials.
Neuroscience, 1996Co-Authors: M.e. Kriebel, F. Llados, J. VautrinAbstract:Abstract Miniature Endplate Potentials were recorded from single frog muscle fibers before, during and after treatment with hypertonic saline (200–500 mM NaCl or Na gluconate added to frog saline). Miniature Endplate Potential amplitude distributions were plotted from small muscle fibers so that the modes and ratios of the skew-Miniature Endplate Potential to bell-Miniature Endplate Potential classes could be defined. Muscle fibers were voltage clamped with two electrodes to determine the input resistance before, during and after treatment. Input resistance increased from two to 100 times during treatment and rapidly fell towards control values (no more than 30% greater) when preparations were returned to normal frog saline. Short duration treatments with 200–300 mM hypertonic salines immediately increased frequencies (100-fold) of both skew-Miniature Endplate Potential and bell-Miniature Endplate Potential classes. Preparations when returned to normal frog saline after a few minutes of treatment showed control Miniature Endplate Potential distributions within minutes. One to two hour treatment left only the skew-Miniature Endplate Potential class and with hour-long recovery periods bell-Miniature Endplate Potentials reappeared and ratios of skew-Miniature Endplate Potential to bell-Miniature Endplate Potential classes returned to control values. Treatment with 500 mM NaCl added to frog saline immediately increased the percentage of skew-Miniature Endplate Potentials (from 2 to 50%) with little or no increase in overall Miniature Endplate Potential frequencies. The mode of the skew-Miniature Endplate Potential class was unchanged after hypertonic treatment, whereas that of the bell-Miniature end plate Potential class either remained about the same size or decreased depending on the duration of treatment. The number and percentage of giant-Miniature Endplate Potentials belonging to the skew-Miniature Endplate Potential class increased as a function of the duration of 200–300 mM hypertonic saline treatments. Most giant-Miniature Endplate Potentials had a slow rising phase with a foot and/or breaks demonstrating a composite structure. Sequentially recorded giant-Miniature Endplate Potentials had similar initial slopes indicating either repetitive releases from single sites or releases from cooperative sites. After hypertonic treatment the bell-Miniature Endplate Potential size was never more than that expected with the increase (under 30%) in input resistance. The results presented here are completely different from those of Yu and Van der Kloot [(1991)J. Physiol.433, 677–704] who reported that the bell-Miniature Endplate Potential amplitude was increased two- to four-fold after hypertonic treatment. The wide range of results in the ratio of skew-Miniature Endplate Potential to bell-Miniature Endplate Potential classes is discussed in regards to the quantal hypothesis which is based on a single class of immutable amounts of transmitter; and, a hypothesis based on a dynamical process that meters transmitter in subunit amounts to control Miniature Endplate Potential size and class during release.
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Miniature Endplate Potentials induced by ammonium chloride, hypertonic shock, and botulinum toxin.
Journal of neuroscience research, 1992Co-Authors: J. VautrinAbstract:Intracellular recordings were made at the neuromuscular junction (NMJ) of the mouse diaphragm to study alteration of Miniature Endplate Potential (MEPP) amplitude and rise time after different treatments. Following either hyperosmotic shock or 3 to 5 min of incubation in 10 to 50 mM ammonium chloride (NH4Cl) (replacing NaCl, a treatment which is known to raise intracellular pH) MEPP frequencies increased and the amplitudes of. MEPPs decreased. These treatments as well as type A botulinum toxin (BoTx) gradually prolonged the rising phase of some MEPPs, which increased their time-to-peak (slow-MEPPs; Vautrin and Kriebel: Neuroscience 41:71–88, 1991) and increased eventually their amplitude. Fasciculation after hyperosmotic shock or during NH4C1 challenge was blocked by D-tubocurarine and was due to large slow-MEPPs that reached threshold for the muscle fiber action Potential. The development of fasciculation provided the time course for the development of giant-MEPPs. Increased frequency of giant MEPP is accompanied by a block of the nerveevoked muscle contraction. Effects of BoTx on spontaneous release were functionally antagonized either by NH4C1 or hyperosmotic shock. NH4C1 delayed BoTx blockage of bell-MEPPs. Data suggest that BoTx alters the formation of transmitter packets gradually but similarly to other treatments which increase incidence of skew-MEPPs.
