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J Marsal

Publications and source records attributed to J Marsal.

At least 73 records · Page 4Linked to original sources
PubMed1990

Type A botulinum toxin disorganizes quantal acetylcholine release and inhibits energy metabolism.

The physiological, morphological and biochemical effects of type A Botulinum toxin (BoTX) were analysed in the electric organ of Torpedo, a modified neuromuscular system. The quantal content of the postsynaptic potential, or electroplaque potential (EPP), was reduced by BoTX but the quantum size remained unchanged till complete failure of the neurally evoked transmission. BoTX also suppressed the occurrence of spontaneous electroplaque potentials (MEPPs) of a quantal size but potentials of a smaller amplitude still kept on occurring in the intoxicated synapses. BoTX inhibited the evoked release of acetylcholine (ACh; biochemically measured) but the rate of spontaneous ACh release transiently increased during the period when evoked release went down. On the other hand, there were no significant change of ACh content, of ACh turnover, of ACh repartition in the vesicular and free compartments, or in the number of synaptic vesicles. Surprisingly, the amount of ATP was reduced to 50% in BoTX treated tissue at the time of transmission failure; also the level of creatine phosphate (CrP) was lowered to less than 20% and the rate of activity of creatine kinase was reduced. It was concluded that, electrophysiologically, BoTX affects synaptic transmission in a very similar way in the electric organ and in the neuromuscular junctions. On the other hand, the shortage of ATP supply found in the present study may play a role in the pathophysiology of intoxication and should be taken into account in investigations designed to see whether BoTX affects various phosphorylations in cholinergic nerve terminals.

Acetylcholine↗
PubMed1990

Location of phosphoglycerate mutase in rat skeletal muscle. An immunocytochemical and biochemical study.

The subcellular distribution of phosphoglycerate mutase was studied by immunogold techniques. With the aid of highly affinity-purified anti-phosphoglycerate mutase antibodies, the enzyme was found in both cytosol and nucleus of rat skeletal muscle. No evidence of interaction with contractile proteins was observed in cytosol. Nuclear location was also confirmed biochemically using purified nuclear preparations from rat skeletal muscle. Only one immunoreactive nuclear band was observed by Western blot experiments and corresponded to that of phosphoglycerate mutase mobility. Activity measurements from nuclear extracts showed that 25% of total specific activity is found in the nuclei.

Animals↗
PubMed1989

GABAergic modulation of acetylcholine release in cholinergic synaptosomes from Torpedo marmorata electric organ.

GABAA- and GABAB-receptor-specific agonists inhibit the depolarization-evoked release of acetylcholine in cholinergic synaptosomes from Torpedo electric organ. Over 60% of the release is inhibited by a 10(-4) M concentration of GABA itself. IC50s for muscimol and baclofen are 1.3 x 10(-4) and 2.2 x 10(-6) M, respectively. The effect of muscimol is totally blocked by the direct antagonist bicuculline methiodide, and also by the allosteric antagonists methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate, picrotoxinin and tert-butylbicyclo-orthobenzoate; the effect of baclofen is blocked by delta-aminovalerate. Furthermore, the inhibitory action of muscimol on acetylcholine release is substantially enhanced by flunitrazepam and pentobarbital. These results suggest the existence of typical GABAA and GABAB receptors in the presynaptic nerve terminals of the Torpedo electric organ regulating the liberation of acetylcholine and therefore the discharge of the electroplaques.

Acetylcholine↗
PubMed1989

Increase in reactive cholesterol in the presynaptic membrane of depolarized Torpedo synaptosomes: blockade by botulinum toxin type A.

We have investigated the redistribution of filipin-cholesterol complexes at freeze-fractured presynaptic membrane of pure cholinergic synaptosomes isolated from Torpedo electric organ during acetylcholine release. After chemical depolarization, filipin-induced lesions increase at the presynaptic membrane. These changes do not take place when synaptosomes are stimulated in a calcium-free medium. Botulinum neurotoxin type A blocks both acetylcholine release and the rearrangement of filipin-induced lesions induced by depolarization. Since botulinum neurotoxin type A does not block either membrane depolarization or calcium entry into the nerve terminal, our results suggest that the redistribution of filipin-cholesterol complexes is linked to the acetylcholine release process.

Acetylcholine↗
PubMed1989

Botulinum toxin type A blocks the morphological changes induced by chemical stimulation on the presynaptic membrane of Torpedo synaptosomes.