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Characteristics of slow-Miniature Endplate currents show a subunit composition
Neuroscience, 1991Co-Authors: J. Vautrin, Mahlon E. KriebelAbstract:Abstract The normal neuromuscular junction shows two classes of spontaneous Miniature Endplate Potentials. These classes are based on a discontinuity in the profile of Miniature Endplate Potential amplitude distributions. The amplitude of one class of Miniature Endplate Potentials from a bell-shaped amplitude distribution and the remaining Miniature Endplate Potentials compose a population which forms a left-hand skew distribution with a mode1/7 to1/10 that of the bell-Miniature Endplate Potentials [Kriebel M. E. and Gross C. E. (1974) J. gen. Physiol.64, 85–103]. Some skew-Miniature Endplate Potentials have a slow time-to-peak and show breaks on the rising phase. Most treatments that alter the Miniature Endplate Potential frequency change the ratio of skew-Miniature Endplate Potentials/bell-Miniature Endplate Potentials [Kriebel M. E. et al. (1976) J. Physiol.262, 553–581]. The time characteristics of Miniature Endplate currents were readily altered in the isolated frog and mouse neuromuscular junctions with several agents known to increase the percentage of slow-Miniature Endplate Potentials (heat, botulinum toxin, 4-aminoquinoline and increases in bath osmolarity). The slow-Miniature Endplate Potential amplitudes were a continuum of amplitudes from skew- to giant Miniature Endplate Potentials. The rising phases of Miniature Endplate Potentials were a continuum from smooth to many with breaks and offsets. In a series of sequentially recorded slow-Miniature Endplate currents, many had congruent rising phases of constant slope regardless of amplitude or of time-to-peak. The rising phases of congruent slow-Miniature Endplate currents which showed a change in slope deviated at similar amplitudes. The least value of the slope of a slow-Miniature Endplate current was that of the sub-Miniature Endplate current; and, Miniature Endplate currents with overall lower slope values showed a wave pattern and/or irregular breaks which suggests summation of sequentially delayed sub-Miniature Endplate currents. Plots of the amplitude vs time-to-peak of Miniature Endplate currents from identified junctions demonstrated that the normal percentage of slow-Miniature Endplate currents was greatly increased with the treatments used here and that the time-to-peak of giant Miniature Endplate currents usually was longer than that of normally occurring bell-Miniature Endplate currents. Giant Miniature Endplate currents with short time-to-peak values are probably from two Miniature Endplate currents occurring, by chance, almost simultaneously. During and/or after treatments, Miniature Endplate currents formed clusters of similar size Miniature Endplate currents, not randomly distributed in time, which graded from distinct Miniature Endplate currents to giant Miniature Endplate currents. Slow-Miniature Endplate currents with and without breaks on their rising phases were simulated with a computer using subunits that were combined by various numbers at various rates. Computer-generated Miniature Endplate currents simulate with fidelity the complete range of normal and treatment-induced slow-Miniature Endplate currents. We conclude that most slow-Miniature Endplate currents ranging in size from skew- to giant Miniature Endplate currents are fast bursts of dependent Miniature Endplate currents of the skew class, and that the treatments used here act in a non-specific manner to increase slow-Miniature Endplate currents.
Cesare Colasante - One of the best experts on this subject based on the ideXlab platform.
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Vesicle size and transmitter release at the frog neuromuscular junction when quantal acetylcholine content is increased or decreased.