The action of botulinum neurotoxin on acetylcholine release, and on the structural changes at the presynaptic membrane associated with the transmitter release, was studied by using a subcellular fraction of cholinergic nerve terminals (synaptosomes) isolated from the Torpedo electric organ. Acetylcholine and ATP release were continuously monitored by chemiluminescent methods. To catch the membrane morphological changes, the quick-freezing method was applied. Our results show that botulinum neurotoxin inhibits the release of acetylcholine from these isolated nerve terminals in a dose-dependent manner, whereas ATP release is not affected. The maximal inhibition (70%) is achieved at neurotoxin concentrations as low as 125 pM with an incubation time of 6 min. This effect is not linked to an alteration of the integrity of the synaptosomes since, after poisoning by botulinum neurotoxin type A, they show a nonmodified occluded lactate dehydrogenase activity. Moreover, membrane potential is not altered by the toxin with respect to the control, either in resting condition or after potassium depolarization. In addition to acetylcholine release inhibition, botulinum neurotoxin blocks the rearrangement of the presynaptic intramembrane particles induced by potassium stimulation. The action of botulinum neurotoxin suggests that the intramembrane particle rearrangement is related to the acetylcholine secretion induced by potassium stimulation in synaptosomes isolated from the electric organ of Torpedo marmorata.

Acetylcholine↗
PubMed1988

Depolarization-stimulated protein phosphorylation in pure cholinergic nerve endings.

Cholinergic synaptosomes obtained from the electric organ of Torpedo marmorata have been used to study chemical stimulation-stimulated protein phosphorylation. Cholinergic synaptosomes were exposed to elevated K+0 concentrations or other chemical depolarizing agents such as gramicidin or secretagogues as the calcium ionophore A23187. During depolarization several synaptosomal proteins increase their state of phosphorylation. This phenomenon depends on the presence of Ca2+ in the external medium. These results suggest that stimulation of protein phosphorylation may be implicated in the acetylcholine release process and could represent a modulation mechanism in the neurotransmitter release machinery at this cholinergic synapse.

Animals↗
PubMed1988

Ouabain induces acetylcholine release from pure cholinergic synaptosomes independently of extracellular calcium concentration.

We have studied the correlation between [3H]ouabain binding sites, (Na+ + K+)ATPase (EC 3.6.1.3) activity and acetylcholine (ACh) release in different subcellular fractions of Torpedo marmorata electric organ (homogenate, synaptosomes, presynaptic plasma membranes). Presynaptic plasma membranes contained the greater number of [3H]ouabain binding sites, in good agreement with the high (Na+ + K+)ATPase activity found in this fraction. Blockade of this enzymatic activity by ouabain dose-dependently induced ACh release from pure cholinergic synaptosomes, either in the presence or absence of extracellular calcium ions. We suggest that one of the mechanisms involved in the ouabain-induced ACh release in the absence of Ca2+o may be an increase in Na+i that could (a) evoke Ca2+ release from internal stores and (b) inhibit ATP-dependent Ca2+ uptake by synaptic vesicles.

Acetylcholine↗
PubMed1988

Tetanus and botulinum toxins block the release of acetylcholine from slices of rat striatum and from the isolated electric organ of Torpedo at different concentrations.

Tetanus toxin, like botulinum toxin type A, blocks cholinergic synaptic transmission at the central and peripheral nervous systems. Nevertheless, the diseases induced by the two toxins are different since tetanus toxin induces a spastic paralysis and botulinum toxin elicits a flaccid paralysis. Thus, we have investigated the sensitivity of a central and a peripheral cholinergic synapse to these two toxins. We have studied the action of both poison on the release of acetylcholine from slices of the rat striatum and from the isolated electric organ of Torpedo, which is homologous to the neuromuscular junction. Acetylcholine release from the rat striatum was continuously monitored by a chemiluminescent method. The secretion of acetylcholine from the electric organ was estimated both by measuring the amplitude of the evoked electrical discharge from stacks of electroplaques, and by continuously monitoring the neurotransmitter release from isolated nerve terminals. Tetanus toxin blocks the electrical discharge of electric organ prisms, and also impairs the release of acetylcholine from the Torpedo electric organ nerve endings. Our results on acetylcholine release show that tetanus toxin is more potent than botulinum toxin type A at the central cholinergic synapse (tetanus/botulinum toxins potency ratio about 100-200) whereas botulinum toxin is the most potent at the peripheral cholinergic synapse (botulinum/tetanus toxins potency ratio about 100).

Acetylcholine↗
PubMed1988

Energy metabolism and quantal acetylcholine release: effects of botulinum toxin, 1-fluoro-2,4-dinitrobenzene, and diamide in the Torpedo electric organ.

In the Torpedo electric organ, a modified nerve-muscle system, type A botulinum toxin blocked the release of acetylcholine (ACh) quanta, both neurally evoked and spontaneous. At the same time, the toxin increased the release of a class of small miniature potentials (the subminiature potentials), reduced the ATP and more the creatine phosphate content of the tissue, and impaired the activity of creatine kinase (CK). Thus, we compared this pattern of changes with those provoked by 1-fluoro-2,4-dinitrobenzene (FDNB), an efficient inhibitor of CK. As expected, FDNB rapidly inactivated CK, which resulted in a profound depletion of ATP whereas the stores of creatine phosphate were preserved. In addition, FDNB caused conspicuous morphological alterations of nerve endings and ACh depletion. This agent also suppressed evoked and spontaneous quantal release whereas the occurrence of subminature potentials was markedly increased. Diamide, a penetrating thiol oxidizing substance, provoked first a transient rise in quantal ACh release and then blockade of transmission with, again, production of a large number of subminiature potentials. Creatine phosphate was depleted in the tissue by diamide, the ATP content reduced, and CK activity partly inhibited. The morphology of nerve terminals did not show obvious changes with either diamide or botulinum toxin at the stage of transmission failure. Although the three poisons acted by different mechanisms, this resulted in a rather similar pattern of physiological changes: failure of quantal release and enhancement of subquantal release. These results and experiments on synaptosomes indicated that CK inhibition was probably a crucial mechanism for FDNB but not for diamide or botulinum intoxication.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1987

Botulinum neurotoxin inhibits depolarization-stimulated protein phosphorylation in pure cholinergic synaptosomes.

Botulinum neurotoxin, a strong blocker of acetylcholine release at peripheral cholinergic synapses, inhibits depolarization-stimulated protein phosphorylation in pure cholinergic synaptosomes isolated from the electric organ of Torpedo marmorata. Moreover, tetrodotoxin has the same effect on protein phosphorylation when cholinergic synaptosomes are depolarized by veratridine. Correlation between presynaptic protein phosphorylation and acetylcholine release is suggested by the fact that botulinum neurotoxin blocks specifically neurotransmitter release without affecting membrane depolarization and calcium fluxes in our synaptosomal preparation.

Animals↗
PubMed1987

ATP release from pure cholinergic synaptosomes is not blocked by tetanus toxin.

Tetanus toxin (TeTx) is a neurotransmission impairing toxin that acts on several neurotransmitter systems. TeTx also inhibits the K+-induced release of acetylcholine (ACh) from synaptosomes isolated from the electric organ of Torpedo. Neither the membrane potential and depolarization, nor the depolarization-induced calcium uptake into cholinergic nerve terminals is modified after TeTx poisoning. On the other hand, it is known that, when cholinergic nerve terminals are stimulated, there is a release of ATP associated with the release of ACh. We have explored the action of TeTx on this co-release, and have found that there is no action of TeTx on the nucleotide release. Thus, TeTx blocks ACh release without modifying ATP release.

Acetylcholine↗
PubMed1987

Structural changes at pure cholinergic synaptosomes during the transmitter release induced by A-23187 in Torpedo marmorata. A freeze-fracture study.

Pure cholinergic synaptosomes isolated from the electric organ of Torpedo marmorata were stimulated by calcium ionophore A-23187. The effect of time course of stimulation on the changes in intramembrane particles (IMPs) on presynaptic membranes was studied by quick-freezing and aldehyde-fixation freeze-fracture. We showed that the decrease of small-particle density at the P-face and the increase of large-particle density at the E-face was maximum after 30 sec of A-23187 stimulation. Later, the density of synaptic vesicles decreased. We suggest that the redistribution of IMPs on the presynaptic membrane and acetylcholine (ACh) release from pure cholinergic synaptosomes have a similar time course when triggered by A-23187.

Acetylcholine↗
PubMed1987

Botulinum toxin inhibits quantal acetylcholine release and energy metabolism in the Torpedo electric organ.