The Journal of Physiology, 2002Co-Authors: William Van Der Kloot, Jordi Molgó, Roger Cameron, Cesare ColasanteAbstract:We investigated whether the synaptic vesicles at the neuromuscular junction change size when their acetylcholine (ACh) content is altered. The size of the Miniature Endplate Potential (MEPP) increased 3- or 4-fold in preparations pre-treated in a hypertonic solution in which the anion was gluconate. We measured the dimensions of synaptic vesicles in such preparations and in controls. The size of the vesicles and size distribution were indistinguishable. Quanta contained about half of the usual amount of ACh in preparations stimulated in the presence of hemicholinium-3, an inhibitor of choline uptake, or in NH(4)(+), which diminishes the proton gradient for ACh uptake into the vesicles. Neither treatment changed the size of the synaptic vesicles. ACh content and vesicle size were both decreased in preparations stimulated in (-)-vesamicol, an inhibitor of ACh uptake in vesicles. Since the other inhibitors decreased ACh content by a similar amount without altering vesicle size, (-)-vesamicol may decrease vesicle size by acting on another target. We also found that a hypertonic solution in which the anion was aspartate increased quantal size similar to gluconate. Both anions have high hydration energy and a large volume. When these treatments increased quantal size the mean 20-80 % rise time of MEPPs recorded with an extracellular electrode was 170 micros. In the controls it was 97 micros. Perhaps some of the added ACh is bound within the vesicles, which slows the rise. Our major conclusion is that ACh content can change notably without any change in the size of the synaptic vesicles.
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Recycling and refilling of transmitter quanta at the frog neuromuscular junction.
The Journal of Physiology, 2000Co-Authors: William Van Der Kloot, Roger Cameron, Cesare Colasante, Jordi MolgóAbstract:1. Fluorescent dyes have been used at the frog neuromuscular junction to label synaptic vesicular membrane. Retrieved membrane is reformed into vesicles, which are released along with pre-existing vesicles. Consequently, if vesicular refilling with acetylcholine (ACh) is depressed by inhibitors, two sizes of quanta should be released: normal and smaller. As recycling continues the fraction of smaller size quanta should increase exponentially. 2. We enhanced the rate of quantal release by elevating the K+ concentration. The principal inhibitors were (-)-vesamicol (VES), hemicholinium-3 (HC3), and NH4+. Quantal size measurements were fitted to one and to two cumulative lognormal probability distribution functions. When two fitted better, the statistical significance assessment took into account the three additional parameters used in calculating the fit. 3. After recycling in the presence of inhibitor, many sets were fitted better by two lognormal functions. As recycling continued, the fraction of the Miniature Endplate Potential voltage-time integrals ( MEPPs) in the larger sub-population decreased exponentially. 4. The size of the releasable pool was estimated by counting the quanta released by carbonyl cyanide m-chlorophenylhydrazone (CCCP). This was compared to pool sizes calculated from the inhibitor experiments. The two estimates of pool size were indistinguishable, with mean values ranging from about 170,000 to 270,000. 5. With all of the treatments tested, the means of the sizes in the smaller sub-population of MEPPs were about 1/3 those of the larger sub-populations. 6. Recycling synaptic vesicles appear to be incorporated into the releasable pool from which they have roughly the same probability of release as the pre-existing vesicles.
Shoei-yn Lin-shiau - One of the best experts on this subject based on the ideXlab platform.
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Potentiation of Miniature Endplate Potential frequency by ATP in Xenopus tadpoles
British journal of pharmacology, 1993Co-Authors: Shiau‐hui Yang, Shoei-yn Lin-shiauAbstract:1. Extracellular application of ATP (1 mM), a substance co-stored and co-released with acetylcholine in peripheral nervous systems, potentiated the spontaneous secretion of acetylcholine (ACh) but had no effect on the amplitude and decay time constant of Miniature Endplate Potentials (m.e.p.ps) at neuromuscular synapses in Xenopus tadpoles. 2. alpha,beta-Methylene ATP (0.3 mM) and GTP (1 mM) were also effective in increasing m.e.p.p. frequency. On the other hand, ADP, AMP and adenosine (all at 1 mM) decreased m.e.p.p. frequency. 3. Unlike the transient effect of ATP analogue and GTP on m.e.p.p. frequency, the phorbol ester TPA (2 microM) which is a protein kinase C activator, increased m.e.p.p. frequency consistently and the effects lasted as long as the presence of TPA. 4. Staurosporine (0.5 microM) and H-7 (10 microM), which are protein kinase C inhibitors, each decreased the basal level of m.e.p.p. frequency and markedly inhibited the effects of both ATP and TPA. 5. These results suggest that there is a basal activity of cytosolic protein kinases in the nerve terminals of Xenopus tadpoles and the effect of ATP is probably mediated by the binding of membrane surface purinoceptors which in turn activates cytosolic protein kinases and increases ACh release.