1. Type A Botulinum toxin (BoTX) blocked nerve-electroplaque transmission in small fragments of Torpedo marmorata electric organ incubated in vitro. The effect was observed either with the crystalline toxin complex (associated with haemagglutinin) or with the purified neurotoxin (molecular weight approximately 150,000). 2. The quantal content of the evoked post-synaptic response was reduced by BoTX but the quantum size remained unchanged till complete blockade of the evoked response. 3. Spontaneous electroplaque potentials were composed of two populations: one with a bell-shaped amplitude distribution (miniature potentials or quanta) and a population of small events with a skewed distribution (subminiatures). In BoTX-poisoned tissue, the bell-distributed miniatures progressively disappeared, but the subminiatures kept on occurring. Occasionally, larger spontaneous potentials with a slow time course were recorded; they were also BoTX resistant. 4. A biochemical assay showed that evoked acetylcholine (ACh) release was impaired by BoTX. During the period when evoked transmission was blocked, spontaneous ACh release transiently increased. 5. At the time of transmission blockade, there was no significant change of ACh content, of ACh turnover, of ACh repartition in the vesicle-bound and free compartments, or of the number of synaptic vesicles. 6. The amount of ATP was reduced to 50% by BoTX, and that of creatine phosphate (CrP) to less than 20%. The ATP-CrP-converting enzyme, creatine kinase, was inhibited in BoTX-poisoned tissue. 7. Thus, the electrophysiological effects of BoTX are very similar at the nerve-electroplaque and the neuromuscular junctions. The present work suggests in addition that suppression of quantal release by BoTX is related to marked alterations of the energy metabolism in the tissue.

Acetylcholine↗
PubMed1985

Modulation of the endogenous acetylcholine release from rat striatal slices.

The dopaminergic modulation of the acetylcholine release from rat striatal slices has been investigated using a chemiluminescent method. Dopamine, more efficiently than apomorphine, decreased the potassium-evoked release of acetylcholine. The effect of dopamine antagonists, haloperidol and sulpiride, has been studied, and haloperidol was a better antagonist than sulpiride to the dopamine effect. Haloperidol elicited an acetylcholine release from striatal slices at 0.1 nM, probably by removing endogenous dopamine from dopaminergic receptors.

Acetylcholine↗
PubMed1985

Large-scale purification of presynaptic plasma membranes from Torpedo marmorata electric organ.

The presynaptic plasma membrane (PSPM) of cholinergic nerve terminals was purified from Torpedo electric organ using a large-scale procedure. Up to 500 g of frozen electric organ were fractioned in a single run, leading to the isolation of greater than 100 mg of PSPM proteins. The purity of the fraction is similar to that of the synaptosomal plasma membrane obtained after subfractionation of Torpedo synaptosomes as judged by its membrane-bound acetylcholinesterase activity, the number of Glycera convoluta neurotoxin binding sites, and the binding of two monoclonal antibodies directed against PSPM. The specificity of these antibodies for the PSPM is demonstrated by immunofluorescence microscopy.

Acetylcholinesterase↗
PubMed1982

Binding of beta-bungarotoxin to Torpedo electric organ synaptosomes. A high resolution autoradiographic study.

Isolated pure cholinergic synaptosomes from Torpedo electric organ were incubated in vitro with beta-bungarotoxin for 15, 30 and 60 min and processed for electron microscopy. It was found that no morphological damage was seen after 15 min but by contrast, severe disruption of synaptosomes was present at 30 or 60 min after incubation with toxin. Synaptosomes were incubated also for 15 min in the presence of 125I-labelled beta-bungarotoxin and the binding was evaluated by electron microscopic autoradiography. The toxin was found to bind to the presynaptic membrane. The surface density of toxin binding sites was calculated to be around 3000/micron2. In a minor population of synaptosomes, the toxin was translocated into large vesicles suggesting that the toxin-receptor complexes underwent endocytosis in such vesicles. These results give further support to the view that inhibition of transmitter release by the toxin is produced by its action on plasma membrane.

Animals↗
PubMed1981

Effects of ouabain and electrical stimulation on the fine structure of nerve endings in the electric organ of Torpedo marmorata.

The cycle of synaptic vesicles was studied in isolated nerve terminals and in the electric tissue of Torpedo marmorata. The synaptosomes, as used in this investigation, were a pure cholinergic subcellar fraction that captured dextran particles as an extracellular marker. This endocytotic phenomenon was enhanced by potassium depolarization. Field electrical stimulation (1 Hz and 10 Hz) of the electric organ induced the appearance of membrane foldings into presynaptic terminals. Morphometric studies showed that the number of synaptic vesicles did not decline until after at least 30 min. On the other hand, at 10 Hz these changes were accompanied by an increase in length of the membrane of the terminal. At 15 min of recovery after prolonged stimulation, there was a great increase in density of synaptic vesicles with a large number of vesicles of small diameter. This increase was accompanied by a decrease of membrane length, suggesting that reformation of vesicles is related to retrieval of membrane. Pharmacological stimulation with ouabain produced changes similar to those of long-term electrical stimulation. These changes in membrane were accompanied by a decrease of the population of synaptic vesicles and a wide variation in their diameters. It is concluded that structural changes reported here could not be correlated with kinetics of the transmitter release.

Animals↗