William Van Der Kloot - One of the best experts on this subject based on the ideXlab platform.
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Vesicle size and transmitter release at the frog neuromuscular junction when quantal acetylcholine content is increased or decreased.
The Journal of Physiology, 2002Co-Authors: William Van Der Kloot, Jordi Molgó, Roger Cameron, Cesare ColasanteAbstract:We investigated whether the synaptic vesicles at the neuromuscular junction change size when their acetylcholine (ACh) content is altered. The size of the Miniature Endplate Potential (MEPP) increased 3- or 4-fold in preparations pre-treated in a hypertonic solution in which the anion was gluconate. We measured the dimensions of synaptic vesicles in such preparations and in controls. The size of the vesicles and size distribution were indistinguishable. Quanta contained about half of the usual amount of ACh in preparations stimulated in the presence of hemicholinium-3, an inhibitor of choline uptake, or in NH(4)(+), which diminishes the proton gradient for ACh uptake into the vesicles. Neither treatment changed the size of the synaptic vesicles. ACh content and vesicle size were both decreased in preparations stimulated in (-)-vesamicol, an inhibitor of ACh uptake in vesicles. Since the other inhibitors decreased ACh content by a similar amount without altering vesicle size, (-)-vesamicol may decrease vesicle size by acting on another target. We also found that a hypertonic solution in which the anion was aspartate increased quantal size similar to gluconate. Both anions have high hydration energy and a large volume. When these treatments increased quantal size the mean 20-80 % rise time of MEPPs recorded with an extracellular electrode was 170 micros. In the controls it was 97 micros. Perhaps some of the added ACh is bound within the vesicles, which slows the rise. Our major conclusion is that ACh content can change notably without any change in the size of the synaptic vesicles.
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Recycling and refilling of transmitter quanta at the frog neuromuscular junction.
The Journal of Physiology, 2000Co-Authors: William Van Der Kloot, Roger Cameron, Cesare Colasante, Jordi MolgóAbstract:1. Fluorescent dyes have been used at the frog neuromuscular junction to label synaptic vesicular membrane. Retrieved membrane is reformed into vesicles, which are released along with pre-existing vesicles. Consequently, if vesicular refilling with acetylcholine (ACh) is depressed by inhibitors, two sizes of quanta should be released: normal and smaller. As recycling continues the fraction of smaller size quanta should increase exponentially. 2. We enhanced the rate of quantal release by elevating the K+ concentration. The principal inhibitors were (-)-vesamicol (VES), hemicholinium-3 (HC3), and NH4+. Quantal size measurements were fitted to one and to two cumulative lognormal probability distribution functions. When two fitted better, the statistical significance assessment took into account the three additional parameters used in calculating the fit. 3. After recycling in the presence of inhibitor, many sets were fitted better by two lognormal functions. As recycling continued, the fraction of the Miniature Endplate Potential voltage-time integrals ( MEPPs) in the larger sub-population decreased exponentially. 4. The size of the releasable pool was estimated by counting the quanta released by carbonyl cyanide m-chlorophenylhydrazone (CCCP). This was compared to pool sizes calculated from the inhibitor experiments. The two estimates of pool size were indistinguishable, with mean values ranging from about 170,000 to 270,000. 5. With all of the treatments tested, the means of the sizes in the smaller sub-population of MEPPs were about 1/3 those of the larger sub-populations. 6. Recycling synaptic vesicles appear to be incorporated into the releasable pool from which they have roughly the same probability of release as the pre-existing vesicles.